We need to learn from our prior mistakes. The example of the 2016 election is just that an example. There is misinformation everywhere. As stated this is not new information to us.

Wow after reading the article linked here, I think Google needs to make some changes to their news algorithm. It's crazy to think that anyone can just create a news site, publish false and possibly dangerous information, and have a site like Google promote it as true. Of course I'm guessing they've already made some fixes, but this hoax tricked a lot of people.

Жить умеренно

It's so infuriating that many Christians justified this

This seems like man compares and tries to dominate himself over others like a competition.

It would be nice if people would stop saying this—and saying it like this—as if it's a badge of honor. jQuery is a fuckin' beast. 10 years ago, the reason that the browser was being brought to a crawl on any given pages often came down to the fact that it was using jQuery. jQuery is the reason that bloated frameworks became normalized and brought us to where we are today. So, enough already with this just-a-little-jQuery stuff.

It seems the goal of most schools is to just teach students the subject they are required to know and not life skills.

cool research tools

A little Ozymandias esque? Not sure if it's meant to be a warning or just a little melancholic

I like this especially because it's a way for a trustworthy business to get some use from seeming trustworthy and not get-rich-quick. I don't want to have to guess whether some awful VC startup is going to last more than a couple years -- but when I have more of a relationship with a smaller entity, I'm more comfortable with that.

Facades could just be a form of integrity. We have to look good at work because we have to look trust worthy and professional. It's just the modern standard, not really a lie. There are still facades that are lies, but most I would say are not lies.

It's easier to say things like this when your name is "anonymous" and you have no identity. This is also just disgusting and embarrassing for someone to say. Though true, the Internet allows for comments like this to happen. Is it the Internet's fault? No. Maybe? I don't know.

it's almost like she's lying to herself and playing make-believe with her life

I feel like it's too kind to the phenomenon to call it "over analytical" or "smart".

Consider how often you've seen a study headline (or pop science piece) with comments below it saying that the stated effect could be explained by XYZ factor that the study authors aren't considering. For a while I had a practice of going and looking at the study when I saw people leaving those comments. A large portion of the time I found the commenter's iamverysmart explanation was explicitly controlled for in the analysis design, and an even larger amount of the time it was at least addressed in the text. I spent a while copy-pasting these things into replies to the (lauded!) commenters, but it was like moving a lake with a spoon. Eventually I stopped checking and just started assuming that substantive counter-analysis to one of these studies would at least be posted independently, not in a comment section. That heuristic's been working pretty okay.

It is in no way "over analytical" to pop off with "well here's my rationalization of this result I don't like the sound of" without reading the source, and I wonder if it's too much US classroom culture making people think their zero-effort "insight" is valid and worthy.

It happens way to often that some people may even say common. Most men think that declining to give someone your number is no big deal, but some men's pride can take turn for the worst.

Kevin Courtney #NEU💝NHS. (2022, January 5). Ventilation isn’t just for Covid.... ...It’s for Education This study looks at the impact of CO2 not just as a marker of pollution but as a pollutant in itself. It shows that as CO2 rises above 700/800 ppm cognitive function begins to be impaired https://dash.harvard.edu/bitstream/handle/1/27662232/4892924.pdf?sequence=1&fbclid=IwAR2kWIHIJfssa_sw72MD6W1hnkDvSm4bikK5FOLxwQxhjYLEYjfPCfzXz3E [Tweet]. @cyclingkev. https://twitter.com/cyclingkev/status/1478778857536860170

I agree that we should not regard the conventional understanding as true—in the sense of treating it as always true. But I disagree with the suggestion of this piece, which is that it is never true.

We shouldn't conclude too much from the existence of "broken" versions that were discarded. Much of the manufacture of clarity that comes out of writing is really just a consequence of putting it down in a way that is accessible to other people (probably, as PG says here, as a result of multiple revisions), but it's not necessary for firsthand understanding. Suppose I write some code, and it is correct. Later, I revise it to be clearer to a wandering party. This takes work, but have I accomplished anything with respect to what PG is saying here? Did I, when originally setting out to put it into a fixed form, find that my understanding was lacking and have to confront this and then only after reconciling the issue truly understand it—whereas I didn't before? No. It was correct before, and it's not more correct now; it's possible that I understood it perfectly well, I just didn't understand how to better explain it to others.

Note: This rebuttal was posted by the corresponding author to Review Commons. Content has not been altered except for formatting.

Learn more at Review Commons

Reply to the reviewers

Note: Figures we made to respond to the referee comments appear to be not supported by the ReviewCommons system.

Reviewer #1 (Evidence, reproducibility and clarity (Required)):

**Summary:**

The authors characterized a new lncRNA locus named FLAIL that controls flowering time in Arabidopsis thaliana. The functional validation of this locus is strongly supported by the use of several different tools (CRISPR-Cas9 deletions, T-DNA insertion, amiRNA gene silencing, and transgene complementation of KO lines). It is also suggested that FLAIL lncRNA works in trans but not in cis. There are strong observations supporting that FLAIL works in trans.

Moreover, it is suggested that FLAIL regulates gene expression by interacting with distant chromatin loci. This was assessed using RNA-Seq and ChIRP-Seq. Yet, the overlap between DEGs in the flail mutant and FLAIL binding sites at the chromatin is very small, with only 12 genes. From those, only 2 flowering genes' expression was rescued by FLAIL transgene complementation. The final conclusion that FLAIL lncRNA represses flowering by direct inhibition of the 2 flowering genes expression is correlative, and lacks genetic validation.

#1.1 We plan to support the conclusions in the manuscript genetically as the reviewer suggests. We started these experiments yet they will require the timeframe of the full revision.

In addition inspection of the supplementary file shows that the ChIRP analysis was done without filtering for the FDR so that some of the positive hits have an FDR of 0,232.

#1.2 We strengthened the manuscript by implementing and FDR filter of ChIRP-seq results. The distribution of FLAIL binding sites in Fig. S7B and Table S4, and overlapping numbers between DEGs and FLAIL-ChIRP in Fig. S8A were correspondingly updated.

In addition, many of the peaks land in intergenic regions with is not mentioned in the text a graph with the position of the peaks in respect to nearby genes would help.

#1.3 Thank you for the suggestion, we strengthened the manuscript with the requested analysis. We implemented the FDR filter, then we used "tssRegion" in ChIPseeker to set distance to the nearest TSS as (-1000, 1000), then most peaks were located in promoter regions (67.24%) and in intergenic regions with 16.38%. Since many papers present the position of the peaks by ChIPseeker (PMID: 32338596, PMID: 28221134, PMID: 31081251, PMID: 32012197, PMID: 31649032, PMID: 32633672) we also applied a similar method to display a distribution of FLAIL binding loci relative to distance from the nearest TSS in Fig. S7C.

In one sentence, the authors used the right model system and methodology, including advanced techniques, to characterize a new trans-acting lncRNA important for controlling the flowering time in Arabidopsis but lack evidence supporting a mechanism of action that goes beyond the interaction with several chromatin loci.

**minor points:**

line#63-64 the authors say the COLDAIR and ASL work on FLC in cis in my view the original papers suggested/showed they work in trans.

#1.4 We increased precision by changing this sentence to ‘Vernalization-induced flowering associates with several lncRNAs such as ____COOLAIR____, COLDAIR____, ANTISENSE LONG (ASL), and COLDWRAP____ that in cis or in trans locally repress gene expression of FLOWERING LOCUS C (FLC), a key flowering repressor at different stages of vernalization’____.

Fig 1B please add some more protein-coding RNAs for the bio-info analysis for comparison

#1.5 ____done.

Order of Supplementary Fig citation is mixed with S2 coming before S1B

#1.6 Thank you, we ordered all figures by appearance in the text. __

__

It would help the reader to have a schematic of the crisper deletions, T-DNA insertion, and position of primers used for the RT-qPCR.

#1.7 We enhanced our presentation of Fig. 1A. It shows a schematic of them as well as positions of primers.

In the supplementary PDF file, some of the text is missing on page 3 beginning and end of lines.

#1.8 we ensured all text in new submission.

Reviewer #1 (Significance (Required)):

The use of several different tools to validate the biological function of FLAIL locus is a major strength of this work.

The authors propose that flowering time and its gene regulation are controlled by sense FLAIL lncRNAs. However, the sense transcription of FLAIL locus is not detected in wild-type plants by TSS-Seq, TIF-Seq, or plaNET-Seq.

#1.9.1 There appears to be some confusions. Transcription of sense FLAIL can be observed in chr-DRS, TSS-seq, TIF-seq in wild type and even in plaNET-seq in NRPB2-FLAG nrpb2-1 plant. We enhanced presentation of Fig. 1 and provided a more clear description in Line 81-99.

If the authors would have explored further the expression of FLAIL transcripts in different stages of development (vegetative and non-vegetative) and in response to different conditions, it would make their claims on the function of FLAIL lncRNAs more convincing. Additionally, flail mutants could have been obtained in the hen-2 background, since it's there where we can observe FLAIL transcription.

#1.9.2 Thank you for the suggestion. We included additional analyses in ____Fig. S2 for FLAIL transcription level in different tissues and different abiotic stress conditions base on 20,000 publicly available RNA-seq libraries (PMID: 32768600). Although many libraries are non-stranded, this analysis determined that sense FLAIL or total FLAIL (including sense and antisense) is broadly expressed over many tissues and induced in response to many abiotic stresses (Fig. S2A-B), therefore suggesting that FLAIL may be needed broadly in Arabidopsis.

FLAIL locus lays on the proximal promoter region of PORCUPINE (PCP), an important regulator of plant development. As flail mutants, pcp mutants display an early flowering phenotype. The authors show no link between FLAIL and PCP from the overlap between re-analysis of published RNA-Seq data for pcp and RNA-Seq and ChIRP-Seq from the authors. This analysis is not enough to exclude the involvement of PCP from the FLAIL function. PCP expression using RT-qPCR should be performed in flail mutants to further support that FLAIL works independently from PCP.

#1.10 We strengthened this conclusion by adding the requested experiment. PCP transcription level in flail3 mutant was provided by RT-qPCR and RNA-seq in Fig. S11A-B.

This work does not hypothesize any molecular mechanism besides the interaction of FLAIL lncRNAs with several chromatin loci. It was recently proposed in Arabidopsis that a trans-acting lncRNA interacts with distant loci via the formation of R-loops. The authors do not comment on that. This work would benefit in correlating FLAIL binding sites with R-loop-forming regions mapped in Arabidopsis, regardless of the results from this analysis. Additionally, the authors could attempt to look for a motif responsible for FLAIL binding.

Check R-loop forming data R-loops (Santos-Pereira and Aguilera, 2015) in Arabidopsis, determined by DRIP-seq (Xu et al., 2017).

#1.11 Thanks very much for this excellent suggestions.

First, we searched for a consensus DNA motif on FLAIL binding regions by Homer. We determined four commonly enriched DNA sequence motifs among FLAIL target genes (Fig. 4G). Notably, the target genes CIR1 and LAC8 contained consensus sequences that matched to all FLAIL binding motifs (Fig. 4G). These data are consistent with a model where FLAIL binds DNA targets through a sequence complementary mechanism. Functionally important sequences are frequently conserved among evolutionarily distant species, we observed three motifs that appeared to cross-species conserved (Fig. S9), suggesting a potential evolutionarily constrained role.

Second, we indeed identified R-loops peaks on several of FLAIL binding sites by DRIP-seq (Xu et al., 2017). For example, we observed R-loop formation over three FLAIL binding motifs at CIR1 locus and one at LAC8 (Fig. R1), indicating that R-loop formation may also be a factor determining FLAIL binding. Even though R-loop peaks are present at several FLAIL targets, full elucidation if R-loop formation determines FLAIL targeting requires further experimental evidence is beyond the scope of the current manuscript.

Fig. R1 Representative tracks at LAC8 and CIR1 showing R-loop formation by DRIP-seq on Watson strand (w-R loops), Crick strand (c-R loops). Undetectable R-loops after RNAse-H treatment was shown as negative control. Four conserved sequence regions of FLAIL binding motifs were indicated by red arrows at LAC8 and CIR1 loci. Gene annotation was shown at the bottom.

Most of the key conclusions are convincing, except for the flowering time control directly through CIR1 and LAC8, which should be mentioned as speculative

____#1.12____ Thank you for finding most key conclusions convincing. We plan strengthen the manuscript with additional genetic evidence to as part of the full revision.

The words locus and loci are latin and they should be written in italic. The word Brassicaceae, referring to the family should be in italic, and should not be "Brassicaceaes". The word analysis has the wrong spelling.

#1.13 We follow conventions given in Scientific Style and Format: The CBE Manual for Authors, Editors and Publishers (1994) Cambridge University Press, Cambridge, UK, 6th edn. The words locus and loci are common Latin terms and should not be italicized. However, should the format of the final prefer these words in italics we will change it later. We improved consistency of using italics. “Brassicaceaes” was changed to “Brassicaceae”.

"How much time do you estimate the authors will need to complete the suggested revisions: this is difficult to answer as it depends to which level the author would like to take their work. In my view, if all new experiments would have to be started from scratch it is too far away to be estimated.

Reviewer #2 (Evidence, reproducibility and clarity (Required)):

In this ms, the authors identified the FLAIL lncRNA that represses flowering in Arabidopsis from a locus producing sense and antisense transcripts. They use an allelic series involving T-DNA insertions, CRISPR/Cas9 and artificial miRNAs to study the role of FLAIL in flowering. A complementation series of constructs of the flail3 allele allowed them to show that the sense FLAIL lncRNA can act in trans. RNAseq revealed a small group of genes linked to the regulation of flowering whose expression is affected in the mutant and restored in the complementation line. To gain further insight into FLAIL function, the authors used a ChIRPeq approach to test whether the lncRNA can recognize potential target genes along the genome and they could show that FLAIL binds specific genomic regions. Clearly, this paper shows very nice evidence that the FLAIL lncRNA can act in trans to regulate gene expression. Nevertheless, there are certain points that need to be clarified to further support the action of the sense FLAIL transcript.

1.According to Fig. 1 A, the antisense FLAIL is "internal" to the DNA genomic area spanning the sense FLAIL. Hence, with direct RT-qPCR is very difficult to distinguish between these molecules as a minor "RT" activity of the Taq polymerase may lead to detection of low levels of antisense, idem if RDRs may generate low antisense levels. Although I think that the plaNET seq brings strong evidence about the start and ends of these molecules, to measure them by RT-qPCR is not trivial and requires the use of strand-specific RT-PCR using a 5' extension of the oligo and amplification with one oligo of the FLAIL sequence (sense or antisense) and the added oligo.

#2.1.1 Thanks for this good suggestion. We tested both sense and antisense FLAIL transcription using oligo linked gene specific reverse primers for RT and a pair of the linked oligo and gene specific forward primer for qPCR. Primer locations were shown in Fig. 1A and new data were in Fig. 1C-D, Fig. 2B-C, and Fig. S4B-C.

It is not clear how they could distinguish precisely sense and antisense particularly when both RNAs correlate as it is the case here in all alleles (Fig. 1 C and 1D). This should be more explicitly mentioned in the materials and methods section.

#2.1.2 We gave a description of strand specific RT-qPCR method in detail in Line 397-402.

2.In Fig. 2, what are the levels of antisense in the complementing lines with the sense transcript? And reciprocally sense levels in antisense constructs?

#2.2 We added this data in Fig. 2B-C and described in Line 136-143. We indeed observed that sense FLAIL transcripts in the transformed asFLAIL construct or asFLAIL transcripts in the transformed sense FLAIL construct was similar to the control 35S:GUS (Fig. 2B-C), validating that NOS terminator inhibits antisense transcripts. We also noted that the transformed 35S:GUS and sense FLAIL construct expressed higher asFLAIL compared to the flail3 mutant (Fig. 2C). This may be caused by a T-DNA insertion of the resulting transgenic plants.

This will definitively demonstrate the assumption that the T-NOS termination will not allow any expression on the other strand. At present, only one of the lncRNAs is measured in each experiment?

#2.3 We appreciate the next-level reflection of this reviewer, with so many regions initiating cryptic antisense transcription it is an interesting challenge to identify a 3´- terminator that initiates no or poor antisense transcription.

First, previous published data argue that the NOS terminator is largely abolishing initiation of antisense transcription (PMID: 33985972, PMID: 30385760, PMID: 27856735). All these studies address roles of antisense transcription by generating mutations abolishing antisense lncRNA transcription using the NOS terminator sequences.

Second, to satisfy the curiosity of this reviewer, we provide data below that from another manuscript of the lab in preparation. It’s a screenshot of plaNET-seq in fas2-4 NRPB2-FLAG nrpb2-1 mutant carrying a pROK2 construct. The pROK2 T-DNA coincidentally carries a NOS terminator. We mapped plaNET-seq reads to the pROK2 scaffold to display the reads. In pROK2, a NOS promoter activates NPTII expression (red) with NOS terminator as a terminator sequence. No antisense transcription (blue) is detectable by this sensitive method to detect nascent transcripts. Taken together, the selection of the NOS terminator as a region suppressing initiation of antisense transcription represents a valid choice.

Fig. R2 Genome browser screenshot of plaNET-seq at NPTII locus of pROK2 T-DNA vector in fas2-4 NRBP2-FLAG nrpb2-1 mutant. This mutant carries a pROK2 construct, in which a NOS promoter activates NPTII expression with NOS terminator a terminator sequence. Sense strand was shown in red and antisense strand in blue. pROK2 annotation was shown at the bottom.

3.In Fig. 3, it will be important to also show the FLAIL locus in the flail3 mutants (in comparison to the wt) as well as the transgene locus. Here the reads will be strand specific and furthermore this will allow to show that the transgene is not generating antisense transcripts (through RDRs for gene silencing?) and confirm that the sense FLAIL is required for the complementation.

#2.4 Thank you very much for this suggestion. NGS reads for endogenous FLAIL and transgenic FLAIL both map to the FLAIL locus, so we show the FLAIL locus in Fig 3B. This representation shows that sense FLAIL transcripts were significantly reduced in flail3 and rescued in complementation line comparing to wild type. These data argue against the idea of gene silencing and linked antisense production from the transgene. However, RNA-seq suggests that an isoform of asFLAIL appears to accumulate in flail3. Since we fail to identify this accumulation by strand specific RT-qPCR result in flail3 and in CRISPR-deletion lines, this may be an asFLAIL isoform resulting from the T-DNA insertion.

4.In Fig. S5, the expression of FLAIL is shown in the artificial miRNA lines. Is the antisense FLAIL affected "indirectly" by the cleavage of the amiRNA or remains constant? This is likely the case but should be shown.

#2.5 We added this result in Fig. S4C and expression level of asFLAIL remains constant compared to the transformed empty vector control.

5.The ChIRPseq data adds major novelty to the ms and brings new ideas about the way of action of FLAIL. However, are there any common epigenetic states between ChIRP targets (e.g. histone modifications, antisense RNA production, homologies "detected" in the conserved regions between Camelina and Arabidopsis and the target loci? Or others) that may highlight potential mechanisms leading to repression mediated by FLAIL of these loci? There are many databases that could be explored (even during flowering) to search for potential relationships. Although precise description of the mechanism is out of the scope of this ms, this can be discussed in more detail to further expand on the nice data obtained.

#2.6 We searched for a consensus DNA motif on FLAIL binding regions by Homer. We determined four commonly enriched DNA sequence motifs in target genes. Notably, the target genes CIR1 and LAC8 contained consensus sequences that matched to all FLAIL binding motifs (Fig. 4G). These data are consistent with a model where FLAIL binds DNA targets through a sequence complementarity mechanism. Functionally important sequences are frequently conserved among evolutionarily distant species, we observed three motifs that appeared to cross-species conserved (Fig. S9), suggesting a potential evolutionarily constrained role.

**Minor comments:**

6.In Fig. S3, a global alignment between FLAIL and two loci in Arabidopsis and Camelina is sown. What is the extent of homology? How conserved is this sequence at nucleotide level (small or very long?) to support the conservation of this lncRNA. Are there potential structures conserved among these lncRNAs?

#2.7 T____wo consensus regions of ____FLAIL____ sequences among eleven disparate Brassicaceae genomes were shown in Fig. S9. ____Camelina sativa_ shared 98-nucleotide_ conserved sequences with Arabidopsis thaliana. In the future, it will be interesting to explore evolutional conserved structures among Brassicaceae genomes. However, these analyses are beyond the scope of the current manuscript.

7.In Fig. S4B, arrows may help to understand which seeds were selected.

__#2.8 Thanks. Arrows were included.____

__

Reviewer #2 (Significance (Required)):

This paper is a very nice piece of work and demonstrate the action of a long non-coding RNA (lncRNA) in trans on specific targets involved in the regulation of a developmental process, flowering. There is growing evidences that the non-coding genome hides large number of lncRNAs and there is little detailed genetic support for the action of lncRNAs globally. In contrast to many descriptive papers in the field, this ms demonstrates genetically, through an allelic series and complementation experiments, that this lncRNA locus is involved in flowering regulation and that its sense lncRNA recognizes target loci genome-wide, bringing interesting perspectives on potential new mechanisms of transcriptional regulation mediated by non-coding RNAs.

Reviewer #3 (Evidence, reproducibility and clarity (Required)):

In the manuscript by Jin et al authors characterize the FLAIL DNA locus in Arabidopsis (using a wide array of publicly available datasets), which produces a set of sense and anti-sense lncRNAs.

While our work on the FLAIL manuscript was ongoing we published the manuscripts where we presented these novel genomics methods and related data to capture nascent transcription and cryptic isoforms. We shared most data with TAIR, so we are happy to hear that these data are considered publically available.

Authors determined that the sense FLAIL lncRNA (or a set of sense lncRNAs, which isn't fully clear from the way the data are presented) is involved in flowering time in Arabidopsis based on the fact that the several flail mutants lead to the early flowering phenotype and this flowering defect is complemented by transgenic FLAIL DNA, meaning that FLAIL lncRNA acts in trans.

A series of experiments lead us to conclude that the sense isoform of FLAIL is responsible for the effect. We improved the data representation and writing of the manuscript to enhance accessibility.

The T DNA flail3 - mutant results in expression changes (up or down) of 1221 genes, including twenty genes linked to flowering in various ways. Expression of a group of these flowering-related genes could be either fully (for eight genes) or partially (for five genes) rescued by transgenic FLAIL. Authors also conducted the ChIRP-seq to determine which genes are physically bound by FLAIL lncRNA genome-wide. It was found that 210 genes in the genome are bound by FLAIL lncRNA. Comparison of the dataset of differentially expressed genes in the T-DNA flail3 mutant with the ChIRP-seq dataset of genes that are bound by FLAIL lncRNA revealed the 12 overlapping genes.

Among these twelve overlapping genes, four were found to be functionally connected to flowering with expression of these four genes being down in flail3 T-DNA mutant. Two out of these four genes were ruled out from being involved in the regulation of flowering by FLAIL. Authors conclude that the two other genes (Cir1 and Lac8) are responsible for the late flowering phenotype of flail mutants based on the three lines of evidence: (i) these genes expression is reduced in the flail mutant, (ii) FLAIL lncRNA directly interacts with these genes chromatin, (iii) the mutants of these genes were previously reported by others to display early flowering phenotypes too. While I find many of the findings reported in the manuscript very interesting, building a good foundation on which to expand the study and providing a very good leads for follow up experiments, I also have serious concerns about the manuscript in its current form.

Most importantly, this reviewer doesn't think that the mechanism of FLAIL lncRNA action was convincingly demonstrated. The main question would be how FLAIL lncRNA works and this question wasn't fully answered. It is great that FLAIL lncRNA binds directly to the two flowering-related genes, but what does it mean? Does it change any chromatin context of these genes quantitatively or qualitatively to affect the transcription? Or does it bind any components of transcriptional machinery and thus controls the transcriptional output?

#3.1 This manuscript addresses an important question in the field question: what is the evidence for functional elements in non-coding regions of genomes? Despite many efforts, convincing genetic support for these functions often remained limited. In addition to our strong genetic data, we provided new evidence that FLAIL recognizes targets with evolutionally conserved sequence motifs as part of the revision in Fig 4F and Fig. S9. Additionally, we plan to do ChIP-qPCR to identify histone modifications on FLAIL targets.

Additionally, flail3 T-DNA mutant affects the expression of 1221 genes and FLAIL lncRNA physically interact with 210 genes, so how can authors be fully sure that FLAIL lncRNA has only direct effect on these two genes and doesn't also contribute to the regulation of the upstream to Cir1 and Lac8 genes or even components of the transcriptional machinery that regulate these genes?

#3.2 We agree with this opinion. It is the reason why we felt stating this exact conclusion in our previous manuscript was justified. We improved accessibility of our manuscript in the revision, these clarify our model, that the trans-acting lncRNA sense FLAIL can interact with the chromatin regions of its target genes to directly or indirectly regulate gene expression changes involving flowering (Line 274).

Theoretically, doing RNA-seq in the amiR-FLAIL sense lncRNA mutant might have a chance of reducing the number of affected DEGs, making it easier to analyze the FLAIL targets, even if the allele can't be used for complementation experiments.

#3.3 Thanks for this suggestion. We plan to confirm key gene expression changes using amiRNA-FLAIL in full revision.

Also, auhors totally neglect putting the Cir1 and Lac8 genes into the context of flowering regulation, but it is something that needs to be done.

#3.4 ____We discussed roles of CIR1 and LAC8 in flowering regulation in Line 260-272. Flowering is fine-tuned to maximize reproductive success and seed production and by endogenous genetic cues and external environmental stimuli such as photoperiod. Nevertheless, many details of the flowering pathways and their integration remain to be investigated____. CIR1 is a circadian clock gene, induced by light and involved in a regulatory feedback loop that controls a subset of the circadian outputs and thus determines flowering time. Our GO analysis supports that a subset of DEGs are connected to the response to red or far red light that contains among other key flowering genes such as ____phytochrome interacting factor____ 4____ (PIF4) and CONSTANS (CO)____. FLAIL also binds the chromatin region of LAC8. LAC8 is a laccase family member that mainly modulates phenylpropanoid pathway for lignin biosynthesis____. Similar to flail, lac8 mutants flower early. While intermediates in this pathway or dysregulation of lignin-related genes could promote flowering in plants, the molecular connections of reduced LAC8 expression to effects on flowering time will require further investigation.

Lastly, the paper needs to be totally rewritten to be even properly evaluated. In its current state it reads like a very short draft.

#3.5 We reorganized the structure of manuscript, improved clarity and provided new mechanistic evidence in Fig. 4G and Fig. S9 to present a more complete manuscript.

The Abstract is weak, the Introduction is written in a such telegraphic style that it is barely readable, in many places there is no connections between sentences leading to the information appear to be presented as random, even if it isn't.

#3.6- We strengthened the Abstract by providing new evidence and improved for the Introduction.

The Results section is written rather rudimentary with information not being sufficiently provided to describe the results but rather scattered between the Results and Figure legends.

#3.7 Thanks for your suggestions, we described each FLAIL length and all constructs in detail in Results, put a schematic of T-DNA and CRISPR mutants in Fig. 1A, moved comparative genomics data to the end of Results and ensured all figures in order.

The Discussion is the best written part of the manuscript.

Thanks for your appreciation of the Discussion.

The Conclusion section carries no specific information and reads more like a little summary suitable for a review article rather than experimental paper.

#3.8 We agree this opinion, this paragraph fits Discussion better and Conclusion was removed.

Therefore, this reviewer thinks that regardless of how authors will choose to proceed with the current experimental version of the manuscript, it'd be in the authors' best interests to at least fully revise the paper before resubmitting anywhere. I'd also advise authors to seek professional editorial help specifically using an editor with the background in the plant sciences.

Authors might also want to consider moving Fig.3 into the Suppl. as it doesn't carry much weight or significance and perhaps make existing figures more meaningful and comprehensive and by including a better diagram of the locus (e.g., Fig. S1), etc.

#3.9 We thank this helpful suggestion. Fig.3 represents the RNA-seq data. In combination with supporting data in the supplementary material, it gives an easy visual readout of the reproducibility of the findings in replicates of stranded RNA-seq. In a new submission, we moved it to Fig. S5B and highlighted 13 differentially expressed flowering genes as well as sense FLAIL in flail3 that were rescued in complementation line in Fig. 3A. Moreover, we gave screenshots of FLAIL itself and four flowering related FLAIL targets in RNA-seq with a clear schematic representation of each locus. We believe these revisions improve Fig. 3.

It's not practical to list all issues with the writing as the paper requires total re-writing, so I can just make a few suggestions without any specific order to help authors improve the paper:

We are happy to improve our manuscript with the help of the reviewers. We addressed all comments including from reviewer #3 with a constructive spirit. However, since colleagues and reviewers #1 and #2 found the manuscript comprehensible to the point where they could make expert-level comments that illustrate understanding of the manuscript, a total re-writing did not feel like the most constructive suggestion to improve the manuscript.

--There is no statement anywhere that states the goal of the study.

#3.10____ We stated the goal of the study in line 50-69 and we think this is a misunderstanding. We summarized three issues currently exist in characterization of functional lncRNA in the last sentence of the first three paragraphs in Background: 1 in Line 50, the broad range of candidate hypotheses by which lncRNA loci may play functional roles call for multiple approaches to distinguish alternative molecular mechanisms. 2 in Line 59, functional characterization of trans-acting lncRNAs remains a key knowledge gap to understand the regulatory contributions of the non-coding genome. 3 in Line 69, the contribution of trans-acting lncRNAs to the regulation of distant flowering genes is currently unclear. So in the last paragraph of the background, we claimed that our goals are to address these questions through characterization of functional FLAIL lncRNA in flowering repression using multiple genetic approaches and various genomic data.

--No rational is provided on why authors decided to examine this specific genomic locus.

#3.11 For several years, our lab studies the rules and roles of non-coding transcription. We characterized and are characterizing several loci with evidence of non-coding transcription in a range of species. Early experiments suggested that FLAIL functioned in flowering, this manuscript clarifies that the function is executed as trans-acting lncRNA of the sense FLAIL isoform.

--Typically, the significance is in studying the function of lncRNA or a group of lncRNAs produced from a genomic locus, I don't think I ever encountered the instances when it was exciting to study just a specific genomic locus. If the locus does indeed have any significance for initiating the study, it needs to be explained.

#3.12 This study is remarkable in many aspects. We fully discuss key strengths in the discussion. First, we ____exhibit a trans-acting lncRNA FLAIL that represses flowering by promoting the expression of floral repressor genes as discussed in Line 247-281_; Second, in Line 284-306, we informed that this study provide a compelling model about how to apply _series of convincing genetic data____ to functionally characterize lncRNA loci. Third, in Line 307-312, evolutionary conserved FLAIL sequences across species is key to characterize the functional _microhomology in other _Brassicaceae.

--The locus can produce lncRNAs, but it can't harbor them.

#3.13 We clarified this confusion by enhancing ____presentation of Fig. 1 and providing a more clear description of each sequencing method and results in Line 81-99. Although we provided evidence that transcription of both sense and antisense FLAIL are more stable in hen2-2, they were clearly observed in chr-DRS in wild type and plaNET-seq in NRBP2-FLAG nrpb2-1 and sense FLAIL was even detected in TSS-seq and TIF-seq in wild type.

--No length of FLAIL lncRNAs or their range is provided in the first section of Results.

#3.14 We gave the length of sense FLAIL in Line 82 and antisense FLAIL in Line 86.

--On many occasions authors don't state rational for doing experiments, which leads to information often flowing as random.

#3.15 we enhanced clarity of the rational for each experiment and made some connections between sentences to make more fluent. For example, in sentences in Line 99, Line 113, Line 126, Line 159, Line 183, Line 214, and Line 219.

--What do authors mean by the subtitle "FLAIL characterizes a trans-acting lncRNA repressing flowering"? How can lncRNA FLAIL or FLAIL locus characterize lncRNA?

#3.16 We changed it to “FLAIL represses flowering as trans-acting lncRNA” in Line 112.

--Check all figures. E.g., Fig. 3B-E mentions only accession numbers for the genes.

#3.17 The systematic gene IDs are a valid way to represent data, in particular for genomics data since it facilitates cross-comparisons. To make it more accessible we also show systematic names of each gene in Fig. 3A-F, Fig. S6 and Table S3.

--It is not clear where exactly the T-DNA insertion is located relative to sense FLAIL in flail3 mutant (Fig. S4).

#3.18 We moved the schematic to clarify this to revised Fig. 1A and the exact T-DNA insertion site is mentioned in the legend.

--- What is the length of the complementing sense FLAIL lncRNA?

#3.19 We now include the length of the complementing sense and antisense FLAILs in Line 351-352.

--Check the description of each and every construct used and provide explanation for each in the Results. E.g., the pFLAIL:gFLAIL18/88 and pasFLAIL:gasFLAIL18/39 constructs aren't explained in Results, and can only be found in Fig. 2 legends.

#3.20 We described each construct including pFLAIL:gFLAIL18/88 and pasFLAIL:gasFLAIL18/39 constructs in Line 133, amiR-FLAIL-11 and amiR-FLAIL-11 in Line 149.

Reviewer #3 (Significance (Required)):

Tens of thousands of lncRNAs have been identified in various eukaryotes, but their biological roles have been shown only for a small fraction of them, and the mechanisms of their action are delineated for only a very few of them. Most of the advances on the field of lncRNAs are reported in metazoan, while the field of lncRNAs in plants is lagging far behind in terms of knowledge about lncRNAs with assigned biological functions or lncRNAs with delineated mechanisms of action. From this point of view, this reviewer is always excited to see any new functional plant lncRNAs for which either biological or mechanistic functions have been determined, and deems the information on this subject significant. The manuscript's findings are potentially very interesting and present a decent body of work that lays a very solid groundwork for future experiments. My main concern about the manuscript's significance in its current form is the fact that no real solid mechanism of action for the described lncRNA or a set of lncRNAs (?) has been demonstrated. The best mechanistically studied lncRNAs in Arabidopsis are involved in the regulation of flowering time, particularly those that function in the vernalization flowering pathway and to lesser extent in autonomous pathway. The new FLAIL lncRNA or lncRNAs (?) described in this manuscript also appear to regulate the flowering time in Arabidopsis, however more experiments would be needed to provide a definite conclusion about how direct FLAIL's effect is and how exactly it functions. That unfortunately obviously diminishes the significance of the manuscript and makes it potentially interesting only to researches studying flowering in Arabidopsis and even then the manuscript results would be incomplete to make solid conclusion.

Lots of functional phenotype have

Additionally, the manuscript requires complete re-writing.

We thank this reviewer for the appreciation of __a decent body of work and a very solid groundwork for future experiments. We are confident that our revisions make the manuscript more comprehensible to highlight the qualities of our manuscript more accessibly.____

__

Note: This preprint has been reviewed by subject experts for Review Commons. Content has not been altered except for formatting.

Learn more at Review Commons

Referee #3

Evidence, reproducibility and clarity

In the manuscript by Jin et al authors characterize the FLAIL DNA locus in Arabidopsis (using a wide array of publicly available datasets), which produces a set of sense and anti-sense lncRNAs. Authors determined that the sense FLAIL lncRNA (or a set of sense lncRNAs, which isn't fully clear from the way the data are presented) is involved in flowering time in Arabidopsis based on the fact that the several flail mutants lead to the early flowering phenotype and this flowering defect is complemented by transgenic FLAIL DNA, meaning that FLAIL lncRNA acts in trans. The T-DNA flail3 mutant results in expression changes (up or down) of 1221 genes, including twenty genes linked to flowering in various ways. Expression of a group of these flowering-related genes could be either fully (for eight genes) or partially (for five genes) rescued by transgenic FLAIL. Authors also conducted the ChIRP-seq to determine which genes are physically bound by FLAIL lncRNA genome-wide. It was found that 210 genes in the genome are bound by FLAIL lncRNA. Comparison of the dataset of differentially expressed genes in the T-DNA flail3 mutant with the ChIRP-seq dataset of genes that are bound by FLAIL lncRNA revealed the 12 overlapping genes.

Among these twelve overlapping genes, four were found to be functionally connected to flowering with expression of these four genes being down in flail3 T-DNA mutant. Two out of these four genes were ruled out from being involved in the regulation of flowering by FLAIL. Authors conclude that the two other genes (Cir1 and Lac8) are responsible for the late flowering phenotype of flail mutants based on the three lines of evidence: (i) these genes expression is reduced in the flail mutant, (ii) FLAIL lncRNA directly interacts with these genes chromatin, (iii) the mutants of these genes were previously reported by others to display early flowering phenotypes too. <br /> While I find many of the findings reported in the manuscript very interesting, building a good foundation on which to expand the study and providing a very good leads for follow up experiments, I also have serious concerns about the manuscript in its current form. Most importantly, this reviewer doesn't think that the mechanism of FLAIL lncRNA action was convincingly demonstrated. The main question would be how FLAIL lncRNA works and this question wasn't fully answered. It is great that FLAIL lncRNA binds directly to the two flowering-related genes, but what does it mean? Does it change any chromatin context of these genes quantitatively or qualitatively to affect the transcription? Or does it bind any components of transcriptional machinery and thus controls the transcriptional output? Additionally, flail3 T-DNA mutant affects the expression of 1221 genes and FLAIL lncRNA physically interact with 210 genes, so how can authors be fully sure that FLAIL lncRNA has only direct effect on these two genes and doesn't also contribute to the regulation of the upstream to Cir1 and Lac8 genes or even components of the transcriptional machinery that regulate these genes? Theoretically, doing RNA-seq in the amiR-FLAIL sense lncRNA mutant might have a chance of reducing the number of affected DEGs, making it easier to analyze the FLAIL targets, even if the allele can't be used for complementation experiments. Also, authors totally neglect putting the Cir1 and Lac8 genes into the context of flowering regulation, but it is something that needs to be done. Lastly, the paper needs to be totally rewritten to be even properly evaluated. In its current state it reads like a very short draft. The Abstract is weak, the Introduction is written in a such telegraphic style that it is barely readable, in many places there is no connections between sentences leading to the information appear to be presented as random, even if it isn't. The Results section is written rather rudimentary with information not being sufficiently provided to describe the results but rather scattered between the Results and Figure legends. The Discussion is the best written part of the manuscript. The Conclusion section carries no specific information and reads more like a little summary suitable for a review article rather than experimental paper.

Therefore, this reviewer thinks that regardless of how authors will choose to proceed with the current experimental version of the manuscript, it'd be in the authors' best interests to at least fully revise the paper before resubmitting anywhere. I'd also advise authors to seek professional editorial help specifically using an editor with the background in the plant sciences. Authors might also want to consider moving Fig.3 into the Suppl. as it doesn't carry much weight or significance and perhaps make existing figures more meaningful and comprehensive and by including a better diagram of the locus (e.g., Fig. S1), etc.

It's not practical to list all issues with the writing as the paper requires total re-writing, so I can just make a few suggestions without any specific order to help authors improve the paper:

--There is no statement anywhere that states the goal of the study.

--No rational is provided on why authors decided to examine this specific genomic locus.

--Typically, the significance is in studying the function of lncRNA or a group of lncRNAs produced from a genomic locus, I don't think I ever encountered the instances when it was exciting to study just a specific genomic locus. If the locus does indeed have any significance for initiating the study, it needs to be explained.

--The locus can produce lncRNAs, but it can't harbor them.

--No length of FLAIL lncRNAs or their range is provided in the first section of Results.

--On many occasions authors don't state rational for doing experiments, which leads to information often flowing as random.

--What do authors mean by the subtitle "FLAIL characterizes a trans-acting lncRNA repressing flowering"? How can lncRNA FLAIL or FLAIL locus characterize lncRNA?

--Check all figures. E.g., Fig. 3B-E mentions only accession numbers for the genes.

--It is not clear where exactly the T-DNA insertion is located relative to sense FLAIL in flail3 mutant (Fig. S4). --- What is the length of the complementing sense FLAIL lncRNA?

--Check the description of each and every construct used and provide explanation for each in the Results. E.g., the pFLAIL:gFLAIL18/88 and pasFLAIL:gasFLAIL18/39 constructs aren't explained in Results, and can only be found in Fig. 2 legends.

Significance

Tens of thousands of lncRNAs have been identified in various eukaryotes, but their biological roles have been shown only for a small fraction of them, and the mechanisms of their action are delineated for only a very few of them. Most of the advances on the field of lncRNAs are reported in metazoan, while the field of lncRNAs in plants is lagging far behind in terms of knowledge about lncRNAs with assigned biological functions or lncRNAs with delineated mechanisms of action. From this point of view, this reviewer is always excited to see any new functional plant lncRNAs for which either biological or mechanistic functions have been determined, and deems the information on this subject significant. The manuscript's findings are potentially very interesting and present a decent body of work that lays a very solid groundwork for future experiments. My main concern about the manuscript's significance in its current form is the fact that no real solid mechanism of action for the described lncRNA or a set of lncRNAs (?) has been demonstrated. The best mechanistically studied lncRNAs in Arabidopsis are involved in the regulation of flowering time, particularly those that function in the vernalization flowering pathway and to lesser extent in autonomous pathway. The new FLAIL lncRNA or lncRNAs (?) described in this manuscript also appear to regulate the flowering time in Arabidopsis, however more experiments would be needed to provide a definite conclusion about how direct FLAIL's effect is and how exactly it functions. That unfortunately obviously diminishes the significance of the manuscript and makes it potentially interesting only to researches studying flowering in Arabidopsis and even then the manuscript results would be incomplete to make solid conclusion. Additionally, the manuscript requires complete re-writing.

I have a tendency to do this. I keep aspiring to do bigger things once I reach a certain ideal, but I never do and so I just procrastinate away from achieving that ideal as a form of self-sabotage. It's such a foolish mechanism for self-sabotage

Schönes Tool für Gist.

I totally agree that providing good instruction makes students willing to use the app. However, I believe it's not just about the good instruction as such but also the layout and how it is presented. Personally speaking, when I need to use an app I'm not familiar with I immediately go to Youtube to find a tutorial; perhaps this works for me because I'm much of a visual learner.

the egregore of technology

how much of privacy is related to safety? how much of differentiated worldviews are emergent from privacy / psychological safety: to think, to create, to question -- and find resonance and develop our own views?

In my opionion, creating another name for science done by non-european ancient people is...I can't think of a word that doesn't sound super combative...but not ideal. All science (and all things ever) that any human has ever done has been in a cultural context. Why is it 'ethnoscience' when an Aboriginal person did it but it's just 'Science' when Pythagoras did it?

It's weird that I never really thought of other cultures creating different constellations. I knew of course that cultures have different names and stories for the same constellations. But for some reason I never really considered that different cultures created different constellations. It makes sense, it's just not something I ever learned or considered.

He really didn't say or do anything for the last two weeks. Just bided his time while people blamed the entire lack of response on the prime minister and mayor of Ottawa.

.c1

Author Response

Reviewer #2 (Public Review):

The visual system must extract two basic features of visual stimuli: luminance, which we perceive as brightness, and contrast, the change in luminance over space or time (this paper focuses on changes over time). Contrast is separately processed by ON and OFF pathways, which encode luminance increments or decrements, respectively. Contrast must be robustly detected even if the overall luminance changes rapidly, as might occur if an animal is moving in and out of shadows. This paper addresses how such a luminance correction occurs in the fly.

In the fly, three types of first-order interneurons - L1, L2, and L3 - transmit information from photoreceptors to the medulla, where ON and OFF encoding emerges. Previous work suggested that all three interneurons primarily encode contrast signals and that they project to distinct pathways: L1 to the ON pathway and L2 and L3 to the OFF pathway. Ketkar et al. show that, contrary to this model, these interneurons encode both contrast and luminance in specific ways and are not cleanly segregated into ON versus OFF inputs.

This study reveals several new insights into early visual processing that are interesting and well-supported by the data:

1) The authors show that behavioral responses to ON stimuli can compensate for rapid changes in luminance. However, the purported sole input to the ON pathway, L1, shows activity that is highly dependent on luminance. This suggests that a luminance correction must arise downstream of L1. These results are analogous to findings previously made by the same group regarding the OFF pathway (Ketkar et al., 2020). The previous paper showed that L2 provides contrast information to the OFF pathway, and L3 provides luminance information to allow for a luminance correction in downstream contrast encoding. But unlike the multiple inputs to the OFF pathway, the ON pathway was thought to only receive input from L1, provoking the question of whether L1 is able to provide both contrast and luminance information.

2) Using well-designed calcium imaging studies, the authors surveyed the responses of the three interneurons and found that they encode different stimulus features: L1 encodes both contrast and luminance, L2 purely encodes contrast, and L3 purely encodes luminance (with a different dependence than L1). These are interesting and important findings revealing how both contrast and luminance encoding are distributed across the three interneurons.

3) Using neuronal manipulations, the authors dissected the contributions of the three interneurons to ON and OFF behavior under changing luminance. These experiments showed that L1 and L3 are required for the luminance correction in the behavior. Moreover, the finding that all three interneurons contribute to both ON and OFF behavior contrasts with the existing model of segregated pathways. Thus, this paper could change the way we think about early visual processing in the fly: rather than relaying similar information to distinct downstream pathways, first-order interneurons relay distinct information to common pathways.

Overall, the major claims of this paper are important and supported by the experiments. There are just a few concerns that I would note:

Thank you for the overall positive evaluation of our work, as well as for the constructive criticism, which we are going to address below.

1) The authors state that they have shown luminance invariance in ON behavior (e.g. line 376-377 of the Discussion), but this is not entirely accurate: the ON behavior decreases as luminance increases. This is still an interesting effect since it's the opposite of what L1 activity does, so it's clear that the circuit is implementing a luminance correction, but it is not "luminance invariance".

As pointed out in response to essential comment #2, we carefully edited the manuscript to talk about ‘near’ luminance invariance, or data approaching luminance invariance. More prominently, we rephrased the text to highlight the need for a luminance gain to scale behavioral responses to contrast, even if the resulting behavior is not entirely luminance invariant.

2) The visual stimuli presented for most imaging experiments (full-field) are not the same as those presented for behavior (moving edges). It is possible neuronal responses and their encoding of luminance and contrast may differ if tested with the moving edge stimuli (if so, this would be concerning). The authors did image L1 with both types of stimuli and could compare these responses. Also, testing behavior at 34º and imaging at 20º presents a possible discrepancy in comparing these data.

We use moving ON edges in Figure 1, and these data suggest that the transient response of L1 scales with step changes in luminance, consistent with data in Figure 2B. Although we did not point this out in the paper, the L1 responses in Figure 1 also decay to different response levels, consistent with the luminance-sensitive component that static stimuli reveal in Figure 2. Furthermore, for other ongoing projects in the lab, we have for example measured physiological responses in L2 with the same stimuli used in behavior, and there is no discrepancy with the data reported here. Overall, there is no reason to believe, following a vast amount of literature in Drosophila and other flies, that LMCs would respond any different to moving vs. static stimuli.

We can additionally point out that the behavioral data of L3 silencing (at 34ºC) nicely correlate with physiological contrast responses of L1 and L2 (at 20ºC, predicted from electrophysiological recordings for LMCs in Ketkar et al. 2020, measured for L1 here). Many previous studies, for example in motion detection, have linked data from physiological recordings at room temperature with behavioral experiments done at higher temperature (e.g., Ammer et al., 2015; Clark et al., 2011; Creamer et al., 2019; Fisher et al., 2015; Leonhardt et al., 2017; Salazar-Gatzimas et al., 2016; Serbe et al., 2016; Silies et al., 2013; Strother et al., 2017). We therefore do not think that these are major concerns.

3) I find it puzzling that silencing L1 has little effect on ON behavior at 100% contrast and varying luminance (Figure 3A), but severely affects ON behavior to 100% contrast (and lower values) when different contrasts are interleaved (Figure S1). The authors note this but do not provide a clear explanation of why this might be the case. Aside from mechanism, it is not clear whether the difference is due to varying luminance in the first experiment or varying contrast in the second one (e.g. they could test 100% contrast without varying luminance).

The two stimulus sets used here do not allow us to pinpoint why the L1 silencing phenotype differs between them, since they comprise more than one difference as discussed above (see point 4) in “Essential Revisions”). We now include two additional experiments that dissect the role of different stimulus parameters (Supp. Figure 2). To understand whether the difference is due to varying luminance, we tested responses to ON edges of fixed (100%) contrast and luminance at the same stimulus parameters (motion duration, speed) as used in Figure 3, and did not find reduced turning responses when silencing L1. Thus, varying luminance does not change the effect of L1 on ON behavior. However, when repeating this experiment with a bright inter-stimulus interval, L1 silencing lead to a strong response deficit. Therefore, differences in the interval luminance explain the differences in the L1 silencing phenotype observed not only in this study but also across studies. Although we hypothesize a role of contrast adaptation that may function differently with altered contrast statistics, a more detailed investigation would be necessary to understand the mechanism. Nevertheless, our experiments allow us to conclude that L1 is not the sole major input to the ON pathway, even though it is required under certain stimulus conditions.

4) I do not entirely agree with the authors' interpretation of the L1 ort rescue experiment for OFF behavior. They state that rescue flies "responded similarly to positive controls". However, the graph shows that the rescue flies generally fall in between the mutant and heterozygote control flies; they resemble the controls at low luminance but resemble the mutants at high luminance. One may conclude that L1 is sufficient to enhance OFF behavior at low luminance, but it is a stretch to say it's a complete rescue.

Sorry, we just meant to say that they “responded similarly to positive controls (...) at low luminance”, but the sentence was badly written. We corrected this to: “L1 ort rescue flies responded similarly to positive controls at low luminances, rescuing responses to OFF edges at dim backgrounds.”

5) The authors typically use t-tests to analyze experiments with 2 variables (genotype and luminance) and 3 or more conditions per variable. This is not the most appropriate statistical test; typically one would use a two-way ANOVA. At the least, it should be clear whether they are performing corrections for multiple comparisons if performing many t-tests on the same dataset.

Thank you for the suggestion, we now use a two-way ANOVA followed by corrected pairwise comparisons and state this clearly in the figure captions (also addressed above in essential comment #5).

Reviewer #3 (Public Review):

Ketkar et al combine calcium imaging and behavioral experiments to investigate the encoding of luminance and contrast in 3 first-order interneurons in the Drosophila lamina: L1, L2, and L3, as well as the role of these signals in moving ON edge behavior across luminance. The behavioral experiments are well performed. The rescue experiments are particularly interesting. Together with silencing they support and nicely extend previous work showing that L1/2/3 are not simply segregated between ON and OFF pathways. My main issue is the link that the authors make between the cellular responses and the behaviors performed and therefore the overall conclusions and claims of the paper about the roles of contrast vs luminance encoding of each neuron type (particularly L1) in the behaviors.

Major concerns:

1) The authors state that the main behavior they study, namely optomotor response to moving light edges at 100% contrast, is "luminance invariant". A strict definition of this would be that behavioral responses are constant with increasing luminance. However, there are very few plots in this paper where this is the case. In almost all examples, the response is decreasing with respect to increasing luminance. The authors do qualify a "nearly" invariant behavior, but this does not change the fact that interpretation of the data in the context of the framing of the paper is often problematic.

We thank the reviewer for this critical comment. The main point (that we apparently failed to make clear enough) is that there is a clear requirement for a luminance gain. Physiological LMC responses measured using calcium imaging to ON stimuli in Figure 1, or predicted from previous electrophysiological recordings to OFF stimuli in (Ketkar et al., 2020) cannot account for any of the (control) behavioral data. We now edited the text to tone down statements about luminance invariance, and instead highlighted the need for a luminance gain.

2) The manuscript would benefit from clear definitions of luminance and contrast, as well as an explanation of how contrast and luminance sensitivity can be inferred from experiments. In particular, the authors use transient vs. sustained response properties in L1, L2, and L3 as indicators of contrast and luminance sensitivity, but this is not stated clearly. It would be important to explain this to the reader early on.

We now added definitions of general terms to the introduction and added data and analysis to the manuscript (Figure S1, and Figure 2B-D) to more clearly test which component of the neurons’ responses encode contrast or luminance.

3) In the manuscript, it is often stated that "calcium imaging experiments reveal that each first order interneuron is unique in its contrast and luminance encoding properties" (line 110). This was shown clearly for L2 and L3 in their previous work in Ketkar et al. 2020, with a welldesigned two-step stimulus that was able to tease apart contrast vs. luminance invariance. Unfortunately it does not seem that this level of experimental detail and analysis is applied to L1 here. In particular, the authors state " L1 encodes both contrast and luminance in distinct response components." Line 112, in the summary of their findings. I would not agree that the authors have actually shown this properly in this manuscript.

Addressed above, in point 6 of “Essential Revisions”

4) The results as they are stated, are at times not well supported by the data. The manuscript would benefit from a careful assessment of the accuracy and precision of the language used to interpret the data. Sometime just moving some conclusions to the discussion and explaining the assumptions made to reach a particular conclusion would be enough. A few of examples:

We carefully edited the entire manuscript, in addition to addressing the specific points below.

o Figure 2: "Lamina neuron types L1-L3 are differently sensitive to contrast and luminance". It is overall true that from the raw traces, the response are different. However the quantification in C-E only pertains to luminance.

As stated above, we now did further analysis on the contrast encoding properties of L1 and L2 and pointed out the major differences between these neurons (Figure 2B-D).

o Figure 3: "L1 is not required but sufficient for ON behavior across luminance". The data convincingly shows this. I would however point out that the statement "this data [..] highlights its behavioral relevant role of its luminance component" line 231 is an overstatement.

We deleted this statement at the end of the paragraph.

o Figure 6: "L1 luminance signal is required and sufficient for OFF behavior" the data presented shows convincingly that when L1 is inactive the behavior becomes (more) intensity variant. However, it does not show that it is the "luminance signal" in L1 that is required for this effect. In general, because L1 has a sustained and a transient response, it is difficult to strictly implicate one or the other in supporting any behavior, short of manipulating L1 to make it fully transient or fully sustained.

We agree. The figure title now reads “L1 function is required and sufficient for OFF behavior”.

o It is often not clear which conclusions stem from this work and which from their previous work Ketkar et al. 2020, or even other previous work on contrast sensitivity in particular. Clarifying this might help with my concern about statements not well supported by the data in this paper, and also justify their overall novelty. In general the manuscript assumes familiarity with this previous work, which is not always helpful for the reader.

As stated above, we now more clearly separate previous findings from novel findings in the abstract, and throughout the text. We also expanded the introduction to better explain the core concepts that are needed to understand this work, without having read Ketkar et al. 2020.

like in 1984, it's revealed that the "war" is not really a war and is just a way of keeping the citizens in each state under control. there will never be an end to the war but they encourage patriotism and hatred of the enemy to foster hope that there will be

https://dancohen.org/2019/07/23/engagement-is-the-enemy-of-serendipity/

Dan Cohen talks about a design change in the New York Times app that actively discourages exploration and discovery by serendipity.

This is similar to pulling out digital copies of books you're looking for instead of going to the library, tracking down the book on the shelf and in the process seeing and experiencing the books on the shelf which are nearby, or even the book that catches your eye across the aisle, wasn't in your sphere of search or interest, but you pick it up anyway.

How can we bring this sort of design back to digital experiences?

It's not just the algorithmic feeds which are narrowing our interests and exposure, but the design of our digital spaces as well.

It's interesting to see how far back exaggeration in the media has been a relevant thing. Today we have different social media sites, blogs, and podcasts, etc. that modernly display "yellow journalism", and that's just something that I thought wasn't a thing during the 1800s and 1900s.

It's challenging to come to terms with the fact that even at the disposal of tools like critical fabulation, the limitations (caused by archival violence) still persist. Sometimes, you just can't know.

One of the reactions I found the most surprising to The Prestige Trap was how many people disagreed with this explanation of prestige.

A decent chunk of readers took the materialist position that prestige is simply a function of how much money a job pays. While the two are strongly correlated, I think the latter is a lagging indicator of the former more than anything else.

This is made evident by the fact that there are plenty of jobs which are pretty high prestige but pay terribly (e.g., people who work on the Hill as interns comes to mind). The natural response is that those low-wage jobs lead to high-wage opportunities in the future. But if you talk to people on the ground who are actually thinking through these career decisions, it's rarely as simple as that. The path to the high-paying exit ramps is not as linear for the 21-year-old mind as you'd think.

I should have acknowledged that there are a lot of other reasons that prestige is valuable beyond it just guaranteeing a modicum of approval from your peers. For example, a point a lot of people made to me is how prestige is useful for getting noticed when you apply to other jobs down the road.

That's true, but I want to bracket that idea—prestige is only useful when it's a selection criterion that interviewers care about. So yes, prestige will open doors—but those doors themselves will be more of the same.

And that's what motivated me to write this essay in the first place—my friends were coming back from these high-prestige jobs and feeling miserable. What's the point of keeping a path open if you know you don't enjoy it?

Note: This preprint has been reviewed by subject experts for Review Commons. Content has not been altered except for formatting.

Learn more at Review Commons

Referee #1

Evidence, reproducibility and clarity

Summary:

Embryogenesis is characterized by rapid cell divisions without gap phases. How these cells achieve successive rounds of chromosome segregation in dozens of minutes without failure is of great interest to cell and developmental biologists. A key aspect of rapid divisions is the oscillatory nature of centrosome assembly, which aids in building the mitotic spindle during mitosis, and centrosome disassembly during mitotic exit. Polo kinase activation and localization to the centriole is essential for centrosome dynamics, but its molecular targets, timescales of activation and deactivation, and overall mechanism of action is still not fully determined.

This paper aims to build a mathematical model to tease out the mechanism of Polo recruitment to centrioles and transformation of centrosome scaffold proteins (Spd2/Cep192 and Cnn/CDK5RAP2) from inactive forms to functional, multimeric platforms. The authors posit that several features are critical to describe the dynamics of Polo, Spd-2, and Cnn: 1) a negative feedback loop that releases Polo from a centriole receptor, 2) a kinetic relay that allows Spd-2 to assemble, followed by Cnn, and 3) a disassembly mechanism driven by de-phosphorylation. They validate their model in several ways, most notably by introducing an Ana1 mutant that inhibits Polo binding to centrioles: their model predicts a delay in Polo accumulation which bears out in vivo.

The cell biology experiments of this paper are of high quality and well quantified, and I have no concerns there. However, the mathematical model elevates this study to the next level, and thus deserves greater scrutiny. I'm not concerned that the model doesn't get everything right, or that all of the parameters are correct. This is new territory. I think the value of models is their power to predict, rather than their power to explain existing data. The authors are giving the field a great hypothesis generator which we can use to plan experiments for the next 5 years. Then the model will be updated to be more accurate. Thus, this work represents a significant achievement.

Still, some key validations regarding phosphorylation rates are missing that could be easily tested. Furthermore, the study would be strengthened by greater understanding of the PCM disassembly. mechanism. Addressing these two points will improve my confidence in the mathematical model.

Major Comments.

1. This study builds a model that relies heavily on rates of phosphorylation and de-phosphorylation. Further, de-phosphorylation is assumed to be the key disassembly mechanism, but this has not been rigorously studied in fly embryos. Thus, two critical aspects of the model remain unverified.

Surely, the authors could test how changing phosphorylation rate (kcat S and kcat C) and de-phosphorylation rate (kdis) affects the recruitment and departure of Spd-2, Polo, and Cnn in vivo. This could be achieved by 1) titrating an inhibitor of Polo (e.g., BI-2536) or introducing a mutation in the T-loop of Polo (the equivalent of T210D or T210V in flies; T210D should raise kcat, while T210V should lower kcat; https://doi.org/10.1021/bi602474j), and 2) inhibiting a phosphatase such as PP2A, which is the presumed antagonist of Polo according to several C. elegans studies.

If their model adequately predicts the outcome of these two experiments (changing phosphorylation and dephosphorylation rates), I will be more convinced.

1. The models focus on Polo and Spd-2 pulses during mitosis, but ignore the disassembly phase of Cnn. Do Cnn levels drop during mitotic exit? Can this drop in Cnn be described by any of the authors' proposed models?
2. These models are described as the "simplest possible" yet have many unknown parameters. For example, Model 1 has 12 parameters, none of which have been determined experimentally. How did the authors land on these values? Is it possible that one could alter any combination of these parameters and achieve a similar outcome? Or, if the Kcat of Polo is changed two-fold, does the whole model fall apart (see above)?

Experimentally determining these parameters would greatly strengthen this paper, but I think that would require gargantuan effort that is beyond the scope of the current work. Instead, it is therefore critical to test how robust the model is by probing the parameter space. For example, could the authors show us what the model predicts (e.g., as in Figure 2C) when each parameter is changed by 2-fold? Presumably the authors have already done this, but I would like to see the outcomes.

Minor Comments.

-Figure 1. The authors should include representative images of centrosomes for the plots in panels A,C and D. The x-axes could have more informative labels (e.g., time relative to NEB). - Figure 1A. Much of Figure 1 has already been performed in C. Elegans, yet this fact is not mentioned until the discussion. For example, the pulsed nature of Polo and SPD-2 appearance and disappearance has been reported in C. elegans in Mittasch et al. 2020 and Magescas et al., 2019. These findings, and their implications for evolutionary conservation, should be mentioned in the main text (e.g. page 6 or 7). - Figure 2. It's hard to envision how a scaffold can both flux outward and be structurally strong. The mere fact that there is outward movement of scaffold chunks implies breaking of bonds, which indicates overall structural weakness. Are the authors talking about strength of the entire PCM, or just strength of the chunks? It would be great if the authors could clarify this. -Figure 2. One would think that scaffold flux and strength are anti-correlated. Perhaps this is the case? As far as I'm aware, previous studies of Cnn flux were performed primarily in S-phase, when there is presumably less need for PCM strength. What about during mitosis during chromosome segregation? Does the PCM become stronger during mitosis? Does Cnn flux decrease during mitosis? - Figure 2B. I would prefer a legend in the actual figure indicating what the different symbols mean. I found it difficult glancing back and forth between the text and the figure. - "We also allow the rate of 𝐶∗ disassembly to increase as the size of the 𝐶∗ scaffold increases, which appears to be the case in these embryos (Conduit et al, 2010)."

I can't find any analysis of PCM disassembly in this study. What are the authors referring to as "disassembly"? Do they mean departure of Cnn from the PCM in S-phase? Or, disassembly of the whole PCM during mitotic exit?

-"If the centriole and PCM receptors (Ana1 and Spd-2, respectively) recruit less Polo, the centriole receptor (Ana1) will be inactivated more slowly."

Is Ana1 a known substrate of Polo? This seems highly speculative. The authors should note that deactivation of Ana1 could be through various other mechanisms. Furthermore, Polo could be locally degraded as shown in human cells doi: 10.1083/jcb.200309035.

-"We note that our mathematical models are purposefully minimal to reduce the number of parameters and test possible mechanisms rather than to mimic experimental data." I appreciate this statement.

Significance

This paper aims to build a mathematical model to understand the cyclic nature of centrosome assembly and disassembly in fruit flies. Due to the conserved nature of the components (proteins in the system, such as Polo Kinase, Spd-2, and Cnn/CDK5RAP2), this model could likely be extended to a broad swath of eukaryotes. This approach is quite unique in the centrosome field, as only one other study (Zwicker et al., PNAS 2014) has tried seriously to model the growth kinetics of PCM, the outermost part of a centrosome. The field has been dominated by genetics and cell biology approaches, so implementing a mathematical model will advance the field and generate hypotheses, even if the model is not yet fully fleshed out. This paper represents a significant advance.

This study will be of broad interest to the centrosome field.

Expertise: centrosome biogenesis, mitosis, biophysics

Note: I am not sufficiently qualified to evaluate the mathematics underlying the model.

But maybe you don't need to do that. Maybe it would be instructive to take lessons from the traditional (pre-digital) publishing industry and consider how things like "print runs" and "reissues" factored in.

If you write a blog post in 2017 and the page is acceptable*, and then five years later you're publishing a new post today in 2022 under new design norms and trying to retrofit that design onto your content from 2017, then that sounds a lot like a reprint. If that makes sense and you want to go ahead and do that, then fair enough, but perhaps first consider whether it does make sense. Again, that's what you're doing—every time you go for a visual refresh, it's akin to doing a new run for your entire corpus. In the print industry (even the glossy ones where striking a chord visually was and is something considered to merit a lot of attention), publishers didn't run around snapping up old copies and replacing them with new ones. "The Web is different", you might say, but is it? Perhaps the friction experienced here—involved with the purported need to pick up a static site generator and set your content consistently with templates—is actually the result of fighting against the natural state of even the digital medium?

* ... and if you wrote a blog post in 2017 and the page is not acceptable now in 2022, maybe it's worth considering whether it was ever really acceptable—and whether the design decisions you're making in 2022 will prove to be similarly unacceptable in time (and whether you should just figure out the thing where that wouldn't be the case, and instead start doing that immediately).

I think that housing prices growing at this fast of a rate is terrifying. Housing prices are skyrocketing yet wages are not. There is a huge barrier for new home buyers because most young people cannot afford such an expensive payment for housing. I think it is crazy that most people cannot afford to pay for a home so they are forced to waste more money renting.

Being in a drought affects many different things and people. It affects farmers who need water to grow their crops. It affects ecosystems that rely on water such as the Great Salt Lake. It also impacts summer fun activities because there is not enough water in the lakes to boat on or play in.

This paragraph resonated with me because it is something that still happens everywhere. People are judged and put worth to themselves by their skin color or race. It is sad to think that our society is so shameless that it can treat other human beings as nothing just because they look different or just aren't consider to be "white". The paragraph really makes you think of the injustice that many people of color experience in the hands of society because they are not consider to be desirable, and black communities are those that suffer the most. I think it is important to educate people of the effects of racism and that youths shouldn't be suffering or having to worry whether society thinks they are bad people or criminals when they are still figuring themselves out.

I know that French speakers like this line. Juliette Gréco, for whom this song was written, specifically said of this line " C'est superbe."

Which I'm sure it is. But — having spent weeks thinking about it — I'm still not sure what it's actually saying.

"J'avoue" — easy enough: I confess.

"J'en ai bavé" — literally, to dribble, to drool, to slobber, to drip, to foam at the mouth (makes me think of another great song of this era, "L'Eau à la bouche"!). But figuratively — clearly the sense here — to suffer, to fall on hard times, to have a rough go of it.

But "pas vous"? With no punctuation, it's pretty baffling. Like, "I admit, I had a hard time of things... but you didn't, you were fine!" Or is it in the sense, "pas vous?," like "didn't you also?"

Anyway, I think the idea is more just to get as many possible V sounds into the line as possible. So I will allow myself to stop thinking about it.

Excellent - here, she is bringing this vision full circle to share how she came to be a climate writer, the very moment(s) that impacted her decision to enter this line of work. And, we get to understand that she became a writer (not just writer-adjacent) after all.

This foundational contradiction of capitalist production has to do with it's rigidity, and humanity's rejection of rigidity? I am struggling to understand this central point (but perhaps it's just too early for me to grasp it).

I notice this with the students at Cornell. Many of them do the readings like shit because they think if they just read it they can check it off. When you are responsible for your own learning your realize that there is no point in doing the reading badly because it's your learning and not the instructors. After the course is over you will regret not trying harder because you will have forgotten everything.

To situate these discussions in a contemporary contexts, I'm really intereted in how narratives of health shift over time, and what the political and social underpinnings of these shift are. It's surprising to read about sugar's historical roots as a medicinal, "health promoting" substance in the context of today's culture which heavily demonizes sugar as "unhealthy." Perhaps this shift could be accounted for by tracing sugar as a class signifyer. When sugar was a commodity that reflected wealth and colonial dominance, it gets marked as "healthy," but now as fatness and sugary diets are racialized markers of lower class, it is unhealthy and morally bad. Just some musings...have no idea if this actually tracks historically.

A single mom on disability struggles to provide food and clothing for her teenage daughter. A recent college graduate forgoes therapy. A young professional puts off buying a home and taking the next steps in his life. And a 74-year-old in a senior living community knows her monthly Social Security budget down to the cent.

The lives of these Utahns are all being shaped by spending around 50% or more of their income on housing each month.

Their challenges are part of a larger statewide housing crisis, one that is being blamed on both a shortage of homes and sluggish income growth that isn’t keeping pace with soaring real estate prices.

While past spikes in housing costs have priced people out of home ownership in Utah, the current affordability crisis is more all-encompassing — so it’s also stretching renters to the breaking point, said James Wood of the University of Utah’s Kem C. Gardner Policy Institute.

“I speak from personal experience,” said Wood, a senior fellow at the Gardner Institute. “I have people in my basement, and I’ve tried to help them find places. It’s really tough.”

Nearly one in five renters in Utah is severely cost-burdened, meaning they spend at least half their income on housing and often struggle to pay for food, transportation and other bills, according to federal data for 2013 to 2017. And more than 63% of the state’s lowest-income residents fall into this category, this data shows.

[Read more: Do you spend more than half your income on rent? Here are resources that can help.]

The disparities are particularly acute for Utahns of color, with a recent Gardner analysis showing that Black and Hispanic renters are more likely to face severe housing cost burdens. The research found that 32% of Black renters in the state spend more than half of their income on housing, making them almost twice as likely to face severe cost burden as white renters.

For a minimum wage worker in Utah, a rental home would have to cost $377 per month or less in order to be affordable, according to an analysis by the National Low Income Housing Coalition. But the average rent for a one-bedroom Salt Lake City apartment is nearly triple that, at $1,099 a month, according to a June report from the popular rental website, Zumper.

Wood said some of the state’s lowest-income residents receive public housing assistance, but there’s not enough money to reach everyone who needs help. Without government support, this group of Utahns lives on the brink of homelessness, with any additional hardship potentially pushing them over the edge.

“Whether it’s domestic violence, or whether it’s the loss of job or a health incident or a traffic accident,” he said. “That’s a disaster.”

Tara Rollins, executive director of the Utah Housing Coalition, notes that it’s easier to prevent people from losing their housing than it is to get them off the streets. The coalition advocates for increased wages and additional units of deeply affordable housing, to help people in this position before they’re pushed into homelessness.

Even those who are moderately cost-burdened — meaning they spend more than a third of their income on housing — face challenges, Rollins noted. But she doesn’t think that many Utahns and policy makers are paying enough attention to this swath of Utahns who are barely keeping their heads above water.

“Unless they feel it, see it, they don’t get it,” she said. “You can’t see somebody’s wallet and how empty it is.”

(Rick Egan | The Salt Lake Tribune) Jan Aus, a 74-year-old apartment resident in Sandy, says her rent keeps rising.(Rick Egan | The Salt Lake Tribune) Jan Aus, a 74-year-old apartment resident in Sandy, says her rent keeps rising. (Rick Egan/)

‘Nobody has my back’

When Jan Aus, 74, moved into a senior living community in Sandy seven years ago, she was shelling out $720 for rent each month.

“And then they raised it $10 two or three years after that,” she recounted. “And then, bing, they hit me with $65 a year.”

Today, Aus is paying $925 to live in her one-bedroom apartment ― a figure that sucks up the bulk of her Social Security check. She knows the amount she has to budget each month down to the cent: $1,251.80.

Aus said there are people who are “worse off than I am,” noting that she’s receiving government assistance available to low-income Utahns to help pay for electricity and food. She owns her car and considers her health insurance “good,” as long as she makes sure to get generic prescriptions.

Still, she said the amount of money she’s putting toward rent has become stressful, especially as she waits to see whether the apartment complex where she lives will raise her rent again this fall.

“It scares me,” she said. “And like I said, they’re going to hit me in September ... and it scares me to think they’re going to raise it again. I just feel like nobody has my back.”

If not for the federal pandemic stimulus checks, Aus said, she wouldn’t have any kind of savings, money she’s socked away in hopes that she can put a security deposit down on a more affordable apartment soon.

The problem, she said, is that there’s very little available in her price range of $800 to $900 a month, other than a room in someone else’s house.

“I don’t see myself going that way,” she said. “That to me is kind of scary. I think we need more affordable housing, I really do. Because it’s not going to get any better. To me, it’s going to get worse.”

‘I want to take care of more of my health’

Jazmin May has cut back her therapy sessions from once a week to once a month. The 24-year-old Salt Lake City resident can’t go in for an eye exam as soon as she’d like. And she’s had to ask her parents to chip in some money when her car needed repairs.

That’s all because rent claims about 50% of her income, and she has to stretch the rest to pay her other bills.

“For right now, it works for me,” she said. “I do wish I had more money left over in my paycheck just to be able to afford other things. I want to take care of more of my health.”

May says many of her other friends from college are also struggling, as they strain their early-career salaries to cover the cost of housing. Some have found it impossible and have gone back to live with their parents, she said.

She and a friend signed a lease for their two-bedroom apartment near Liberty Park in 2019, but the pandemic that arrived just months later quickly jeopardized the living arrangement. Her friend lost a retail job and had to move back to her family home in Ogden.

May said she considered looking for another roommate and decided it would be better for her mental health if she lived alone for a while.

Taking on the entire rental payment meant accepting a job at an area museum rather than continuing to hop between political campaigns — work that she loves but is too unreliable for her right now.

“I feel like I have sacrificed, in a way, my passion, to be able to afford housing, because I love campaigns and politics and outreach,” she said. “But campaigns are also not a stable job, and often you don’t get benefits. So I decided to just take a break from politics for a little.”

Even with the more predictable salary, May said home ownership seems like an unattainable goal at this point, especially since she’s worried that housing costs will always be one step ahead of her income growth.

She peruses rental listings for fun sometimes, but she’s not convinced she could find something cheaper, especially considering the pet fees she’d have to pay for her cat, Lilith. Her only other option, she said, would probably be to move into her parents’ home, as some of her friends have done.

‘You just kind of want to be an adult’

Fresh out of college 10 years ago, Orem resident Eric Wilson set a long-term goal of saving enough for a down payment on a home.

He’s passed up concerts he wanted to attend, vacations he wanted to take and movies he wanted to see. He’d love to buy the latest tech gadgets and the newest iPhone, but he’s socking away every extra penny in his investment portfolio instead.

Still, the 31-year-old marketing specialist said he doesn’t feel much closer to buying a house than he did a decade ago — and perhaps even further away, as he watches home values grow at warp speed compared to his slow-and-steady savings. So he can’t help but wish Utah’s economy would hit the tiniest snag.

“Just a little bit. Not enough to hurt anybody,” he half-jokes. “Just to make house prices go down.”

Making it especially hard to save is his current rent, which eats up nearly half of his salary.

Wilson has lived in the two-bedroom unit since shortly after he graduated from Utah Valley University. He had a roommate initially but opted not to get another one after his friend married and moved out.

“You just kind of want to be an adult and go off and do your own thing and have your own space and not have to worry about marking your milk,” he said. “But at a certain point, if prices keep rising, it’s not really feasible.”

Wilson had gotten about a fifth of the way to his goal of saving $100,000 when COVID-19 struck and his marketing agency had to cut jobs, including his. His unemployment lasted six months, forcing him to deplete the nest egg he’d spent so long accumulating.

He keeps browsing online real estate listings, despite knowing how far away he is from becoming a homeowner. The hobby is becoming increasingly demoralizing, though, he said.

Three years ago, he toured a modest home in a nice neighborhood that was pretty affordable for him, priced at a bit less than $200,000. Recently, he saw the same place had sold again for $415,000.

Wilson said he likes his apartment and knows he’d have to pay much more if he relocated. But he’s also weary of renting. He’s tired of feeling like he has to put off his life — and delay buying a dog or becoming a foster parent.

“I’m just kind of not at that point where I can do that space-wise,” he said. “But I would love to do that. And having a home would help make that possible.”

(Christopher Cherrington | The Salt Lake Tribune)(Christopher Cherrington | The Salt Lake Tribune)

‘This is where you belong’

Anna, 50, is a single mother supported by disability payments from Social Security and living in an income-restricted apartment complex in Holladay — but with around $700 left over each month after she pays her rent, she said, she’s still struggling.

While her monthly housing costs have ballooned from $850 when she first moved into the two-bedroom apartment in 2016 to $1,077 now, her disability income hasn’t increased at the same rate.

“It’s a huge stress,” Anna said. She fears she might be pushed out of the unit for speaking out about her rent increase, and The Salt Lake Tribune is not publishing her surname.

Among her biggest challenges is making sure her 13-year-old daughter can access nutritious foods, a goal she said is easier thanks to assistance from The Church of Jesus Christ of Latter-day Saints.

“Otherwise, my daughter probably would just be eating rice,” Anna said.

She’s also struggled at times to supply her daughter with new clothes that fit as she outgrows old ones.

Anna said she’s trying to save money, but “life keeps happening,” such as a car problem earlier this year that claimed everything she had saved and more. Sometimes, she worries that one misstep could land her and her daughter on the streets.

Drowning in monthly housing costs, Anna said she’s been searching online for a more affordable apartment in the hopes that she wouldn’t have to stretch so much to make ends meet — but she’s growing increasingly discouraged.

“I do keep on looking,” she said. “I keep hoping maybe I’ll find somewhere that is rent manageable as well as safe that I can move my daughter and I to so we can be able to provide for ourselves without having to rely on governmental programs completely. And basically feel like we’re being pushed into that hole that, well, if you can’t work for yourself, then this is where you belong. That’s how it feels.”

Hours after speaking with The Tribune, Anna received a notice taped to her door that her rent was being increased once again: to $1,122 starting July 1.

Crédito: Taylor Stevens, Bethany Rodgers

Word count: 2156 Copyright The Salt Lake Tribune Jun 10, 2021

Related items Latter-day Saints are overrepresented in Utah’s Legislature, holding 9 of every 10 seats Davidson, Lee. The Salt Lake Tribune; Salt Lake City, Utah [Salt Lake City, Utah]. 14 Jan 2021.

Salt Lake County keeps losing Latter-day Saints, and there are multiple theories as to why Davidson, Lee. The Salt Lake Tribune; Salt Lake City, Utah [Salt Lake City, Utah]. 14 Jan 2021.

Filmmaker gets to Sundance with a dark comedy inspired by his Latter-day Saint mission Means, Sean P. The Salt Lake Tribune; Salt Lake City, Utah [Salt Lake City, Utah]. 28 Jan 2021.

Jana Riess: The big question — Will Latter-day Saints follow their prophet and get the COVID vaccine? Riess, Jana. The Salt Lake Tribune; Salt Lake City, Utah [Salt Lake City, Utah]. 20 Jan 2021.

Sex therapist expelled from Church of Jesus Christ of Latter-day Saints Sarah Pulliam Bailey. The Washington Post; Washington, D.C. [Washington, D.C]. 23 Apr 2021: A.8.

Show more related items Search with indexing terms Subject Cost control Affordable housing Pandemics Home ownership Location Utah Company/organization Salt Lake Tribune Back to top About ProQuestContact UsTerms and ConditionsPrivacy PolicyCookie Policy Cookie Preferences Accessibility

Some pages ago, Timmy had mentioned that he didn't know what direction the play was supposed to be taking —satirical, comedic, etc. I initially disagreed with that thought, but this line right here makes me agree with him now. It's satire, I suppose, but unconvincingly so. So I guess it's just meant to be taken at face value.

It's very interesting that we have written in the stage directions, presumably by Strindberg himself, a departure from more presentational acting styles. Representational acting is often cited as having begun almost 10 years later in 1898 with Stanislavski's direction of The Seagull. Perhaps he wasn't a true innovator, just a popularizer!

A lot of these lines so far are way more verbose than you get used to reading in modern plays. If Mamet wrote this she would probably just say something like "shit" instead of this eight word set of three sentences. Modern playwriting seems to prioritize brevity in a way that these older plays don't do as much. It's not really about plot or content or anything, just how people speak. Maybe it says something about the character. A nervous thing? A sign of dull intellect? A lot of words for no added thought - could be a sign of someone who wants to be appeasing in conversation by affirming how intently they are listening.

My note is more about the stage directions. It's around this point where it dawned on me that the directions are very simple, and that it's affecting my vision of the play because it's so open-ended. There aren't even descriptions of an interiors. The implications for me are that I don't really imagine spaces, just people in a sort of void. I find it off-putting.

This line is funny to me, its like an old way of saying "you only live once"!

I wonder if this is supposed to make the reader question what makes a human, human? And what makes an animal, an animal? How do we separate the terms? Or is there no difference? Maybe this will make us dive into what it means to be human. It's an interesting concept because then I wonder what will the character's do that show their humanity?

What a great set of lines. The stage directions here are so simple - they fight and woyzeck loses, but really taking the time to imagine how a director would block out this interaction makes it so much more terrifying. and then the fart line. christ. Buchner was doing action hero one liners before film was even invented. I feel that everything he did was ahead of its time. It's a theme when I talk about him. As for Woyzeck, this scene makes it worth talking about the randomness of the horror that happens to him. At least in the tragedies of the Greeks and Shakespeare, you know why your life is bad, and usually you earned it. But with Woyzeck, the suffering has no meaning and no apparent cause. It just happens - and this is often how we experience suffering in real life. Like Buchner's illness - it came out of nowhere and unleashed its carnage nonetheless. It's a difficult reality to swallow, but it feels very true.

This is a very interesting thing about how people become great writers. It's a lesson I never forgot. The writer Hunter S Thompson once said that he would retype entire novels, just to get a feel for what the writer was thinking, to understand the method and the prose as each word follows. This is a lesson in greatness I think. When I started joke writing, I would write out a joke word by word and then take it apart. I would consider how important each word was in relation to the joke. It's something I'm still doing and has taught me the patterns, the turn, the punch, the changes. I feel like typing something out forces someone to pay a very close and detailed attention that wouldn't come simply from reading. When Georg Büchner translated Hugo's plays, I think that would have been a great and invaluable lesson in writing.

I thought that this was an interesting point because when a doctor tells you that you are sick and need this medicine that will relieve or cure you of your illness most people will pay whatever just to get better. It's like these drug companies really put you in a position where you have no other choice but to buy their drug in a time of vulnerability, especially if you have a rare disease that apparently that can be helped by this orphan drug.

This means you're a publisher and editor motherfucker. Stop saying you just host content.

you can search for and find information in the google world

but how can we have communities built around shared interests and purpose

don't see yet. That's what IndyLab offers

Link will be provided on launch this week!

IndyLab as a project will be launching on Open collective as a project on https://opencollective.com/open-learning-commons

supporting the https://opencollective.com/stopresetgo

effort to perform trans-disciplinary sensemaking and develop and pilot rapid, bottom-up, whole system change transition strategies for humanity

using indylab to connecting communities to discover and share their learnings in boundaryless collaboration

anti-matter to David Allen

Never getting Things done

Quite right too

Getting Things Done

assumes that you know what IT is that you need to do right here right now

But if you inclined to ask the question what is the IT that we really need to do

GTD is not for you at this stage

Do you think in some ways this is similar to the first time living completely on your own?

Robin Sloan's writing process sounds similar to that of Niklas Luhmann where he chose to work on things that seemed exciting and fun. This is, in part, helped by having a large quantity of interesting notes to work off of. They both used them as stores to fire their internal motivation to get work done.

The Magistrate and men's grievances with Lysistrata and the other women's defiance stems from their assumption that women are insignificant in war and civil duties because they have no stake in it. To combat the claim, Lysistrata explains just what significance women do have, beginning with the reminder of who is fighting the wars in the first place: sons. Were it not for the children women bore, such wars would not be fought. It's also a powerful claim, because it bears with it the loss of a child, and how that affects a mother specifically.

Semel, Jay M. “SEXUAL HUMOR AND HARMONY IN ‘LYSISTRATA.’” CLA Journal, vol. 25, no. 1, College Language Association, 1981, pp. 28–36, http://www.jstor.org/stable/44321687.

It's clear that they are writing the book in the perspective of maximizing body recomposition in whatever way possible. I believe that this is a little misguided for most people as I personally don't feel that good if I eat a ton of protein and nothing else. It makes my stomach feel a little poopy.

ReconfigBehSci. (2022, January 20). @timcolbourn @OmicronData As I said before, it’s not the function of this account to argue/advocate covid policies, but I will comment on the shape of the argument. The use of the frame “just delay” here seems hugely prejudicial. We don’t talk that way about flu or other diseases we might get repeatedly [Tweet]. @SciBeh. https://twitter.com/SciBeh/status/1484074977964011520

This is a common enough feeling (not about Publii, specifically; just the general concern for flexibility and control in static site generators), but when you pull back and think in terms of normalcy and import, it's another example of how most of what you read on the internet is written by insane people.

Almost no one submitting a paper for an assignment or to a conference cares about styling the way that the users of static site generators (or other web content publishing pipelines) do. Almost no one sending an email worries about that sort of thing, either. (The people sending emails who do care a lot about it are usually doing email campaigns, and not normal people carrying out normal correspondence.) No one publishing a comment in the thread here—or a comment or post to Reddit—cares about these things like this, nor does anyone care as much when they're posting on Facebook.

Somehow, though, when it comes to personal Web sites, including blogs, it's MySpace all over again. Visual accoutrement gets pushed to the foreground, with emphasis on the form of expression itself, often with little relative care for the actual content (e.g. whether they're actually expressing anything interesting, or whether they're being held back from expressing something worthwhile by these meta-concerns that wouldn't even register if happening over a different medium).

When it comes to the Web, most instances of concern for the visual aesthetic of one's own work are distractions. It might even be prudent to consider those concerns to be a trap.

It's unfair that she was treated differently because she was a women. If this hadn't had happened she would have been recognized for her hard work.

Author Response:

Reviewer #2:

Weaknesses:

The competition assay used in this study may not truly reflect the competitiveness of SSIMS males. The mating assay used 20 virgin WT females and 4 males (including both WT and SSIMS), resulting 5:1 sex ratio so the males are not really competing for females. A more competitive ratio (such as WT females: WT males: SSIMA males at 1:1:1) should be designed to address this. Also, the sperm competition assay mixed the mated WT females with SSIMS males for 12 days, allowing plenty of time for the females to remate with these males. Therefore, it's more like a sperm replacement assay rather than competition assay. The authors should either repeat it with a strict time control, or soften their statements for sperm competitiveness.

We have repeated the experiment at a 1:1:1 ratio as suggested. The new results are reported in the revised Figure 3. It is not clear to us how the timing of the mating experiments differentiates sperm competition versus sperm displacement, but we agree that sperm displacement is a better term to describe what we did. We have repeated the sperm displacement experiment with strict time control based on several published literature precedents and describe the results in the revised manuscript.

Some necessary information or statistics are not shown or mis-presented. For example, the alternative splicing diagram in Figure 1c likely was taken from the original transformer gene, but here it's the tTA gene so the male intron should be removed since it's not in the construct;

We have revised text in the manuscript to clarify some of these points. First of all, the male intron is still in the construct, even though we fused the intron to the tTA gene. The alternative splicing between males and females is caused by use of alternative 5' splice sites, which means the intron that is spliced out in males is just a smaller section of the intron that is spliced out in females. Use of an alternative 5' splice site in males means that a protein-coding sequence with multiple stop codons is incorporated to the mature mRNA. We do not support the precise splicing mechanism with empirical data in this paper, but this has been done in a number of previous publications (https://doi.org/10.1016/j.ibmb.2014.06.001; https://doi.org/10.1371/journal.pone.0056303).

Because the construct works as predicted (100% female lethality in the absence of tetracycline), and we did not change the genetic design in a way that would impact the mechanism of female lethality, we think there is little reason to believe that the splicing is occurring in a different way.

the panels of Figure 2 were not consistent to the legend and confusing; the statistics for different tetracycline concentration tests were not shown in Figure 2 or text to answer their hypothesis "(to) optimize rearing of SSIMS stock, …..we titrated Tet in the food";

We re-wrote the text describing Figure 2 to make the results more clear. We clarified in the legend that the symbol signifies p<0.0001 (we were not trying to imply that all experiments had this level of significance, only the ones marked with the symbol in the figure). We removed the word ‘optimize’ from the main text. Optimization was not the true aim of the experiment, and as the review points out, we did not statistically determine an optimal concentration of Tet. Our main goal was to show a dose- dependent response in the number of females surviving on Tet-free medium, which the data supports and which does not require statistical support.

Figure 3b shows 5-8 day old females were used but in the text it's 5-6 day, and it didn't mention the duration of the first crossing and time lag until the second crossing which are critical in such experiments; the conclusion and statistics for Figure 3c among tests with mixed males should also be mentioned.

We have corrected the figure (now Figure 3c) to indicate that the females were 5-6 days old. The first mating was for 5-6 days and there was no lag time between being co-housed with different males. We have performed multiple new experiments in revision that have been added to Figure 3. We have revised the discussion of these new experiments (and how they relate to the originally performed experiments) in the revised submission.

The discussion is largely towards the merits of SSIMS but missing some key points that might decide how it can be translated into applications or transferred to other species. First, the actual basis for tTA lethality that employed in this study is still unknown which is subject to suppression by a pre-existing inherent variation in the targeted field population. The very phenomenon may also be true for any gene-overexpression-based lethality including EGI lines generated here. Second, the complete penetrance observed from the relatively small sample size here can be hardly used to predict field or mass-rearing condition. Previous study showed that mutations in such lethal construct could occur at a one out of 10,000 frequency, and typical SIT program release millions of sterile insects every week. Third, while the authors claimed SSIMS is "one of the most complex engineered systems in insects", they also proposed that "the genetic design is likely to be portable to other species" without mention any potential obstacles along the way. Therefore, efforts should be made to give full picture of SSIMS including rain and sunshine.

We have added discussion of possible failure modes for this genetic biocontrol approach to the discussion section. We have also added text to discuss how the complexity of SSIMS is a potential obstacle to its translation to non-model organisms.

Peer review report

Reviewer: Nidia Bañuelos Institution: University of Wisconsin-Madison email: nbanuelos@wisc.edu

General comments

Questions of how and why grading practices change over time, how students, faculty, and administrators respond to grades, and the external pressures on grading practices (e.g. war, graduate school requirements) are inherently interesting! The authors have clearly done a careful job of tracking these – often minute, and likely, difficult to follow – changes at Dalhousie University. The manuscript is well-written and relatively easy to follow.

My biggest concern, reflected in the more detailed comments below, is that the authors could do a better job of explaining to the reader why these changes are interesting, important, and relevant to historians of higher education more broadly – even those who aren’t at Dalhousie. They do some of this at the very end of the paper and, indeed, this summing up of their findings and explanation of their relevance was my favorite part of the manuscript. I would suggest reorganizing the paper so that these bigger takeaways appear in the introduction and so that the reader is reminded of them at each major section break of the paper. For example, when the authors present a quote from a student who is concerned that grades have little to do with learning outcomes, they might remind us that one of their main arguments is that “decisions about university grading schemes had very little to do with actual pedagogy” (p. 15).

As it is written, the manuscript sometimes reads like a list of facts about grading changes. But, I think a reframing that focuses on the general importance of these changes could make the entire piece more engaging. More on this below…

Section 1 – Serious concerns

• Do you have any serious concerns about the manuscript such as fraud, plagiarism, unethical or unsafe practices? No

• Have authors’ provided the necessary ethics approval (from authors’ institution or an ethics committee)? not applicable

Section 2 – Language quality

• How would you rate the English language quality? High quality

Section 3 – validity and reproducibility

• Does the manuscript contain any objective errors, fundamental flaws, or is key information missing?

While I don’t notice any “objective errors”, I do think the paper has a major flaw (i.e. little explanation of the broader significance of this case study) and could benefit from additional information about the institutional context, the archival material, and external influences on grading trends. (Please see below.)

Section 4 – Suggestions

• Based on your answer in section 3 how could the author improve the study?

I. Most importantly, I would like to see an introduction that explains the authors’ general arguments about grading changes – including the trajectory of these changes at Dalhousie and why this arc contributes to our knowledge of the history of higher education more broadly. Then, the authors might continually remind us of the arc they present at the outset of their paper – especially when they are highlighting a piece of evidence that illustrates their central argument. To me, the quotes from students and faculty responding to grading changes are among the most interesting parts of the paper and placing these in additional context should make them shine even more brightly!

II. I’d like to read a little more about Dalhousie itself – why it is either a remarkable or unremarkable place to study changes in grading policies. Is it representative of most Canadian universities and thus, a good example of how grading changes work in this national context? Is it unlike any other institution of higher education and thus, tells us something important about grades that we could not learn from other case studies? I don’t think this kind of description needs to be particularly long, but it should be a little more involved than the brief sentences the authors currently include (p.3, paragraph 1) and should explain the choice of this case.

III. I’d also like to know more about the archival materials the authors used. The authors mention that they drew from “Senate minutes, university calendars, and student newspapers” (p. 3), but what kinds of conversations about grades did these materials include? At various points, the authors engage in “speculation” (e.g. p.4) about why a particular change occurred. This is just fine and, in fact, it’s good of the authors to remind us that they are not really sure why some of these shifts happened. But, they might go one step further and tell us why they have to speculate. Were explicit discussions of grading changes – including in inter- and intradepartmental letters and memo, reports, and other documents – not available in these archives? Why are these important discussions absent from the historical record?

IV. At various points, the authors make references to the outside world – for example, WWII (p. 5), the Veteran’s Rehabilitation Act (pp. 6-7), and British versus American grading schemas (p. 6). But, these references are brief and seem almost off-handed. I know space is limited, but putting these grading changes in their broader context might help make the case for why this study is interesting and important. Are the changes in the 1940s, for example, related to the ascendance of one national graduate education model over another (e.g. American versus British)? Are there any data on how many Canadian undergraduates enrolled in British versus American graduate programs over time? If so, I would share any information you might have on these broader trends.

Similarly, the authors make brief mention of the internal reaction to grade changes – quoting students or faculty minutes. But, it would be wonderful (space permitting) to have even more information the internal impact of these changes. Did they change faculty instructional practices? Did they seem to have any effect on students’ orientation to their learning? Did standardization reflect an increasing interdependence of departments, or did it contribute to their lessening autonomy? If the archival record doesn’t permit us to know these things, then this might be a limitation the authors note at the end of the manuscript. I noticed that the authors reference a secondary source on Dalhousie student experiences repeatedly (Waite, 1998). Even a little more from this text or another secondary source like it could help the reader better understand the impact of grade changes.

• Do you have any other suggestions, feedback, or comments for the Author?

This is a very nitpicky concern that doesn’t fit well elsewhere, so please take it with a grain of salt. I was surprised at the length of the reference list – it seemed quite short for a historical piece! I wonder, again, if more description of the archival material - including why you looked at these sources, in particular, and what was missing from the record – would help explain this and further convince the reader that you have all your bases covered.

Section 5 – Decision

Requires revisions: The manuscript contains objective errors or fundamental flaws that must be addressed and/or major revisions are suggested.

There are hundreds of possible search results when someone does a search, so how does the search engine choose which information to display? Is a blog article made on a regular basis and  maintaining a regular schedule a good way to ensure that Google crawls your pages more frequently and saves more possible query outcomes for future users? Maybe this is what causes some pages to be buried in other pages on Google searches.

This really gets to the heart of the matter, it is justifiable that Employees are the lowest of the priorities for an executive.

Based on the article priorities are: 1. External Customers - They bring money into the company 2. Your boss - They being money into you 3. Internal Customers (stakeholders or peers) - They make things work for external customers and your boss 4. Employees - They are paid to work for the company and are the lowest of the four priorities if you have to stack rank

it's really interesting and also nice that the author did include a lot of different examples and didn't just focus on violence that affected women that was done by men, but also different instances that include LGBTQ+ people and other men as well. i think it does a good job on further emphasizing how vital it is to have a system in a college that actively prevents SA and takes care of those who tell their stories because things like this can literally happen to anyone regardless of age, race, sex, and/or sexual orientation

i feel like porn has contributed a lot to this because all women are now expected to be the same way when engaging in sexual activity to the point where when women are having sex, it's likely they're going to feel subconscious about their acts/behaviors rather than just enjoying the pleasure of it but wondering if they're doing a "good job."

it's so interesting but also so horrible to see how for a man, sex is literally just seen as a reward

Author Response:

Reviewer #1 (Public Review):

Edmondson et al. develop an efficient coding approach to study resource allocation in resource constrained sensory systems, with a particular focus on somatosensory representations. Their approach is based on a simple, yet novel insight. Namely - to achieve output decorrelation when encoding stimuli from regions with different input statistics, neurons in the sensory bottleneck should be allocated to these regions according to jointly sorted eigenvalues of the input covariance matrix. The authors demonstrate that, even in a simple scenario, this allocation scheme leads to a complex, non-monotonic relationship between the number of neurons representing each region, receptor density and input statistics. To demonstrate the utility of their approach, the authors generate predictions about cortical representations in the star-nosed mole, and observe a close match between theory and data.

Strengths:

These results are certainly interesting and address an issue which to my knowledge has not been studied in-depth before. Touch is a sensory modality rarely mentioned in theoretical studies of sensory coding, and this work contributes to this direction of research.

A clear strength of the paper is that it demonstrates the existence of non-trivial dependence between resource allocation, bottleneck size and input statistics. Discussion of this relationship highlights the importance of nuance and subtlety in theoretical predictions in neuroscience.

The proposed theory can be applied to interpret experimental observations - as demonstrated with the example of the star-nosed mole. The prediction of cortical resource allocation is a close match to experimental data.

We thank the reviewer for the feedback. Indeed, demonstrating an ‘interesting’ effect in even such a simple model was one of the main aims.

Weaknesses:

The central weakness of this work are the strong assumptions which are not clearly stated. In result, the consequences of these assumptions are not discussed in sufficient depth which may limit the generality of the proposed approach. In particular:

1.The paper focuses on a setting with vanishing input noise, where the efficient coding strategy is to reduce the redundancy of the output (for example through decorrelation). This is fine, however, it is not a general efficient coding solution as indicated in the introduction - it is a specific scenario with concrete assumptions, which should be clearly discussed from the beginning.

2.The model assumes that the goal of the system is to generate outputs, whose covariance structure is an identity matrix (Eq. 1). This corresponds to three assumptions: a) variances of output neurons are equalized, b) the total amount of output variance is equal to M (i.e. the number of of output neurons), c) the activity of output neurons is decorrelated. The paper focuses only on the assumption c), and does not discuss consequences or biological plausibility of assumptions a) and b).

We have clarified the assumptions in the revised version. The original version did not distinguish clearly between assumptions that were necessary to allow study of the main effect, and assumptions that were included to present a full model but that could have been chosen otherwise without affecting the results. This has now been made much clearer. Regarding the noise issue (point 1), we have clarified the main strategy pursued by the model namely decorrelation, we acknowledge other possible strategies, and we make clear whether and how noise could be incorporated into the model. Regarding the biological plausibility of our assumptions (point 2).

Reviewer #2 (Public Review):

The authors propose a new way of looking at the amount of cortical resources (neurons, synapses, and surface area) allocated to process information coming from multiple sensory areas. This is the first theoretical treatment of attempting to answer this question with the framework of efficient coding that states that information should be preserved as much as possible throughout the early sensory stages. This is especially important when there is an explicit bottleneck such that some information has to be discarded. In this current paper, the bottleneck is quantified as the number of dimensions in a continuous space. Using only the second-order statistics of the stimulus, and assuming only the second-order statistics carrying information, the authors use variance instead of Shannon's information. The result is a non-trivial analysis of ordering in the eigenvalues of the corresponding representations. Using clever mathematical approximations, the authors arrive at an analytical expression -- advantageous since numerical evaluation of this problem is tricky due to the long thin tails of the eigenvalues of the chosen covariance function (common in decaying translation-invariant covariances). By changing the relative stimulus power (activity ratio), receptor density (effectively the width of the covariance function), and the truncation of dimensions (bottleneck width), they show that the cortical allocation ratio, surprisingly, is a non-trivial function of such variables. There are a number of weaknesses in this approach, however, it produced valuable insights that have a potential to start a new field of studying such resource allocation problems all across different sensory systems in different animals.

##Strengths

*A new application of the efficient coding framework to a neural resource allocation problem given a common bottleneck for multiple independent input regions. It's an innovation (initial results presented at NeurIPS 2019) that brings normative theory with qualitative predictions that may shed new light to seemingly disproportionate cortical allocations. This problem did not have a normative treatment prior to this paper.

*New insights into allocation of encoding resources as a function of bottleneck, stimulus distribution, and receptor density. The cortical allocation ratios have nontrivial relations that were not shown before.

*An analytical method for approximating ordered eigenvalues for a specific stimulus distribution.

##Weaknesses

The analysis is limited to noiseless systems. This may be a good approximation in the high signal-to-noise ratio regime. However, since the analysis of allocation ratio is very sensitive to the tail of eigenvalue distribution (and their relative rank order), not all conclusions from the current analysis may be robust. Supplemental figure S5 perhaps paints a better picture since it defines the bottleneck as a function of total variance explained instead of number of dimensions. The non-monotonic nonlinear effects are indeed mostly in the last 10% or so of the total variance.

We agree that the model is most likely to apply in the low-noise regime, as stated in the Discussion. The robustness of the results is indeed a worry, and indeed we have encountered some difficulties when calculating model results numerically due to the issue pointed out by the reviewer, and this led us to focus on an analytical approach in the first case. However, to test model robustness we have now included numerical results for several other covariance functions to demonstrate that, at least qualitatively, the results presented in the paper are not simply a consequence of the particular correlation structure we investigated.

In case where the stimulus distribution is Guassian, the proposed covariance implies that the stimulus distribution is limited to spatial Gaussian processes with Ornstein-Uhlenbeck prior with two parameters: (inverse) length-scale and variance. While this special case allowed the authors to approach the problem analytically, it is not a widely used natural stimuli distribution as far as I know. This assumed covariance in the stimulus space is quite rough, i.e., each realization of the stimulus is spatially continuous isn't differentiable. In terms of texture, this corresponds to rough surfaces. Of course, if the stimulus distribution is not Gaussian, this may not be the case. However, the authors only described the distribution in terms of the covariance function, and lacks additional detail to fill in this gap.

We would argue that somewhat ‘rough’ covariance structure might be relatively common, for example in vision objects have clear borders leading to a power law relation and similarly in touch objects are either in contact with the skin or they are not. In either case, we have now extended the analysis to test several other covariance functions numerically. We found that, qualitatively, the main effects described in the paper were still present, though they could differ quantitatively. Interestingly, the convergence limit appeared to depend on the roughness/smoothness of the covariance function, indicating that this might be an important factor.

The neural response model is unrealistic: Neuronal responses are assumed to be continuous with arbitrary variance. Since the signal is carried by the variance in this manuscript, the resource allocation counts the linear dimensions that this arbitrary variance can be encoded in. Suppose there are 100 neurons that encode a single external variable, for example, a uniform pressure plate stimulus that matches the full range of each sensory receptor. For this stimulus statistics, the variance of all neurons can be combined to a single cortical neuron with 100 times the variance of a single receptor neuron. In this contrived example, the problem is that the cortical neuron can't physiologically have 100 times the variance of the sensory neuron. This study is lacking power constraint that most efficient coding frameworks have (e.g. Atick & Redlich 1990).

We agree that the response model, as presented, is very simplistic. However, the model can easily be extended to include a variety of constraints, including power constraints, without affecting the results at all. Unfortunately, we did not make this clear enough in the original version. The underlying reason is that decorrelation does not uniquely specify a linear transform and the remaining degrees of freedom can be used to enforce other constraints. As the allocation depends only on the decorrelation process (via PCA), we do not explicitly calculate receptive fields in the paper and any additional constraints (power, sparsity) would affect the receptive fields only and so were left out in the original specification. We have now added clearer pointers for how these could be included and why their inclusion would not affect the present results.

The star-nosed mole shows that the usage statistics (translated to activity ratio) better explains the cortical allocation than the receptor density. However, the evidence presented for the full model being better than either factor is weak.

We agree that the results do not present definitive evidence that the model directly accounts for cortical allocations and as we state in the paper, much stronger tests would be needed. Our idea here was to test whether, in principle, the model predictions are compatible with empirical evidence and therefore whether such models could become plausible candidates for explaining neural resource allocation problems. This seems to be the case, even though the evidence in favour of the ‘full model’ versus the ‘activity only’ model is indeed not overwhelming (though this might be expected as the regional differences in activity levels are much greater than those in density). We have now added additional tests to show that the results are not trivial. We would also like to note that it is not obvious that the ‘full’ model would perform better than the ‘activity only’ model: for either we choose the best-fitting bottleneck width (as the true bottleneck width is unknown), and therefore the degrees of freedom are equal (with both activity levels and densities fixed by empirical data).

Reviewer #3 (Public Review):

This work follows on a large body of work on efficient coding in sensory processing, but adds a novel angle: How do non-uniform receptor densities and non-uniform stimulus statistics affect the optimal sensory representation? The authors start with the motivating example of fingers and tactile receptors, which is well chosen, as it is not overstudied in the efficient coding literature. However, the connection between their model and the example seems to break down after a few lines when the authors state that they treat individual regions as independent, and set the covariance terms to zero. For finger, e.g. that would seem highly implausible, because we typically grasp objects with more than one finger, so that they will be frequently coactivated.

Our aim was to take a first stab at a model that could theoretically account for neural resource allocation under changes in receptor density and activity levels, and by necessity this initial model is rather simple. Choosing a monotonically decreasing covariance function along with some other simplifications allowed us to quantify the most basic effects, and do so analytically. Any future work should take more complex scenarios into account. Regarding the sense of touch, we agree that the correlational structure of the receptor inputs will be more complex than assumed here, however, whether and how this would affect the results is less clear: Across all tactile experiences (not just grasps, but also single finger activities like typing), cross-finger correlations might not be large compared to intra-finger ones. Unfortunately, there is currently relatively little empirical data on this. That said, we agree with the broader point that complex correlational structure can be found in sensory systems and would need to be taken into account when efficiently representing this information.

The bottleneck model posited by the authors requires global connectivity as they implement the bottleneck simply by limiting the number of eigenvectors that are used. Thus, in their model, every receptor potentially needs to be connected with every bottleneck neuron. One could also imagine more localized connectivity schemes that would seem more physiologically plausible given the observed connectivity patterns between receptors and relay neurons (e.g. in LGN in the visual system). It would be very interesting to know how this affects the predictions of the theory.

We agree that the model in its current form is not biologically plausible. While individual receptive fields can be extremely localised, the initial allocation of neurons to regions we describe in the paper relies on a global PCA, and it is not clear how this might be arrived at in practice under biological constraints. However, our aim here was to specify a normative model that generates the optimal allocation and thereby answer what the brain should be doing under ideal circumstances. Future work should definitely ask whether and how these allocations might be worked out in practice and how biological constraints would affect the solutions.

The representation of the results in the figures is very dense and due to the complex interplay between various factors not easy to digest. This paper would benefit tremendously from an interactive component, where parameters of the model can be changed, and the resulting surfaces and curves are updated.

We have aimed to make the figures as clear as possible, but do appreciate that the results are relatively complex as they depend on multiple parameters. The code for re-creating the figures is available on Github (https://github.com/lauraredmondson/expansion_contraction_sensory_bottlenecks), making it easy to explore scenarios not described in the paper.

For parts of the manuscript, not all conclusions made by the authors seem to follow directly from the figures: For example, the authors interpret Fig. 3 as showing that activation ratio determines more strongly whether a sensory representation expands or contracts than density ratio. This is true for small bottlenecks, but for relatively generous ones it seems the other way around. The interpretation by the authors, however, fits better the next paragraph, where they argue that the sensory resources should be relatively constant across the lifespan of an animal, and only stimulus statistics adapt. However, there are notable exceptions - for example, in a drastic example zebrafish change their sensory layout of the retina completely between larvae and adult.

We have amended the text for this section in the paper to more closely reflect the conclusions that can be drawn from the figure. These are summarised below. The purpose of Fig. 3B is to show that knowledge of the activation ratio provides more information about the possible regime of the bottleneck allocations. We cannot tell the magnitude of the expansion or contraction from this information alone, or where in the bottleneck the expansion or contraction would occur. Typically, when we know the activation ratio only, we can tell whether regions will be expanded or contracted or whether both occur over all bottleneck sizes. For a given activation ratio (for example, a = 1:2, as shown in the 3B), we know that the lower activation region can be either contracted only or both expanded and contracted over the course of the bottleneck. In this case, regardless of the density ratio, the lower activation region cannot be contracted only. Conversely, for any density ratio (see dashed horizontal line in Fig. 3B), allocations can be in any regime.

In the final part of the manuscript, the authors apply their framework to the star nosed mole model system, which has some interesting properties; in particular, relevant parameters seem to be known. Fitting to their interpretation of the modeling outcomes, they conclude that a model that only captures stimulus statistics suffices to model the observed cortical allocations. However, additional work is necessary to make this point convincingly.

We have now included a further supplementary figure panel providing more details on the fitting procedure and results for each model. Given that we fit over a wide range of bottleneck sizes, where allocations for each ray can vary widely (see Figure 6, supplement 1A), we tested an additional model to confirm that the model requires accurate empirical density and/or activation values for each ray to provide a good fit to cortical data. Here we randomise the values for the density and activation of each ray within the possible range of values for each. We find that with this randomisation of the values the model performs poorly on fitting even with a range of bottleneck sizes. This suggests that the model can only be fitted to the empirical cortical data when using the empirically measured values.

This seems really simple, but it kind of feels tough to me, too. I think it's like that thing that was talked about in class the other day, it's just hard to tackle a completely new way of thinking and organizing at first.

https://medium.datadriveninvestor.com/3-improvements-to-your-pkm-system-just-by-no-longer-calling-your-digital-brain-a-second-brain-e5099188854

Words matter. Don't call your personal knowledge management system a "second brain" as it others something that is a part of you and your thinking.

(Not to mention that it's a marketing term for Tiago Forte's system. See: https://boffosocko.com/2021/07/03/differentiating-online-variations-of-the-commonplace-book-digital-gardens-wikis-zettlekasten-waste-books-florilegia-and-second-brains/#Second%20brain)

relating something pleasurable as we know it (chocolate) and ascribing a negative connotation to it just for the sake of colorizing and de-valuing people... It's interesting because here, "spice" takes on such a powerful division that we still see today except it's been subverted. That it, the indignation of white people when they are jokingly made fun of for not being able to take spice and how often they might perceive that as reverse racism when it is rooted in racism against other people...

"I'm just a developer."

This is probably an interesting quote because we use language so much that we forget what it is for, myself included. Its effect when it was invented and I think just like Baldwin said, it's just a tool and if not used properly, it will obscure the meaning that the speaker wants to convey.

The problem almost certainly starts with the conception of what we're doing as "building websites".

When we do so, we mindset of working on systems

If your systems work compromises the artifacts then it's not good work

This is part of a broader phenomenon, which is that when computers are involved with absolutely anything people seem to lose their minds good sensibilities just go out the window

low expectations from everyone everyone is so used to excusing bad work

sui generis medium

violates the principle of least power

what we should be doing when grappling with the online publishing problem—which is what this is; that's all it is—is, instead of thinking in terms of working on systems, thinking about this stuff in such a way that we never lose sight of the basics; the thing that we aspire to do when we want to put together a website is to deal in

documents and their issuing authority

That is, a piece of content and its name (the name is a qualified name that we recognize as valid only when the publisher has the relevant authority for that name, determined by its prefix; URLs)

that's it that's all a Web site is

anything else is auxiliary

really not a lot different from what goes on when you publish a book take a manuscript through final revisions for publication and then get an ISBN issued for it

so the problem comes from the industry

people "building websites" like politicians doing bad work and then their constituents not holding them accountable because that's not how politics works you don't get held accountable for doing bad work

so the thing to do is to recognize that if we're thinking about "websites" from any other position things that technical people try to steer us in the direction of like selecting a particular system and then propping it up and how to interact with a given system to convince it to do the thing we want it to do— then we're doing it wrong

we're creating content and then giving it a name

This is interesting as it says there is more than one type of privilege. As a person who's has been told that they are privileged without any context on concept. This helps show that there can be more than just the surface level privilege

I feel like this is incredibly true for people that don't like to acknowledge their privilege. I feel that when people are asked to address the idea of privilege they feel as if someone is blaming them for the privilege they possess, when really they are just being asked to acknowledge that it's real.

This was interesting to me because part of growing to understand privilege is not just what we benefit from it or our advantages from it, but it's also learning about those who are put at disadvantage from it or don't share the same privilege.

It's the teachers job to be mindful of who has spoken and who has not. You need to make sure that not just one or two students are giving most of the opinions as otherwise the discussion will become very skewed.

The students will want to share more if you build some efficacy by asking them follow up questions, summarize, and make their opinion feel welcome. This should be done especially if it's the students first time talking. As soon as you get them to share once, latch on to it and make them want to do it again.

It's always easy to think a issue is very basic and one sided when just your opinion is in the mix. It's only after we hear other peoples perspectives who have grown up in completely different environments and settings that we realize how wrong we truly were!

I still find that crazy. I can't believe that protestors aren't able to speak their mind without getting gassed and shot with rubber bullets. It's not just like that in the United States, protestors in France, Hong Kong, and a lot of other countries get the same treatment because they ask for change.

I don't agree with this assessment. It's very short-sighted.

Just as a reminder, since it's a new term: value independent from anything outside themselves. They are not just means to be employed for human purposes (that would be instrumental value); they are ends in themselves.

Sorry but I don't think the table works well in the html - it's a very long table, you can't scroll through years until you get to the bottom and can't see everything all in one screen. I suggest you just include a link to the excel spreadsheet, as it's easier to get a overall view of it that way.

Reviewer 2. Rhonda Bacher It is good to have alternative workflows for single-cell analysis, and I am glad to see the authors have submitted the package to Bioconductor. I hope the authors maintain the package and update with new methods as necessary such as if new normalizations or batch corrections are developed. I only have two comments that I hope the authors try to clarify further:

1. The statement starting with "Optionally, after batch-to-batch normalisation, we also..." should not be in that location. It seems to suggest to readers that this is the recommended method, whereas later that is not the case. In these sentences the manuscript also claims that this normalization approach is more "robust" without providing any evidence or citation.
2. It's still not completely clear to me how the authors extension of the sc-qPCR method is different from MAST. The same authors of the qPCR method extended it here: "MAST: a flexible statistical framework for assessing transcriptional changes and characterizing heterogeneity in single-cell RNA sequencing data". MAST is also an LRT, but I am assuming that here you are not using the detection rate as a covariate? That's OK if true, it just needs to be clear to the reader. I imagine this could be a frequently asked question by users down the road, so even a sentence on how it is different from (or similar to) MAST would help. Suggestion only: I may have missed it, but it might be helpful to include a statement that says something like "Statistical methods for single-cell analysis are constantly evolving. Here we have implemented XX. The flexibility of ascend allows it to adapt as future methods are developed and prove useful".

She thinks there are a lot of others that are fighting to stop violence

I think it is important for everyone to understand that we all have different forms of privilege. Even though some may have more than others, or some some of us may feel that we have none, it is important to for us to be aware of the perceptions other people will have of us, and if we can do this, we all become much easier to work with.

I greatly connected with this quote. As a white woman I don't face much discrimination, but I realize that the privileges I do have I benefit from. As a white woman there are probably several instance where privileges I have benefitted me. While I come from a small town I see a privilege I get there is that everyone knows my mother, while some may have this same instance but it may come as discrimination against them.

It's interesting to see a president acknowledge the need for renewable energy at that time, as I tend to assume that the conversation regarding climate change has only begun recently. It's quite disappointing and a little concerning that he didn't follow through on his goals though. With the dates that we will be "past the point of reversing climate change" constantly changing, it feels like we are never making progress fast enough. I fear that just as Carter's goals were prophetic, the ones we are setting in this age will be as well, but I'm also optimistic that the more outlined goals such as the Biden Administration's plan cited earlier in the piece will have more success.

People believe so much in certain miracle sures that it's like a religion for them. It will take a lot to sway sellers of BOO away from it. I really hope that they are able to shut down this company before people ingesting it become ill. Lead and arsenic are very dangerous, especially to children. I shudder to think about the baby soaking in a bathtub of lead-filled BOO.

These topics seem common problems with health misinformation. For years the anti-vaccine debate was centered around autism. It's sad that people keep falling for the same misinformation and scare tactics.

"Products can't be bought in stores." Another red flag. Why not? Buyers of this product should've further questioned how safe and legitimate BOO was because they aren't a built up brand, like Native deodorant for example, where they've built up trust, earning credibility for how effective and safe their deodorant is. It's the equivalent of buying a product from an unknown individual in an unfamiliar town on Ebay. If you wouldn't do that, why would you buy BOO or anything else from an individual you don't know? You may as well buy the $19.95, plus shipping and handling, fat burner you saw on the shopping channel at midnight that can only take your debit card as form of payment. It would be just as credible, safe, and beneficial.

To me, this sentence seems to foreshadow a greater main point to this article, besides the obvious success of Black Oxygen Organics. This sentence hints that not only have the pandemic and its affects allowed for many products, not just BOO, to find success, but that it's also created a space for misinformation and conspiracy theories to thrive.

I find it interesting that there are groups devoted to debunking the false claims of MLMs just as there are fact checkers all over social media. In this case, activists are using old fashioned journalism by ebedding themselves in the culture and documenting what is happening. This infiltration tactics are great because it allows first hand accounts and direct source reporting to give insights into what actual sellers are taliking about in these group chats. It's also using the same social media outreach tactics to find like-minded people who want to debunk false claims, in the same vein group organizers try to recruit new customers and people to sell high priced dirt.

The image of maintaining a health livestyle has been glamorized more than ever. With direct sellers taking advantage of the rush toward natural remedies just to boost their sales during a pandemic really makes me question our well being as the so called "consumer's" we are in the eyes of our nation. It's clear they do not care about our well being all they care about is the money at the end of the day. So why should we trust our government or the business'es that are in place to have our interest at heart?

It's not just a daycare option, and I appreciate the fact that it is not viewed as such. Parental involvement is not simply donating to the seasonal fundraiser or being a member of the PTO/A.

I want to know more about how you go about this. I have experienced this many times in books like “Youth in Revolt '' which made me laugh and feel happy. While other books like “IT“ made me feel fear through descriptive language and an intriguing story. But when I'm writing something I notice that I tend to go numb and I find it hard to convey my feelings. I'm not sure if it's me trying to use big words to make myself smarter or if I'm just not good at sharing my feelings but I would like to know how to do this. I enjoy books much more when they make me feel a certain way (good or bad).

but it's not just one single vote that is in the ballot

This is true. Getting rid of it doesn't mean it's gone forever; it'll just induce other problems that human go through.

This statement hit because it is soooo true. We're constantly regulating ourselves and just deeming that others will accept that and refuse to think otherwise. There are so many regulations going on today (although I think the vaccine mandate is important it's also a prime example) that we just threw out there and hoped it would stick. Obviously not

I agree that there is constant suffering. That saying "If it's not one things it's another" is prime example of the human race rolling with the punches and just blocking or dodging as we go along. Life is a constant battle and either you win some or lose some.

This goes back to the classic bullying concept. Knowing that you are upsetting someone else makes some people almost feel better about themselves. They know they can set someone off just by leaving a single comment, even just 1 word at some times. They seem to feel more important because others disagree with them. It's not necessarily about being right, but about knowing its bothering other people and getting a reaction out of them.

This quote not only relates back to comment sections, be even something a little more personal like text messages, which I think just shows how important it really is. It's so simple for people to misinterpret messages like "okay." because you don't have that tone and gesture. Some may read it simply as okay, and some may read it as anger or annoyance. Without the face to face contact, it is incredibly hard to communicate the emotion behind conversations, and in comment sections, where people are often seeming to look for drama, misinterpretations are even more prevalent.

I feel healthcare and gun rights are the fact the biggest social media argument wars between people. Arguing on those cases is like lighting the fire and it won't stop somewhere you prefer too because it's just so many people have been hurt from the cases or are dealing with something with it. So I think those cases are very sensitive and people still tend to argue about it which would eventually lead to emotionally hurting someone hundreds miles away from you. For my own personal sake, i never try to argue with this and take my own space from it.

It's not enough for systems / institutions to just diversify their students and staff, they have to actually commit to a systemic changes that better distributes power

The overall website is extremely cluttered with different updates, subscription options, promotions, game scores and articles being on the same page. This is a bad example of website accessibility as it may be a sensory overload for some individuals and a bit difficult to understand especially for those individuals using audio softwares that read the contents of the page out loud.

This stands out to me because I have noticed this a lot in my personal life recently. Perspective is everything and affects what we say and do. I have been practicing being more understanding towards other people in my life lately so this quote resonated with me because it is something I've been focusing on.

Wow, this goes to show that being a good listener is more important than giving good advice. We don't always need to FIX something. Most times people just need us to LISTEN and UNDERSTAND.

Good name for this. "Maximizer" sounds good by its name. But when you see what the maximizer does—it's actually not good at all.

Author Response:

Reviewer #1 (Public Review):

The underlying data are dominated by data from the UK Biobank, which means that, in effect, only few samples for the 25-50 age group are available. This may not be a big issue in terms of estimating smooth trajectories, but may limit comparisons to the reference model in certain cases (e.g. early disease onset) where this age range may be of particular interest.

We show per site evaluation metrics, cross validation, and additional transfer examples. These additional analyses show that the model performance is not driven solely by the UKB sample. However, we agree with this comment and have also updated the limitation section (in the Discussion) regarding the overrepresentation of UKB and included a statement regarding the known sampling bias of UKB.

The manual QC data is somewhat limited as it is based on a predominantly younger cohort (mean age ~30yrs). Furthermore, the number of outcome measures (cortical thickness and subcortical volume) and the number of data modalities (only structural MRI) are limited. However, as the authors also state, these limitations can hopefully be addressed by incorporating new/additional data sets into the reference models as they become available.

We have added further details regarding the quality checking procedure to the methods section and improved the clarity of directions for implementing the scripts, including an interactive link to view an example of the manual QC environment, on the QC GitHub page to enable others to reproduce our manual QC pipeline.

Reviewer #2 (Public Review):

1. The evidence that the model will generalize ("transfer" as per the authors) to new, unseen sites, is very limited. To robustly support the claim that the model generalizes to data from new sites, a cross-validation evaluation with a "leave-one-site-out" (or leave-K-sites-out) folding strategy seems unavoidable, so that at each cross-validation split completely unseen sites are tested (for further justification of this assertion, please refer to Esteban et al., (2017)). The "transferability" of the model is left very weakly supported by figures 3 and 4, which interpretation is very unclear. This point is further developed below, regarding the overrepresentation of the UK Biobank dataset.

We thank the reviewers for this suggestion and have addressed the concern regarding generalizability in several ways. First, we ran an additional 10 randomized train/test splits of the data in the full sample. These new analyses show the stability of our models, as there is very little variation in the evaluation metrics across all 10 splits. These results are visualized in Figure 3 – Supplement 2. However, the static Figure 3 – Supplement 2 is challenging to read, simply because there are many brain regions fit into a single plot. Therefore, we also created an online interactive visualization tool that shows the image of the brain region and the explained variance when you hover over a point (see the screenshot of the online tool below). This interactive visualization was created for all supplemental tables for easier exploration and interpretations and we now recommend this tool as the primary method to interrogate our findings interactively. Second, we updated and expanded the transfer data set to include 6 open datasets from OpenNeuro.org (N=546) and we provide this example dataset on our GitHub with the transfer code. This simultaneously provides a more comprehensive evaluation of the performance of our model on unseen data and more comprehensive walk-through for new users applying our models to new data (sites unseen in training). Finally, we added per-site evaluation metrics (Figure 3 – Supplement 3) to demonstrate that performance is relatively stable across sites and not driven by a single large site (i.e., UKB). As site is strongly correlated with age, these visualizations can also be used to approximate model performance at different age ranges (i.e., 9–10-year-old performance can be assessed by looking at ABCD sites evaluation metrics, and 50–80-year-old performance can be assessed by looking at UKB evaluation metrics). Moreover, we would also like to emphasize that we should not expect that all sites achieve the same performance because the sampling of the different sites is highly heterogeneous in that some sites cover a broad age range (e.g., OASIS, UKB) whereas other sites have an extremely narrow age range (e.g., ABCD).

1. If I understand the corresponding tables correctly, it seems that UK biobank data account for roughly half of the whole dataset. If the cross-validation approach is not considered, at the very (very) least, more granular analyses of the evaluation on the test set should be provided, for example, plotting the distribution of prediction accuracy per site, to spot whether the model is just overfitted to the UKB sample. For instance, in Figure 4 it would be easy to split row 2 into UKB and "other" sites to ensure both look the same.

We have addressed this comment in response to Reviewer 1 above.

1. Beyond the outstanding work of visually assessing thousand of images, the Quality Control areas of the manuscript should be better executed, and particularly lines 212-233): 3.a. The overall role of the mQC dataset is unclear. QC implies a destructive process in which subpar examples of a given dataset (or a product) are excluded and dropped out of the pipeline, but that doesn't seem the case of the mQC subset, that seems a dataset with binary annotations of the quality of the FreeSurfer outcomes and the image.

We have addressed this in response to Reviewer 1 above. We included the manual QC in this work, because in prior work by our group (https://www.biorxiv.org/content/10.1101/2021.05.28.446120v1.abstract) that leveraged big data and relied on automated QC, reviewers often criticized this approach and claimed our results could be driven by poor quality data. Thus, in this work we wanted to compare the evaluation metrics of a large, automated QC data set with the manual QC dataset to show very similar performance.

3.b The visual assessment protocol is insufficiently described for any attempt to reproduce: (i) numbers of images rated by author SR and reused from the ABCD's accept/reject ratings; (ii) of those rated by author SR, state how the images were selected (randomly, stratified, etc.) and whether site-provenance, age, etc. were blinded to the rater; (iii) protocol details such as whether the rater navigated through slices, whether that was programmatic or decided per-case by the rater, average time eyeballing an image, etc; (iv) rating range (i.e., accept/reject) and assignment criteria; (v) quality assurance decisions (i.e., how the quality annotations are further used)

These details have been added to the methods section where we describe the manual QC process. We have also updated the QC GitHub with more detailed instructions for using and include a link to view an example of the manual QC environment.

3.c Similarly, the integration within the model and/or the training/testing of the automated QC is unclear. The responses to Reviewer 1 above and our revisions to the methods section should also clarify this. In brief, QC was performed on the data prior to splitting of the data to assess generalizability.

Additional comments

• Repeated individuals: it seems likely that there are repeated individuals, at least within the UKB and perhaps ABCD. This could be more clearly stated, indicating whether this is something that was considered or, conversely, that shouldn't influence the analysis. We have clarified in the methods section that no repeated subjects were used in the dataset.
• Figure 3 - the Y-axis of each column should have a constant range to allow the suggested direct comparison. We have changed Figure 3 to have a constant range across all test sets.
• Tables 5 through 8 are hard to parse - They may be moved to CSV files available somewhere under a CC-BY or similarly open license, and better interpreted with figures that highlight the message distilled from these results.

We agree with the reviewer about the difficulty in summarizing such a large number of results in an easily digestible manner and that tables are not the optimal format to achieve this. Therefore, we have created interactive visualizations for Tables 5-8 that make exploring the evaluation metrics much easier. All the CSV files are also hosted on our GitHub page in the metrics folder (https://github.com/predictive-clinical-neuroscience/braincharts/tree/master/metrics).

• Lines 212-214 about the QA/QC problem in neuroimaging are susceptible to misinterpretation. That particular sentence tries to bridge between the dataset description and the justification for the mQC sample and corresponding experiments. However, it fails in that objective (as I noted as a weakness, it's unclear the connection between the model and QC), and also misrepresents the why and how of QC overall.

We have considerably expanded upon our motivation for using a manual QC approach and the steps this entails, which should address this issue.

• The fact that the code or data are accessible doesn't mean they are usable. Indeed, the lack of license on two of the linked repositories effectively pre-empts reuse. Please state a license on them. We thank the reviewer for this suggestion. We have updated both repositories to include a license file.
• Figure 1 - caption mentions a panel E) that seems missing in the figure.

We have corrected this mistake in the caption of Figure 1.

• There is no comment on the adaptations taken to execute FreeSurfer on the first age range of the sample (2-7 yo.).

We did not make adaptations of the Freesurfer pipeline for this age range and have added this to the limitation section.

• Following up on weakness 3.c, while scaling and centering is a sensible thing to do, it's likely that those pruned outliers actually account for much of the information under investigation. Meaning, EC is a good proxy for manual rating - but Rosen et al. demonstrate this on human, neurotypical, adult brains. Therefore, general application must be dealt with care. For example, elderly and young populations will, on average, show substantially more images with excessive motion. These images will go through FreeSurfer, and often produce an outlier EC, while a few will yield a perfectly standard EC. Typically, these cases with standard ECs are probably less accurate on the IDPs being analyzed, for example, if prior knowledge biased more the output for the hidden properties of this subject. In other words, in these cases, a researcher would be better off actually including the outliers.

This is an important point to raise. We agree with the reviewer that the Euler Characteristic is likely correlated with pathology in addition to data quality (e.g., due to movement artefacts) and this is important to consider when modeling clinical populations but also ensure high quality data. First, we point out that the inclusion threshold is mostly important for the estimation of the normative model, which in our work – like Rosen et al – is based on healthy control data. It is easy to repeat predictions for subsequent clinical samples using a more lenient inclusion threshold (or none at all) in cases where this consideration might be operative. Second, in an effort to strike the right balance, we have chosen the EC threshold quite conservatively in that it excludes subjects that are very far into the tail of the (rescaled and centered) EC histogram. This means that we are likely dropping only subjects with true topological defects. This is also an important motivation for the careful manual QC procedures we describe above. That said, we acknowledge that any heuristic is necessarily imperfect, which we acknowledge in the limitations section and in the methods.

• Title: "high precision" - it is unclear what precision this is qualifying as high. Is it spatial effectively granularity for a large number of ROIs being modeled or is it because the spread of the normative charts is narrow along the lifespan and as compared to some standard of variability.

We refer to spatial precision in terms of the granularity of the regions of interest that we estimate models for. We have revised the manuscript throughout to make this more explicit.

Gutes Projektmanagement

Thinking through the context: McWhorter is kind of hard to categorize in terms of political ideology. He claims he's a liberal, but is pretty comfortable criticizing leftist thinking that he disagrees with, and that's largely what he seems to be doing in this essay.

I agree that the conclusion the authors come to in the study McWhorter discusses is probably flawed--it's just a huge generalization (there are probably many reasons language and the ways we use it might change) and unless there's more to the study than JM is letting on, simply noting that there has been a change does not mean we can draw conclusions about what is at the root of the change. That seems more like an anthropological investigation.

Also, I would say there are contextual things to consider when determining why so much language is self-focused; the last forty years has seen a fairly large technological shift which gave many more people the tools to speak for themselves, and social media may have exacerbated this. That this wasn't considered (either by McWhorter or the authors of the study) seems a pretty big over sight.

Here, he basically sums up his position on why language is more self-focused than it used to be--it's just how we talk, it's not how we think

This is an example of how a person in a position of power opposes ethnic studies because they think that "it has a very toxic effect" that promotes victimhood.

I Like this. So it often doesn’t seem like there is a clear goal we are working towards. It seems so obvious but simply saying something like “we want to have 50% of course curriculum guides with at least one open educational resource (OER)” focuses on that. Everything else can be ignored outside of that goal. Whenever there are meetings, and they talk about what has been accomplished, it’s like a popcorn kernel popped in that goals were just met. For example, they recently moved all the student emails at my school to a different domain name. They made an announcement they did so after the fact like I was entirely in the dark for the whole process rather than being updated along the way.

It's just weird to me that the same word can mean different things in different parts of the world. It makes me more aware of how careful you have to be when you go to different places/

Meaning, you can come up with a solution to an NP problem in polynomic time (something represented by an exponential), if you take the optimal choice every time there is a choice (non-deterministic). Which is the same as verifying that a given solution is correct, by just following it's steps.

This passage is interesting. I think we need to discuss it further. I'm not quite sure I understand what he's getting at. It seems like they are looking at really old art (like egyptian paintings on a wall) and talking about how strange people and life was back then. Plato says "they're like us", referencing that we're just as clueless now as the people they are looking at. We're unable to see our own potential and future.

Before it stated that speech was not important and we know that to be a lie. Speech is very important it's the way humans communicate.

LETS FUCKING GOOOO

The Culture Industry: Mass Deception in Dialectic of Enlightenment

The culture industry is any industry that is producing cultural products: news, beauty, music, fashion industry has one goal: generate profit. They will produce things that are produced like a factory. They are intended to produce consumerism. In the way that sex sells, rebellion also sells. The industries that are making our culture are feeding us our news so are corporaterized. Adorno and Horkheimer would not be suprised about YouTube. That cultural adversary may be dialectical.

Academia. You know more about less and less.

Backdrop context: Shaky ground of liberal democracy in the beginnings of WW2,rise of social movements, rise of nationalism and facism.

Englightment: Reason & Individual liberty Bacon: a larger system that synthesized knowledge and power as one; a flipping of nature over man and man over nature; but A/H thought that this played out in a human global scale.

If Enlightenment was supposed to create logic and reason why did we experience WW2, for A/H if we're to take Bacon seriously we have to consider DOMINATION in newer notions of freedom.

Regression: Enlightenment as Myth. Englightenment becomes totalitarian it ABSTRACTS. Ex. Hitler youth, a difference among others yet they become homogenous sameness among each other.

The result of the sacrifice continues that is far more reaching that MArx's alienation. --> UNFETTERED ACCEPTANCE

Adorno/H say that positivism: 1) a system of philiospgy were every assertion can be proved 2) as ideoogy where eveyrthing is true by default and questioning it goes against objective foundations.

Kant saw a short-sighted view of self-reason

IDEOLOGY of defintiion #2 the process of Englightenment is brought into analysis of the Culture Industry.

Context: AMerica is becoming global powerhouse, Soviet underwent their own Industrial Revolution, change was everywhere and revolutionary change with one large outlier thus the immediat question. Despite a global population of workers revolitng around the globe then why didnt the United States or Western Countries embrace similar post-capitalist systems seen across teh globe? And how is Soviet existing as a post-capitalist system alongside the capitalist system?

Art is being systematized, newer technology is being synthesized into one and the same narrative. The same repackaged story; subject and authority. This mimics the governance of a few over many.

A/H utilize Kant's idea of schemtaism (being how the midn communicates with objects and other structures aorund us, how to reason and cogantate; synthesize and bring Froyd into the mix, how we suppress ourselves and our desires to how we fit into society. We cognitively pick up how the world is presented to us. Viewership creates the bounds within which we can do art. A psychological realism that is difficult to break for a person who works a 9-5pm; to break it in a capitalist system is used to labor and build wealth. Art becomes the same even when it tries to stand out. There is a unifornm aethetic if you wnat to be different, the sameness and constant

Art is now abstracted a fulfilmmnet of mere numbers rather than aesthetic work and utility. Regarding art in newer creation, A/H bring up autombiles as an example, a film must have a romatic sequence that the industry now demands. This is part of Mechanical reproduction. Art will become tailored to your class relation as well. But it's not only class and poleconomy but it's not just corporate art is entirely based oof profit but rather there is a cyclcial ideology that reinforces this ideology. see

The focus isn't on marvel movies (for their thrist for profit) or to tell consumers to not consume this art but its the industry itself that reproduces the

"Thirst" is not conscious it is a result of the structure that is capitalism the culture industry modling of our desires in the first place.

It's not good enough to tell individuals to not consume said tailored art. Example anti-semitism is a result of essentialist notions of race. Largest incubators of anti-semitism stem from the bourigeoise themselves but a ruling class of people attempting to hide a ruling class domination. Here the bouregoise know the struggles of individuals and pins this to Jewish individuals.

Thus its not really just pushing Jewish people out of certain borders this marks the downfall of bouregoise property.

Look up: > Cultural education became....

The whole world is made to pass through the filter of the culture industry...

Culture is a paradoxical commodity. It is so completely subject to the law of exchange that it is no longer exchanged; it is so blindly equated with use that it can no longer be used. For this reason it merges with the adver­tisement. The more meaningless the latter appears under monopoly, the more omnipotent culture becomes. (pg 131).

Unending sameness also governs the relationship to the past. What is new in the phase of mass culture compared to that of late liberalism is the exclusion of the new.

We live in a society, where those that need help have to fight tooth and nail to get it. In medicine an ambulance drive is $1700, older people who need long term care have to have slaved their whole lives just to get one person to do the bare minimum for them. This is in great contrast to other societies that do take care of the sick and elderly with the upmost of care because at the end of the day regardless of where a person is in life or in groups we are all people. It's sad that America does not remember that basic fact.