Le conseil d'école vote le règlement intérieur de l'école, établit le projet d’organisation pédagogique de la semaine scolaire, dans le cadre de l'élaboration du projet d'école à laquelle il est associé, donne tous avis et présente toutes suggestions sur le fonctionnement de l'école et sur toutes les questions intéressant la vie de l'école (par exemple, intégration des enfants handicapés, activités périscolaires, restauration scolaire, hygiène scolaire, sécurité des enfants, art. D. 411-2 du code de l’éducation).

Task 2 Briefly not the main topics throughout each section of the text. This section explains the differences in philosophy from other types of science. It shows how they collect knowledge and that there is not a definite answer to philosophical questions. Restate the main point of th opening paragraph in your own words. The opening paragraphs main point is that philosophy is the study of value. Many time it is undermined by other types of science that claim it is pointless. These people do not allow any kind of growth outside of said science. Estate the main point of the closing paragraph in your own words Philosophy’s goal is not to come to a conclusion but instead expand you thoughts. This is the knowledge that makes it possible to grow how we think. Everyone find and define two vocabulary words Unalloyed-complete and unreserved Dogmatism-the tendency to lay down principles as incontrovertibly true.

DOI: 10.1016/j.devcel.2021.07.021

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What is this?

DOI: 10.1016/j.devcel.2021.07.021

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Curator: @Naa003

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What is this?

DOI: 10.1016/j.devcel.2021.07.021

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Curator: @Naa003

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What is this?

DOI: 10.1016/j.devcel.2021.07.021

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What is this?

DOI: 10.1016/j.devcel.2021.07.021

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What is this?

DOI: 10.1016/j.devcel.2021.07.021

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Curator: @Naa003

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What is this?

DOI: 10.7554/eLife.66078

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Curator: @scibot

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What is this?

Hillus, David, Tatjana Schwarz, Pinkus Tober-Lau, Hana Hastor, Charlotte Thibeault, Stefanie Kasper, Elisa T. Helbig, et al. “Safety, Reactogenicity, and Immunogenicity of Homologous and Heterologous Prime-Boost Immunisation with ChAdOx1-NCoV19 and BNT162b2: A Prospective Cohort Study,” June 2, 2021. https://doi.org/10.1101/2021.05.19.21257334.

DOI: 10.7554/eLife.61733

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What is this?

DOI: 10.7554/eLife.61733

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Curator: @evieth

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What is this?

Nasreen, Sharifa, Hannah Chung, Siyi He, Kevin A. Brown, Jonathan B. Gubbay, Sarah A. Buchan, Deshayne B. Fell, et al. “Effectiveness of COVID-19 Vaccines against Variants of Concern in Ontario, Canada,” July 16, 2021. https://doi.org/10.1101/2021.06.28.21259420.

Prof. Devi Sridhar. “Rest of the World Watching Closely: ‘In Scotland, Estimated That 92.5% of Adults Would Have Tested Positive for Antibodies against SARS-CoV-2 on a Blood Test in the Week Beginning 12 July 2021’- Is This Enough to Dampen Transmission & Protect under 12s from Infection? Under 18s?” Tweet. @devisridhar (blog), August 4, 2021. https://twitter.com/devisridhar/status/1422852550957617157.

DOI: 10.7554/eLife.15797

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What is this?

DOI: 10.7554/eLife.44431

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What is this?

DOI: 10.7554/eLife.44431

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What is this?

DOI: 10.1016/j.cub.2018.11.059

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What is this?

DOI: 10.1002/cne.24001

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What is this?

Bixler, D. (2020). SARS-CoV-2–Associated Deaths Among Persons Aged 21 Years — United States, February 12–July 31, 2020. MMWR. Morbidity and Mortality Weekly Report, 69. https://doi.org/10.15585/mmwr.mm6937e4

I like the differentiation that Jared has made here on his homepage with categories for "fast" and "slow".

It's reminiscent of the system 1 (fast) and system2 (slow) ideas behind Kahneman and Tversky's work in behavioral economics. (See Thinking, Fast and Slow)

It's also interesting in light of this tweet which came up recently:

I very much miss the back and forth with blog posts responding to blog posts, a slow moving argument where we had time to think.

— Rachel Andrew (@rachelandrew) August 22, 2017

Because the Tweet was shared out of context several years later, someone (accidentally?) replied to it as if it were contemporaneous. When called out for not watching the date of the post, their reply was "you do slow web your way…" #

This gets one thinking. Perhaps it would help more people's contextual thinking if more sites specifically labeled their posts as fast and slow (or gave a 1-10 rating?). Sometimes the length of a response is an indicator of the thought put into it, thought not always as there's also the oft-quoted aphorism: "If I Had More Time, I Would Have Written a Shorter Letter".

The ease of use of the UI on Twitter seems to broadly make it a platform for "fast" posting which can often cause ruffled feathers, sour feelings, anger, and poor communication.

What if there were posting UIs (or micropub clients) that would hold onto your responses for a few hours, days, or even a week and then remind you about them after that time had past to see if they were still worth posting? This is a feature based on Abraham Lincoln's idea of a "hot letter" or angry letter, which he advised people to write often, but never send.

Where is the social media service for hot posts that save all your vituperation, but don't show them to anyone? Or which maybe posts them anonymously?

The opposite of some of this are the partially baked or even fully thought out posts that one hears about anecdotally, but which the authors say they felt weren't finish and thus didn't publish them. Wouldn't it be better to hit publish on these than those nasty quick replies? How can we create UI for this?

I saw a sitcom a few years ago where a girl admonished her friend (an oblivious boy) for liking really old Instagram posts of a girl he was interested in. She said that deep-liking old photos was an obvious and overt sign of flirting.

If this is the case then there's obviously a social standard of sorts for this, so why not hold your tongue in the meanwhile, and come up with something more thought out to send your digital love to someone instead of providing a (knee-)jerk reaction?

Of course now I can't help but think of the annotations I've been making in my copy of Lucretius' On the Nature of Things. Do you suppose that Lucretius knows I'm in love?

Lopez Bernal, Jamie, Nick Andrews, Charlotte Gower, Eileen Gallagher, Ruth Simmons, Simon Thelwall, Julia Stowe, et al. “Effectiveness of Covid-19 Vaccines against the B.1.617.2 (Delta) Variant.” New England Journal of Medicine 0, no. 0 (July 21, 2021): null. https://doi.org/10.1056/NEJMoa2108891.

DOI: 10.7554/eLife.37024

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What is this?

DOI: 10.7554/eLife.68755

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What is this?

Most murders are system 1-based and spur-of-the-moment.

System 2-based murders are even more deplorable because in most ethical systems it means the person actively spent time and planning to carry the murder out. The second category includes pre-meditated murder, murder-for-hire as well as all forms of capital punishment.

DOI: 10.1002/cne.24042

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What is this?

DOI: 10.1016/j.cell.2019.08.001

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What is this?

DOI: 10.1016/j.cell.2019.08.001

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What is this?

DOI: 10.1016/j.cell.2019.08.001

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Curator: @ethanbadger

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What is this?

PINHO, A. C. (2021, March 11). EMA recommends COVID-19 Vaccine Janssen for authorisation in the EU [Text]. European Medicines Agency. https://www.ema.europa.eu/en/news/ema-recommends-covid-19-vaccine-janssen-authorisation-eu

Staff, Fox News. “McEnany Encourages People to Get COVID Vaccine: It Reduces Your Risk of Transmitting the Virus.” Text.Article. Fox News. Fox News, July 21, 2021. https://www.foxnews.com/media/mcenany-encourages-people-to-get-covid-vaccine-variant.

Tada, Takuya, Hao Zhou, Marie I. Samanovic, Belinda M. Dcosta, Amber Cornelius, Mark J. Mulligan, and Nathaniel R. Landau. “Comparison of Neutralizing Antibody Titers Elicited by MRNA and Adenoviral Vector Vaccine against SARS-CoV-2 Variants.” BioRxiv, July 19, 2021, 2021.07.19.452771. https://doi.org/10.1101/2021.07.19.452771.

Barouch, Dan H., Kathryn E. Stephenson, Jerald Sadoff, Jingyou Yu, Aiquan Chang, Makda Gebre, Katherine McMahan, et al. “Durable Humoral and Cellular Immune Responses 8 Months after Ad26.COV2.S Vaccination.” New England Journal of Medicine 0, no. 0 (July 14, 2021): null. https://doi.org/10.1056/NEJMc2108829.

DOI: 10.1016/j.cell.2019.04.030

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What is this?

DOI: 10.1016/j.celrep.2018.06.117

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Curator: @evieth

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What is this?

DOI: 10.1016/j.celrep.2019.03.013

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Curator: @ethanbadger

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What is this?

DOI: 10.1016/j.celrep.2019.03.013

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Curator: @ethanbadger

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What is this?

DOI: 10.1016/j.celrep.2019.03.013

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Curator: @ethanbadger

SciCrunch record: RRID:ZFIN_ZDB-ALT-110721-2

What is this?

DOI: 10.1242/dev.105718

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Curator: @mpairish

SciCrunch record: RRID:ZFIN_ZDB-ALT-061204-2

What is this?

DOI: 10.1016/j.stemcr.2015.02.006

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Curator: @mpairish

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What is this?

DOI: 10.1371/journal.pgen.1005934

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What is this?

DOI: 10.1016/j.cub.2017.11.018

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Curator: @evieth

SciCrunch record: RRID:ZFIN_ZDB-ALT-170509-2

What is this?

DOI: 10.1016/j.cub.2017.11.018

Resource: (ZFIN Cat# ZDB-ALT-131122-2,RRID:ZFIN_ZDB-ALT-131122-2)

Curator: @evieth

SciCrunch record: RRID:ZFIN_ZDB-ALT-131122-2

What is this?

DOI: 10.1016/j.neuron.2017.06.001

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Curator: @Zeljana_Babic

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What is this?

DOI: 10.1016/j.neuron.2017.06.001

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Curator: @Zeljana_Babic

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What is this?

DOI: 10.1002/cne.23746

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Curator: @bandrow

SciCrunch record: RRID:ZFIN_ZDB-GENO-030619-2

Curator comments: This one does not resolve but should, will check data

What is this?

DOI: 10.1016/j.neuron.2019.05.038

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Curator: @ethanbadger

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What is this?

DOI: 10.7554/eLife.66596

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Curator: @evieth

SciCrunch record: RRID:ZFIN_ZDB-ALT-140424-2

What is this?

DOI: 10.7554/eLife.66596

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Curator: @evieth

SciCrunch record: RRID:ZFIN_ZDB-ALT-070118-2

What is this?

Prunas, Ottavia, Joshua L. Warren, Forrest W. Crawford, Sivan Gazit, Tal Patalon, Daniel M. Weinberger, and Virginia E. Pitzer. “Vaccination with BNT162b2 Reduces Transmission of SARS-CoV-2 to Household Contacts in Israel.” MedRxiv, July 16, 2021, 2021.07.13.21260393. https://doi.org/10.1101/2021.07.13.21260393.

Luoto, Severi, Marjorie L. Prokosch, Marco Antonio Correa Varella, Indrikis Krams, and Corey L. Fincher. “Editorial: Coronavirus Disease (COVID-19) and Its Psychobehavioral Consequences.” Frontiers in Psychology 0 (2021). https://doi.org/10.3389/fpsyg.2021.723282.

This website is what I kept picturing when I heard the words "digital humanities project". This is like most of the websites I have interacted with, with posts on the main page and categorized tabs at the top for anything else you are looking for. I think I could probably branch out and explore different ones now that I know the wide variety out there.

I think that this idea for a digital project is really interesting. I like that they have mapped it out and you can click on a point to filter by state or establishment type to zero in on what interests you. I never thought that a digital humanities project could look like this. I think it is really cool how different all of these projects are on this site, yet they all fall under the umbrella of digital humanities.

I am usually that person who likes to know what they're getting into before they start. I like to know what skills I need as well so I can see if I will be able to do it easily or if it will be more of a learning curve. I definitely did not know anything about digital humanities before I started this class, but I am learning with each exercise we do. I think a lot of people like to be confident in what they are doing so to have some idea of what they are going to do and what skills they need is reassuring.

Blogging is a convenient way to get information out to an audience. Rather than letter-writing or phone, and then into email and chat rooms, blogging allows you to share information and knowledge by posting your thoughts. It can stay up for as long as you like, allowing a variety of people to view it. I think it is a way to share your ideas and get information out faster.

I do not think the term homo faber, or human as maker has become meaningless because of the digital world of the web and 3D printing. The digital is using that term in a different way. Yes, digital things are done online with the help of certain tools and software, but it is still the human behind the screen. It is the human as maker with the ideas and creativity for these new digital concepts and the ability and knowledge to develop them. People can use the digital to enhance their ideas.

I think it is true that for the most part the environments of the humanities have been things like the scholar's desk, lecture halls, campuses, and convention halls. In the last little bit there has been a shift from these environments in that the digital has now come in to play. I agree that the digital tools carry on the same functions as traditional humanities in comparable ways. We are still learning about the humanities by using a digital form, it is just a newer way of presenting them.

DOI: 10.7554/eLife.54894

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What is this?

Antonova, Elena, Karoly Schlosser, Rakesh Pandey, and Veena Kumari. “Coping With COVID-19: Mindfulness-Based Approaches for Mitigating Mental Health Crisis.” Frontiers in Psychiatry 12 (2021). https://doi.org/10.3389/fpsyt.2021.563417.

https://assets.publishing.service.gov.uk/government/uploads/system/uploads/attachment_data/file/991135/3_June_2021_Risk_assessment_for_SARS-CoV-2_variant_DELTA.pdf. (2021). Retrieved 5 July 2021, from https://t.co/u3sOVk65Zi?amp=1

(2021). Retrieved 5 July 2021, from https://assets.publishing.service.gov.uk/government/uploads/system/uploads/attachment_data/file/991343/Variants_of_Concern_VOC_Technical_Briefing_14.pdf

(2021). Retrieved 2 July 2021, from https://assets.publishing.service.gov.uk/government/uploads/system/uploads/attachment_data/file/988458/Weekly_Flu_and_COVID-19_report_w20_v2.pdf

(2021). Retrieved 2 July 2021, from https://assets.publishing.service.gov.uk/government/uploads/system/uploads/attachment_data/file/988458/Weekly_Flu_and_COVID-19_report_w20_v2.pdf

DOI: 10.7554/eLife.63355

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Curator: @scibot

SciCrunch record: RRID:ZFIN_ZDB-ALT-070629-2

What is this?

Testing out the annotate feature. Student 1 will highlight sections according to the prompts, as shown HERE.

For example: "This is me during interviews. I say too much and veer off topic."

Singh, Urvashi B., Mercy Rophina, Dr Rama Chaudhry, Vigneshwar Senthivel, Kiran Bala, Rahul C. Bhoyar, Bani Jolly, et al. “Variants of Concern Responsible for SARS-CoV-2 Vaccine Breakthrough Infections from India.” OSF Preprints, June 3, 2021. https://doi.org/10.31219/osf.io/fgd4x.

Naveca, Felipe Gomes, Valdinete Nascimento, Victor Costa de Souza, André de Lima Corado, Fernanda Nascimento, George Silva, Ágatha Costa, et al. “COVID-19 in Amazonas, Brazil, Was Driven by the Persistence of Endemic Lineages and P.1 Emergence.” Nature Medicine, May 25, 2021, 1–9. https://doi.org/10.1038/s41591-021-01378-7.

DOI: 10.1016/j.cub.2021.05.042

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Curator: @Naa003

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What is this?

DOI: 10.1016/j.cub.2021.05.042

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Curator: @Naa003

SciCrunch record: RRID:ZFIN_ZDB-ALT-150324-2

What is this?

DOI: 10.1016/j.celrep.2021.109255

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Curator: @Naa003

SciCrunch record: RRID:ZFIN_ZDB-ALT-050428-2

What is this?

Cirillo, Nicola, and Richard Doan. “Bell’s Palsy and SARS-CoV-2 Vaccines—an Unfolding Story.” The Lancet Infectious Diseases 0, no. 0 (June 7, 2021). https://doi.org/10.1016/S1473-3099(21)00273-5.

Liu, Jianying, Yang Liu, Hongjie Xia, Jing Zou, Scott C. Weaver, Kena A. Swanson, Hui Cai, et al. “BNT162b2-Elicited Neutralization of B.1.617 and Other SARS-CoV-2 Variants.” Nature, June 10, 2021, 1–5. https://doi.org/10.1038/s41586-021-03693-y.

Javid, B., Bassler, D., Bryant, M. B., Cevik, M., Tufekci, Z., & Baral, S. (2021). Should masks be worn outdoors? BMJ, 373, n1036. https://doi.org/10.1136/bmj.n1036

Richterman, A., Meyerowitz, E. A., & Cevik, M. (2021). Indirect Protection by Reducing Transmission: Ending the Pandemic with SARS-CoV-2 Vaccination. Open Forum Infectious Diseases, ofab259. https://doi.org/10.1093/ofid/ofab259

DOI: 10.7554/eLife.62196

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Curator: @scibot

SciCrunch record: RRID:ZFIN_ZDB-ALT-120117-2

What is this?

Michaelsen, Thomas Y., Marc Bennedbæk, Lasse E. Christiansen, Mia S. F. Jørgensen, Camilla H. Møller, Emil A. Sørensen, Simon Knutsson, et al. “Introduction and Transmission of SARS-CoV-2 B.1.1.7 in Denmark.” MedRxiv, June 7, 2021, 2021.06.04.21258333. https://doi.org/10.1101/2021.06.04.21258333.

Daniels, Curt J., Saurabh Rajpal, Joel T. Greenshields, Geoffrey L. Rosenthal, Eugene H. Chung, Michael Terrin, Jean Jeudy, et al. “Prevalence of Clinical and Subclinical Myocarditis in Competitive Athletes With Recent SARS-CoV-2 Infection: Results From the Big Ten COVID-19 Cardiac Registry.” JAMA Cardiology, May 27, 2021. https://doi.org/10.1001/jamacardio.2021.2065.

Reviewer #2 (Public Review):

This study presents a novel machine learning tool (termed T-REX) for automated analysis of single cell cytometric data that is capable of identifying rare cell populations, such as antigen-specific T cells. This ability to detect low frequency cells is a distinct advantage over existing tools. The demonstration of this ability is appropriately shown by examining antigen-specific CD4+ T cells before and after rhinovirus infection in a challenge study. Useful demonstrations are also included for examining SARS-CoV-2-specific T cells and changes in cellular populations in cancer patients upon treatment. These examples use both mass cytometry and fluorescence-based cytometry. Since both of these are commonly-used single-cell technologies that generate highly complex data sets, new automated analysis methods such as T-REX are needed.

The first data set examined changes in cell phenotype before and 7 days after rhinovirus infection in healthy adults. The flow cytometric staining panel included markers of T cell differentiation and activation as well as rhinovirus-specific tetramers. The results of T-REX convincingly demonstrate "hotspots" that are expanded at 7 days and enriched for tetramer-staining cells. Thus, this study succeeds at demonstrating the utility of this method for identification of rare cells and the authors use this data set to appropriately determine the model parameters. Combining the results of this algorithm with the "Marker Enrichment Modeling (MEM)" method to characterize the markers expressed on those cell populations identified through T-REX is also very informative since this automates the characterization (that traditionally needs to be done by manual investigation).

This first data set is relevant for this demonstration, but in some aspects it represents a best case scenario. "Phenotypic" identification of antigen-specific T cells in this way is only possible because the time point was chosen to capture the relatively narrow window when T cells would be activated, and there was access to a baseline sample for comparison. The authors do address the second point, and perform the analysis comparing day 7 to a later time point, day 28, as an appropriate alternative. The first concern limits the generalizability of this approach. In fact, the second example dataset examining mass cytometry data in patients with COVID-19 does in fact demonstrate limited ability to detect change in cell populations for many study participants.

Reviewer #2 (Public Review):

The use of convalescent plasma (CCP) to treat patients with Covid-19 has changed over the course of the pandemic (from rates as high as 40% of hospitalized patients in October, 2020 to a low of less than 10% by March 2021). To explore the efficacy of CCP therapy and the impact of the drop in CCP use, the authors assess whether there was a link between CCP use and patient mortality rates over time in the U.S. Using information from blood centers to estimate CCP usage and population level information on deaths from public databases, they found a strong inverse correlation between CCP usage per hospital admission and deaths due to Covid-19 after admission. The model estimates that the case fatality rate decreased by 1.8 percentage points for every 10 percentage point increase in the rate of CCP use. The detailed analysis suggests that the observed effect could not be attributed to changes in patient ages over time or the emergence of variant viruses. Other cofounders such as changes in the use of additional therapeutic agents or clinical interventions were not analyzed. The authors acknowledge the main limitation of this type of analysis i.e. that establishing a correlation does not prove a causal role. With that caveat, they conclude that the decline in usage may have resulted in excess deaths, possibly 29,000 to 36,000 over the past year in the U.S. Because the decreased usage of CCP occurred during the time that several randomized clinical trials and some media coverage reported no benefit of CCP, the authors suggest that resultant "plasma hesitancy" may have contributed to increased mortality. These findings add an important perspective to future considerations for clinical care, treatment guidelines and regulatory approvals of CCP. Emphasizing the importance of using high-titer units and administering CCP early in the disease course, the authors urge a more nuanced interpretation of the available evidence and a holistic approach to decisions about the use of CCP in individual patients.

Shapoorji Pallonji Joyville Sector 102 Gurgaon is a premium residential project in close proximity to Delhi. Contemporary construction, state-of-the-art amenities, top-notch security, and huge green expanse make the project highly demanding. Nestled right close to major establishments like hospitals, schools, malls, and offices.

Reviewer #2 (Public Review):

The authors should be commended on the sharing of their data, the extensive experimental work, the experimental design that allows them to get opposite predictions for both hypotheses, and the detailed of analyses of their results. Yet, the interpretation of the results should be more cautious as some aspects of the experimental design offer some limitations. A thorough sensitivity analysis is missing from experiment 2 as the safety margin seems to be critical to distinguish between both hypotheses. Finally, the readability of the paper could also be improved by limiting the use of abbreviations and motivate some of the analyses further.

Major:

1) The text is difficult to read. This is partially due to the fact that the authors used many abbreviations (MA, PO, IMD). I would get rid of those as much as possible. Sometimes, having informative labels could also help FFcentral and FFlateral would be better than FFA and FFB.

2) The most difficult section to follow is the one at the end of the result sections where Fig.5 is discussed. This section consists of a series of complicated analyses that are weakly motivated and explained. This section (starting on line 506) appears important to me but is extremely difficult to follow. I believe that it is important as it shows that, at the individual level, PO is also superior to MA to predict the behavior but it is poorly written and even the corresponding panels are difficult to understand as points are superimposed on each other (5b and e). In this section, the authors mention correcting for Mu1b and correcting for Sig2i/Sig1Ai but I don't know what such correction means. Furthermore, the authors used some further analyses (Eq. 3 and 4) without providing any graphical support to follow their arguments. The link between these two equations is also unclear. Why did the authors used these equations on the pooled datasets from 2a and 2b ? Is this really valid ? It is also unclear why Mu1Ai can be written as the product of R1Ai and Sig1Ai. Where does this come from ?

3) In experiment 1, does the presence of a central target not cue the participants to plan a first movement towards the center while such a central target was never present in other motor averaging experiment. In the adaptation domain, people complain that asking where people are aiming would induce a larger explicit component. Similarly, one could wonder whether training the participants to a middle target would not induce a bias towards that target under uncertainty

4) The predictions linked to experiment 2 are highly dependent on the amount of safety margin that is considered. While the authors mention these limitations in their paper, I think that it is not presented with enough details. For instance, I would like to see a figure similar to Fig.4B when the safety margin is varied.

The sensitivity analysis is very difficult to follow and does not provide the right information. First, this is only done for exp2 and not exp1. For exp1, it would be good to demonstrate that, even when varying the weight of the two one-target profiles for motor averaging, one never gets a prediction that is close to what is observed. It is unclear in the text that the performance optimization prediction simply consists of the force-profile for the center target. The authors should motivate this choice. For the second experiment 2, the authors do not present a systematic sensitivity analysis. Fig. 5a and d is a good first step but they should also fit the data on exp2b and see how this could explain the behavior in exp 2a. Second, the authors should present the results of the sensitivity analysis like they did for the main predictions in Fig.4b. While I understand where the computation of the safety margin in eq.2 comes from, reducing the safety margin would make the predictions linked to the performance optimization look more and more towards the motor averaging predictions. How bad becomes the fit of the data then ? How does the predictions look like if the motor costs are unbalanced (66 vs. 33%, 50 vs. 50% (current prediction), 33 vs. 66% ). What if, in Eq.2 the slope of the relationship was twice larger, twice smaller, etc. The safety margin is the crucial element here. If it gets smaller and smaller, the PO prediction would look more and more like the MA predictions. This needs to be discussed in details. I also have the impression that the safety margin measured in exp 2a (single target trials) could be used for the PO predictions as they are both on the right side of the obstacle.

5) On several occasions (e.g. line 131), the authors mention that their result prove that humans form a single motor plan. They don't have any evidence for this specific aspect as they can only see the plan that is expressed. They can prove that the latter is linked to performance optimization and not to the motor averaging one. But the absence of motor averaging does not preclude the existence of other motor plans.... Line 325 is the right interpretation.

6) Line 228: the authors mention that there is no difference in adaptation between training and test periods but this does not seem to be true for the central target. How does that affect the interpretation of the 2-target trials data ? Would that explain the remaining small discrepancy between the refined PO prediction and the data (Fig.2f) ?

Reviewer #2 (Public Review):

The authors established a comprehensive map of neurogenetic sites with evolutionary conserved neurogenic and postmitotic gene expression in a common octopus, Octopus vulgaris that has been a historically important species in comparative neuroscience and behavioral studies. The selected molecules include representative regulatory genes such as achaete-scute, neurogenin, and neuroD, and also proliferating cell markers such as elav and PCNA. In subsequent experiments by using a fluorescent dye, the authors carefully traced the migratory pathways from the target ectodermal sites surrounding eyes to many developing brain lobes of clearing staged embryos with light sheet microscopy for 3D reconstruction.

They found that the special regions called lateral lip and other special ectodermal areas produced a pool of migratory postmitotic neurons that might contribute a novelty for developing octopus large and complex brains as in mammals, in contrast to those of other invertebrates such as flies or worms.

I find the bodies of evidence convincing. A good study usually opens many new questions. Before publication, I found two major points that may enhance the author's conclusions.

1) Are the migratory cells only neurons, or could they also be glia, neurosecretory, blood, or immune cells? The enlarged views of the migratory cells and the cytological features must be clarified.

2) The expression of canonical neurogenic genes disappears during middle and late stages, meaning that octopus has very unique neurogenic mechanisms compared to mammals? Consider the octopus-specific novelty.

Reviewer #2 (Public Review):

In this work, Corbett and colleagues investigate how value influences speeded decisions. In a random dot motion task with speed pressure, shortly before motion onset it is indicated which of both choices has a higher value if answered correctly. EEG recordings show a buildup of motor beta in response to the cue (earlier for high value choices, steeper for low value choices) and a dip in LRPs for low value choices in response to stimulus onset. A computational model constructed based on these findings provides a good account of the data. The EEG informed modeling is impressive and deserves merit. The paper is well-written, but rather dense.

• I am struggling with the idea that cue-evoked motor beta reflects urgency. As it currently reads, this is more taken as a given than actually demonstrated. Could this claim be corroborated by e.g. showing that response deadlines modulate this signal? Related to that, how can we be sure that the pre-stimulus patterns seen in motor beta feed into the decision making process itself? It is not hard to imagine why left and right pointing arrows directly trigger motor activity (i.e. simple priming), but does that also imply that such activity leaks into the decision process?

• I had a hard time understanding the choice for this specific design. As the authors write they "primarily focused on the value biasing dynamics in common across these challenging regimes" so I wonder whether conditions with different value differences could have been more instructive (e.g., according to the author's hypothesis different levels of value should parametrically affect motor beta, whereas if this reflect a simple priming process value itself should not matter). Alternatively, it should be better explained why these conditions where crucial for the current findings.

• One of the main selling points of the paper is that we currently lack a model that can explain fast value-based decisions, mostly because the constant drift rate assumption in evidence accumulation models seems invalid. This conjecture is very similar to literature on response conflict, where performance in conflict tasks (such as Stroop, Flanker, etc.) is best modelled using a time-varying drift rate. I wonder to what extent current data reflect the same process, i.e. the value cue "primes" a response, which then has to be suppressed in favor of the correct response. A clear difference is that the value remains relevant here, but could e.g. the motor beta effect just reflect priming?

• If I understand correctly the model was fit to all data effectively ignoring between-participant differences. It is unclear why this was this done (rather than fitting data separately per participant or fitting the data using a hierarchical model), because it induces substantial variance in the fits caused by between-participant differences.

Reviewer #2 (Public Review):

In this work, Sobczak et al. suggest that correlations between fMRI and pupil diameter vary over time, and propose an approach to identify distinct clusters of such correlation patterns. The proposed methods are applied to data acquired from anesthetized rats. Based on the clusters obtained, the authors conclude that pupil dynamics are linked with different neuromodulatory centers over different intervals of time.

Overall, I believe that the study is novel and uncovers potential new modes of coupling between neuromodulatory nuclei and pupil diameter. However, additional analysis may be needed to fully support the validity of the derived clusters, and the decoding methods may need some modification before the accuracy values can be properly interpreted. The mechanisms behind the time-varying fMRI-pupil coupling exhibited under anesthesia could also be further clarified. Specifically:

• The clusters appear to involve interpretable brain regions. However, a more formal analysis of reproducibility of these clusters, and statistical testing against an appropriate null model, are not present. Such tests would be useful for establishing the validity of the derived clusters, ensuring that the conclusions are strongly supported. Similarly, the differentiation between power spectral density of each cluster is not yet supported by statistical testing.

• With regard to the decoding models, it appears there could be interdependence between the training and testing data (the PCA step seems to include all scans, and it was not clear if the training/testing sets contained data drawn from the same animal).

• While the paper is motivated by discussion that pupil diameter changes are complex and related to rich behaviors (mental effort, decision making, etc.), this paper examines data from anesthetized rats. The mechanisms behind the time-varying changes in fMRI-pupil coupling in the current data, and the potential impact of anesthesia, were not clear and could be elaborated upon.

Reviewer #2 (Public Review):

The various pathogenic, parasitic, symbiotic, and mutualistic interactions between insects and the microbes they interact with represents a rich area of research. This study by Xiao et al. represents a very interesting example of such a relationship. Overall the study is well designed and executed. The approach they utilize to test their hypothesis is valid and they combined both laboratory and field collected insects to address the question. The RNAseq analysis also provides potential insights into possible mechanisms by which the virus HaDV2 enables enhanced resistance to Bt Cry1Ac. The RNAseq data also represent one of the minor issues. The authors focused on analyzing only development and immune systems, however, they do not report on any other significantly different changes in gene expression other than reporting that there were 1573 significant differences. The authors should at least provide some holistic analysis and report the data in the supplemental results. Focusing on development and immune systems is valid and rationally supported but a complete analysis should be presented. The relationship between H. armigera and HaDV2 is more a mutualistic relationship, thus, the authors should consider changing the titles of the manuscript and the supplementary data. This is an exciting study and is well written and will be of general interest to the field.

Reviewer #2 (Public Review):

Transposable elements (TEs) have been shown to play an important role in genome evolution, in shaping genomic organization, structure, and genome size. The importance of TEs in evolutionary processes such as adaptation, has been rather limited so far. Here, Oggenfuss et al. used intraspecies data from six global populations of the wheat pathogen Zymoseptoria tritici to study the process of TE insertion dynamics, to detect candidate adaptive regions associated with TE insertions, and to show that TE expansion has driven to genome size of some populations in about 25 years. Using publicly available short read sequencing, as well as newly sequenced populations, they created a pipeline to specifically identify TE insertions, then used TE frequency insertions to infer patterns of selection, which they hypothesize is under strong purifying selection for regions into genes. They further tried to detect evidence for positive selection at loci associated with adaptation to new environment or resistance to fungicide, and finally contrasted the TE expansion in a window of 25 years to show that two populations have increased their genome size due to TE expansion.

Strengths:

The dataset used in this study with different global populations at different time makes the authors in an ideal position to detect TE expansion in a short timeframe of 25 years. While it has been shown in multiple eukaryotic genomes that TEs contribute substantially to variation in genome size, the evidence provided in this paper is compelling and shows that it is unlikely due to genome duplication events.

The pipeline to identify TE insertions from short read sequencing provides a well detailed path to apply in other genomes, and provide logical reasoning detecting TEs absent from the reference or absent from the isolates but present in the reference. Providing validation for some of the steps would be desired if well-known regions can be used.

The authors explored the interesting perspective of identifying TEs under positive selection by focusing on loci with increased frequency in specific populations. They also added a level of functional validation to better understand specific TE insertions under selection, which is often not included. They identified three loci that did confer resistance to fungicide according to their assay. These results would benefit from additional key details in the methods and the rationale behind the choice of loci in order to fully measure the impact of this finding.

Weaknesses:

While the paper does have strengths in principle to study the evolution of TE dynamics, the weaknesses of the paper reside in the fact that the manuscript in its current form does not directly support the key conclusions presented here. Additional analyses would be required to support them. Such as :

The presence of a large percentage insertions being singleton TEs coupled with low frequency (based on an arbitrary cutoff) is used to conclude strong purifying selection is acting on these new insertions. The authors should have included evidence to convince the reader the presence of a TE in one isolate is not a case of false positive. Additional analyses such as a subset of the singleton TEs to corroborate these single loci or plotting the relationship between how many isolate with one insertion were found and read depth could be informative. Also, understanding the singleton in the context of chromosome locations (core and accessory) could help reinforce the evidence of purifying selection.

The author's conclusion of relaxed selection in accessory chromosomes and between populations are mainly based on TE density. This conclusion may be better supported by adding quantified information of the relationship between the numbers and the type of TE insertions and genomic features such as recombination, which is often found to be negatively associated with TE content. Showing the recombination rate between the chromosomes (core, accessory) would help strengthen the argument that selection acts more strongly in regions of high recombination. To make their conclusion more robust, the authors could have included information about the recombination landscape.

Overall, the authors should have provided the adequate statistical analyses to support many of the insertion frequencies that are often only mentioned qualitatively (e.g. "less than expected") without having the quantitative test. Also, the contrast between low TE insertion frequencies versus high frequency is used without providing details about what is expected, either supported by the literature or by more detailed analyses. Arbitrary threshold can be prone to give artifact results.

Context : This article comes with a number of recent papers exploiting the population genomics dataset of the major wheat pathogen Zymoseptoria tritici (Fouché et al 2020, Krishnan et al. 2018) to show that TE-mediated insertions have in part helped to colonize host plants and tolerate environmental stress. Here, the authors build upon this knowledge to attempt at characterizing the process of TE insertion dynamics, detect signatures of adaptive evolution and changes in genome size at the population level.

Reviewer #2 (Public Review):

The current study nicely demonstrates that high-order assembly of SMN protein oligomerization is necessary for animal survival and is dependent on a motif exposed to YG zipper dimers. Mutations in the human SMN1 gene have been shown to cause a neurodegenerative disease named Spinal Muscular Atrophy (SMA). About 50% of the SMA-causing mutations are located in the YG zipper domain. The authors used multi-disciplinary approaches such as biophysical, bioinformatic, computational and genetic approaches to demonstrate that a set of YG box amino acids in SMN protein are not involved in dimerization process and formation high-order oligomers is dependent on these residues. Importantly, mutating key residues within this new structural domain impairs SMN dimerization and causes motor dysfunction as well as viability defects in Drosophila. Overall, this is a well-written paper that offers new insights into the structural and functional aspects of SMN protein. The authors should consider addressing the following issues:

1) The authors should discuss the impact of the YG zipper domain mutations on snRNP biogenesis. SMN protein is a master regulator of snRNP biogenesis. It is a little surprising that the authors did not mention snRNP biogenesis in the whole manuscript.

2) The authors should provide evidence that their transgenic lines express the desired transgene. A WB or qPCR would be great (even as supplementary data).

3) Page 12: The authors stated "Both missense mutations display early onset SMA-like phenotypes". Was it age-dependent phenotype? Did adult animals show a more severe motor dysfunction?

4) There are few statements that the authors should consider making clear. Here is an example "Presumably, the structural changes associated with Cys and Val substitutions do interfere with some aspect of SMN biology, leading to the intermediate and severe SMA phenotypes observed". What do you mean by some aspects? Oligomerization, stability or anything else?

5) There are few typos throughout the manuscript that the authors should correct (western should be written as Western).

Reviewer #2 (Public Review):

The article by Breska and Ivry provides a nice, timely, and relevant continuation of their previous recent work on the role of the cerebellum in interval-based (but not rhythm-based) anticipation in time. While in their related prior work (in particular their recent articles in PNAS and Science Advances) the authors used simple reaction time tasks that made it difficult to attribute the observed effects to visual vs. motor anticipatory mechanisms, in the current work they used a perceptual discrimination task with a delayed response to focus on potential contributions of the cerebellum to temporal anticipation specifically for perceptual sensitivity (where the role of the cerebellum is less obvious, given it has traditionally been implicated more in motor control than in perception). They do so by comparing individuals with cerebellar degeneration to controls, and finding a selective impairment of the individuals with cerebellar degeneration to use interval-based temporal predictions to facilitate visual discrimination, while rhythm-based performance benefits are spared (providing a neat comparison and control).

I have no major comments to detail. The short report is well written, complements related work by the authors nicely, and makes an important and novel contribution to the literature on temporal anticipation (while also having relevant implications more generally for views on the role of the cerebellum in cognition).

Reviewer #2 (Public Review):

The manuscript describes a tool to independently tune mean protein expression levels and noise. Light induces dimerization and subsequent activation of transcriptional activator GAVPO. By introducing 5xUAS (a target sequence for dimerized GAVPO) upstream a mRuby reporter gene, the effect of light can be measured on mRuby mean and noise.

By pulsing light at different periods (from 100-400 minutes), the authors reduce the mRuby noise for intermediate average light intensities. Notably, the pulses are all applied at an absolute light intensity of 100 uW/cm2, with the average light intensity being modulated through the light-off time-periods. Therefore, as all periods tend towards 100 uW/cm2 average light intensity, the PWM duty cycles becomes more similar to the 100 uW/cm2 AM case.

Strengths:

The proposed method is an elegant way to independently tune protein mean and noise. This would have a broad application in the field and is much needed to be able to study the consequence of protein expression noise, independently of mean. In addition, the authors use multiple powerful single-cell techniques to try and determine the mechanism underpinning the light-induced noise modulation.

During constant exposure to light, increased light intensity increases the mean expression of mRuby, while decreasing the noise. This high noise is mostly due to observed bimodality in mRuby expression. Through ODEs and by using small molecule inhibitors, the authors show that this bimodality is caused by some cells being stably off, while other cells enter an on state. In this on state a positive feedback can occur where initial binding of dimerized GAVPO induces histone acetylation and chromatin accessibility, and thus stimulates further GAVPO binding. Bistability induced by constant light exposure is disrupted using small molecule inhibitors of CBP/p300 HAT activity, indicating that histone regulation is a cause for this observed bistability. The stable on state is demonstrated to be more active and accessible through ChIP-seq and ATAC-seq respectively.

Weakness:

The single-cell ATAC-seq data indicate that pulsing light induces switching from an accessible (light on) to inaccessible (light off) chromatin state. The authors argue that the switching back into a chromatin inaccessible state prevents the positive feedback to occur and thus reduces noise. However, there are weaknesses in the description of the mechanism by which the pulses modulate (i.e., reduce) noise. Overall, since these sections in the manuscript are not easy to understand, it is difficult to parse what mechanism the authors attributed to the observed noise reduction and to assess if the data supports the conclusions.

The data from the single-mRNA live-cell imaging experiments are somewhat ambiguous and do not necessarily support some of the arguments. The conclusion that transcription, nuclear export, and mRNA degradation flatten the pulsatile chromatin caused by the PWM is not clear from the data. Especially, since most cells do not show any pulsatile behavior both in the single-cell ATAC-seq and the live-cell imaging data.

Reviewer #2 (Public Review):

Motivated behaviors, such as food seeking when hungry, can also occur spontaneously at irregular intervals. Understanding how this irregular expression arises is important for understanding behavior and is relatively little investigated. The present work thus addresses an important and under-investigated area in neurobiology. Its demonstration of a potential cellular mechanism for irregular behavioral production has wide relevance, ranging from how cells make "decisions" to how whole organisms do so.

Intact Aplysia occasionally produce bites even in the absence of food, and isolated buccal ganglia (which contain the biting central generator circuit) will occasionally spontaneously produce fictive bite motor patterns. The activity of central pattern generator networks has almost exclusively been ascribed to the actions of the voltage-gated channels in the network neuron cell membranes and the synaptic connectivity among the network's neurons. Bédécarrats et al. show that a small, highly regular cell membrane voltage oscillation occurs in a neuron (B63) in the biting neural network, and that occasionally this oscillation becomes large enough to trigger a plateau potential in B63 and a single fictive bite from the entire circuit. They show that this oscillation is not due to cell membrane voltage dependent conductances, but instead from process involving the endoplasmic reticulum, mitochondria, or both. Although organelle-driven changes in cellular or tissue activity have been observed in other cell types, this is, to my knowledge, its first observation in a neural network. These data thus are potentially of great importance in understanding how neural networks function, most of which do not show the great regularity of central pattern generated behaviors.

The presented data seem, to me, strong with respect to the small potential oscillations not being generated by voltage-dependent cell membrane conductances, and somehow involving the intracellular organelles. What is less clear to me is how local release of Ca from endoplasmic reticulum or mitochondria would result in changes in ion composition under the cell membrane, which is what gives rise to the cell membrane potential. Ca is highly buffered in the cytoplasm. It is thus unclear to me that free Ca would remain so for any length of time after release. It does, of course, in muscles, but these are evolved for this to occur. The authors themselves raise a variant of these concerns in the Discussion when considering how the B63 cell membrane voltage oscillations are transmitted to neurons electrically coupled to B63, invoking as a possibility Ca activation of second messengers, which would then themselves be responsible for the cell to cell communication. It seems to me that the same concerns arise with respect to how Ca release at sites distant from the cell membrane could charge the membrane's capacitance.

A second remarkable observation is that B63 depolarization and firing does not reset the organelle-derived slow oscillation. B63 firing should result in substantial Ca concentration changes, at least in a shell under the cell membrane, so a possible feedback mechanism can be imagined. Most biological processes contain multiple feedback process that link cause and effect (e.g., the sequential current activations that return a cell to rest after an action potential, the interactions between sympathetic and parasympathetic system activity that maintain functionally proper body activation, the interactions that regulate hormonal levels). One possibility the authors mention is that the organelle-derived oscillation is used only for intermittent bite activities, and in feeding bites are instead generated solely by standard cell-membrane voltage-dependent processes. Regardless, it is a striking observation that merits additional investigation.

These issues, however, do not change the data, which show a clear association of disruption of endoplasmic reticulum and mitochondrial function and cessation of the cell membrane voltage oscillation. Nor is it reasonable to expect an article like this, showing an organelle-driven cell membrane potential oscillation for the first time in a neuron, to describe every aspect of the mechanism by which it occurs. Indeed, it is a measure of the article's interest that it prompts such thinking. It will be very interesting to see the effects of similar organelle-disrupting treatment on the activity of other well-defined neural networks.

Reviewer #2 (Public Review):

Wellington Miranda et al. investigated how acyl-homoserine lactone autoinducer mediated quorum sensing systems evolve. The authors used the statistical covariation method GREMLIN to identify key amino acids that have coevolved in the well-studied LasI/LasR quorum sensing AHL synthase/receptor pair from Pseudomonas aeruginosa PAO1. LasI produces and LasR detects 3OC12-HSL. The authors identify some new and some previously reported residues using the GREMLIN tool; they focus on L157 in LasI and G38, R61, A127, S129, and L130 in LasR as residues that determine selectivity of the acyl-homoserine lactone that is produced and detected, respectively. Quite expectedly, these residues are in or near the ligand-binding pocket of LasI and LasR. The authors further engineer the LasI/R system to produce and detect the non-native 3OC10-HSL autoinducer in addition to 3OC12-HSL, thereby broadening the specificity of LasI/R.

P. aeruginosa is an important pathogen and a powerful system for the study of quorum sensing. The use of GREMLIN to study how autoinducer synthase and receptor pairs coevolve in terms of sensitivity and specificity for a particular autoinducer is impressive. This paper adds an exciting approach to the growing literature on the evolution of sensitivity and promiscuity in quorum-sensing systems. Further, the authors have developed a thin layer chromatography based approach to separate and detect AHLs from nine samples simultaneously. This methodology should be widely useful to researchers.

The current manuscript does not provide any data about the solubitlity and/or stability of the LasI and LasR mutant proteins being studied. For instance, biochemical analyses would be needed to evaluate if the increased sensitivity of LasRA127L compared to wildtype is due to higher affinity for the autoinducer or because the variant is more stable. Further, this work relies solely on reporter assays and does not address the consequences of these LasI and LasR variants to quorum-sensing dependent P. aeruginosa group behaviors such as pyocyanin production and virulence.

Reviewer #2 (Public Review):

The manuscript entitled "The Shu Complex Prevents Mutagenesis and Cytotoxicity of Single-Strand Specific Alkylation Lesions" by Bonilla and colleagues reports that the yeast Shu complex promotes repair of 3meC in single-stranded DNA during S phase. Specifically, the authors show that mutations and cell lethality induced by MMS in csm2∆ cells are suppressed by overexpression of the human ALKBH2. Further, the authors find that the Csm2-Psy3 module of the Shu complex has increased affinity for 3meC-containing DNA relative to unmodified DNA. The authors propose a model, where the Shu complex binds to 3meC-containing DNA to facilitate HR-dependent post-replicative gap-filling.

Reviewer #2 (Public Review):

Mattis et al have used a hemizygous mutant of the gene Scn1a to study changes underlying the severe epilepsy disorder Dravet syndrome. They describe a change in activation of the dentate gyrus in this mouse model, due to altered excitatory synaptic input. They show that this occurs in the age range after normalization of early inhibitory interneuron dysfunction. This provides an interesting potential mechanism by which neural circuit function is altered even after deficits in inhibition are seemingly corrected. They also report that stimulation of inputs to the dentate gyrus increase seizure susceptibility when body temperature is elevated. Overall these findings indicate a new form of circuit dysfunction that may underlie the etiology of this severe genetic epilepsy disorder.

These findings are not fully complete, and the manuscript suffers from some flaws in experimental design.

The most pressing issue is the lack of a counter-balanced design in experiments testing the ictogenicity of DG stimulation. The authors attempt to justify this stating "there is a theoretical concern that seizure threshold on Day 2 (the second consecutive day of stimulation) could be lowered by a seizure 24 hours prior (a "kindling"-like phenomenon)". In the very next sentence, they cite a study in which this phenomenon has been shown (thus the concern is not theoretical). That said, this is not a semantic argument, but a flaw in experimental design. On day 1, the authors perform experiment A. On day 2, they perform experiment A+B. In an attempt to show that performing experiment A on day 1 does not by itself lead to changes in experiment A+B, they use a separate cohort and show that experiment A does not lead to changes in a repetition of experiment A. Unfortunately, this is not an adequate control. Experiment A+B involves a different set of stimuli, to which the response could very well be altered by the day 1 experiment, but this change would not be revealed with the described experimental design. To determine whether the effect shown in experiment A+B requires a more rigorous, counter-balanced experimental design where one group undergoes experiment A followed by experiment A+B, and a second group undergoes experiment A+B followed by experiment A.

The second major issue is a lack of wild type control groups for several experiments. The experiments presented in Figures 4, 6C and F, and 7 all lack the necessary wild type control measures. Wild type controls were done for Figure 6E, but the data are not presented in the figure.

Some of the cell physiology experiments presented were not optimally designed to provide a relevant mechanistic follow-up to the major findings. For the first major finding of the paper, Figure 2 shows clear and interesting changes in DG activation in the mouse model, and Figure 5 reveals changes to synaptic excitation and inhibition in these neurons. Figure 3 and 4 present data showing changes to PV-interneuron intrinsic properties that only reveal themselves under very intense stimulation. While these findings are interesting and worthy of follow-up, the changes aren't relevant to the synaptic stimulation used in Figure 2.

Finally, Figure 2 has missing data points, seemingly due to cropping of panels. Data visualization is problematic for this vital figure. The fit lines for individual experiments overwhelm the color-filled variance of the mean. Thus, the data in this figure are very difficult to read and interpret. The figure would benefit from including all the individual data points and summary data, but removing the individual fits or putting them into a supplement.

Reviewer #2 (Public Review):

In the manuscript entitled "The Crystal Structure of Bromide-bound GtACR1 Reveals a Pre-activated State in the Transmembrane Anion Tunnel", Li et al. analyzed the effect of bromide binding to GtACR1 by X-ray crystallography and electrophysiology. The authors propose that a bromide ion is bound to the intracellular pocket in the dark, inactivated state and induces a structural transition from an inactivated to a pre-activated state.

I agree that some of the amino acid residues in the current crystal structure change their conformations compared to the previous one reported in 2019 (Li et al., 2019), and it is very impressive that the authors determined the structure using state-of-the-art crystallography technique, ISIMX. However, unfortunately, most of the conclusions and claims described in the manuscript are not well supported by the authors' data.

1) The most serious problem is that the evidence of bromide binding is too weak. The authors showed the composite omit map in Supplementary Figure 1A, but they should present an anomalous difference Fourier map to validate the bromide binding. The authors also claim that they replaced the bromide ion to the water, run the PHENIX refinement, and observed a strong positive electron density at the bromide position in the Fo-Fc difference map (Supplementary Figure 1B). However, when I do the same thing using the provided coordinate and map (I really appreciate the honesty and transparency of the authors), I could not reproduce their result; a weak positive electron density is observed between the bromide position and Pro58 in chain A and there is no positive peak at the position in chain B (Fo-Fc, contoured at 3σ). I am wondering the occupancy and B-factor of the water molecule they show in Supplementary Figure 1B.

In addition to the insufficient evidence, the current models of bromide ions have significant steric clashes. The PDB validation report shows that the top 5 serious steric clashes observed in the coordinate are the contacts between the bromide ions and surrounding residues (PDB validation report, Page 10). I analyzed them and found that the distance between the bromide ion and CG and CD atoms of Pro58 in chain A are only 2.43Å and 2.36Å, respectively. The authors claim that such a close proline-halide interaction has also been observed in the structure of the chloride-pump rhodopsin CIR, but in the structure (PDB ID: 5G28), the distances between the chloride ion and CD and CG atoms of Pro45 are much larger (3.43 and 3.91Å, respectively) and there is no steric clash. Moreover, the authors claim that Pro58 changes its conformation by bromide binding, but it is very possible that the PHENIX program just displaces Pro58 to alleviate the steric clash between the proline and the bromide ion, so the authors should carefully check the possibility.

Overall, the authors should analyze the density again, provide more solid evidence for the bromide binding such as anomalous difference Fourier map, and if they could, they should correct the current significant steric clashes in their models.

2) To analyze the functional importance of putative bromide binding, the authors prepared W246E and W250E mutants and analyzed their electrophysiological properties. Because tryptophan and glutamate are so different in terms of volume and charge, they should analyze other mutants as well. The authors claim that bromide is stabilized by a hydrogen bond interaction formed by the indole NH group of W246, so they should at least test the W246F mutant.

3) The authors claim that the bromide binding in the intracellular pocket induces the conformational change of R94, but the causal relationship is doubtful. As mentioned in the manuscript, R94 forms a salt-bridge with D234 in chain A. However, the arginine has a completely different conformation and does not have any interaction with D234 in chain B. If the bromide binds both in chain A and B and induces the conformational change of R94, why only R94 in chain A interacts with D234? The authors change the pH in the crystallization condition compared to their 2019 study (Li et al., 2019), so the pH may affect the protonation state of D223 and/or other titratable residues and induces the conformational change of R94. The authors should provide more solid evidence for the causal relationship between the bromide binding and the conformational change of R94.

4) The authors assume that the conformational change of R94 creates a functional anion binding site with the Schiff base in GtACR1, but it is too speculative. If the anomalous difference Fourier map does not support the idea, they should delete it.

Reviewer #2 (Public Review):

This study aims at elucidating the substrate-dependent conformational dynamics of TonB-dependent transporter BtuB, which is responsible for vitamin B12 transport in the outer membrane of E. coli. Following the pioneering studies from the same lab, the study employs an innovative approach of in-situ site-directed spin labeling for CW EPR spectroscopy and double electron-electron resonance (DEER) distance measurements in intact E. coli. Despite the intricacy of spin labeling and performing DEER measurements in intact cells, the majority of the obtained DEER spectra are of high quality with impressively long dipolar evolution times and signal-to-noise ratio, enhancing the reliability and accuracy of the data. Despite the limited number of distance constraints on one side of the core domain, experiments are well designed to address the relevant questions and the conclusions are justified by the data. The results fully support the main conclusion that the large substrate-induced structural change on the C-terminal side of the core in the presence of the mutations that mimic the breakage of the R14-D316 ionic lock (i.e., R14A, D316A, D316A/R14A), indicates the shift of the substrate-binding loop 3 towards the periplasm and reproduces the state when the transporter is bound to both substrate and TonB. The authors have utilized the deduced information to assess the currently proposed transport mechanisms. This study provides evidence for a transport mechanism that does not require a mechanical pulling or rotation by TonB as previously proposed. This model is also compatible with the structure of BtuB in complex with TonB that indicates the TonB-dependent release of the ionic lock. It is notable that these results are not seen in reconstituted membranes further highlighting the significance of in-situ structural dynamics studies in general and specifically for the field of EPR spectroscopy. I see substantial advance with respect to, both the mechanism of membrane transport by the TonB-dependent transporters and the application of this innovative approach.

Reviewer #2 (Public Review):

This short report by Yeh et al. reveals the presence of sfRNA in mosquito saliva and that it might enhance DENV infection in human Huh7 cells. By referring to literature, the authors propose that salivary sfRNA is secreted by EVs, and is immunosuppressive. The salivary sfRNA might facilitate DENV transmission and disease prevalence in nature.

Strength: The methods are rigorous, results are clearly presented and the manuscript is well written.

Weakness: sfRNA has long been recognized to interfere with the immune system in the flavivirus field. This study represents a modest advance. Additionally, even as a short report, the study fails to provide sufficient self-standing evidence to support its key claims. The study depends heavily on published literature to support its key conclusions.

Reviewer #2 (Public Review):

Inamdar and colleagues present a convincing manuscript identifying the unique role of IRSp53 in the successful assembly of HIV-1 particles. The study provides insight into the molecular mechanisms underlying the membrane curvature generation associated with virion budding from infected cells. Notably, the authors postulate a model in which the HIV-1 machinery "hijacks" host functions to generate fully-assembled viral particles by recruiting a central virion-assembly factor, HIV-1 Gag, to the luminal extremities of nascent extracellular vesicles generated by endogenous IRSp53. This is achieved through interactions between the two proteins, resulting in supramolecular complexes containing host and pathogen factors.

A significant positive aspect of this study is the implementation of an experimental approach that encompasses complementary techniques, namely biochemistry, super-resolution microscopy, and advanced computational analysis and modelling. This is particularly relevant because several critical studies in the field of HIV-1 often rely on either one or the other set of methods and consequently lack the depth and cross-validation power achieved here. Also, the authors take advantage of experimental models that fall into opposite sides of the natural-artificial spectrum and use them adequately to test hypotheses and make conclusions.

Reviewer #2 (Public Review):

An intact myocardium is essential for cardiac function, yet much remains unknown regarding the cell biological mechanisms maintaining this specialized epithelium during embryogenesis. In this manuscript, Gentile and colleagues discover a novel role for the repressive transcription factor Snai1b in supporting myocardial integrity. In the absence of Snai1b, cardiomyocytes exhibit an enrichment of intermediate filament genes, including desmin b. In addition, the authors detect mislocalization of Desmin, along with adherens junction and actomyosin components, to the basal membrane in snai1b mutant cardiomyocytes, and these mutant cells exhibit an increased likelihood of extrusion from the myocardium. Ultimately, the authors put forward a model wherein Snai1b protects cardiomyocytes from extrusion at least in part by regulating the amount and organization of Desmin in the cell, thereby supporting myocardial integrity.

Overall, the authors highlight an important aspect of epithelial maintenance in an environment that experiences significant biomechanical stress due to cardiac function. By generating a promoter-less allele of snai1b, the authors have created a clean genetic model in which to work. Coupled with beautiful microscopy and transcriptomics, this story has the potential to enlighten both cell biologists and cardiovascular biologists on the underpinnings of myocardial integrity. However, clarifications regarding the overall model would be particularly beneficial for the reader.

1) A clearer discussion of the proposed molecular mechanism for Snai1b function would aid a reader's overall contextualization of this work. At one point, the authors suggest that Snai1b regulates N-cadherin localization to adherens junctions, thereby stabilizing actomyosin tension at cell junctions. Later, it is suggested that Desmin activates the actomyosin contractile network at the basal membrane. It is unclear whether the authors believe that these are separate events or whether they may be coupled, perhaps through Desmin disruption at the lateral membranes, leading to modifications in nearby adherens junctions. A more thorough investigation of the phenotype resulting from desmin b overexpression may clarify this relationship.

2) It appears that extruded cells do not bud off from the myocardium, but rather remain on the apical surface of the existing myocardium. However, it is unclear whether this change in tissue architecture affects cardiac function or the overall morphology of the chamber. A brief discussion of these possibilities would have helped to contextualize the significance of this phenotype.

3) The authors show that cardiomyocyte extrusion is most prevalent near the atrioventricular canal, and they suggest that this regionalized effect is due to the different types of extrinsic factors, like biomechanical forces, that this region experiences. However, it is also possible that regional differences in certain intrinsic factors are involved, such as junctional plasticity, actomyosin activity at the basal membrane, etc. To distinguish between these possibilities, it would have been informative to know whether the extent of N-cadherin/α-18/p-Myosin/Desmin mislocalization varies depending on the regional location of cardiomyocytes within the snai1b mutant heart. For example, do cardiomyocytes near the atrioventricular canal exhibit more extreme effects on N-cadherin/α-18/p-Myosin/Desmin localization than cardiomyocytes in further away portions of the ventricle? Or, do these cells exhibit similar degrees of protein mislocalization, but cells near the atrioventricular canal have a lower threshold for extrusion?

Reviewer #2 (Public Review):

In the manuscript 'A novel mechanosensitive channel controls osmoregulation, differentiation and infectivity in Trypanosoma cruzi' the authors show conclusive evidence that TcMscS is a mechanosensitive channel. They also show that TcMscS has additional roles outside of mechanosensation, likely playing a role in the infectivity of T. cruzi. This manuscript is well written with data that clearly supports the authors hypothesis. The evidence provided in the manuscript clearly shows that TcMscS gates in response to tension and that when knocked out of the genome there is a reduction in infectivity. This work will be impactful to both all researchers studying mechanosensation as it shows that mechanosensitive channels have roles outside of tension sensation.

Recommendations:

A) In the section 'Electrophysiological characterization of TcMscS', the authors present compelling evidence that TcMscS gates in response to tension in the membrane. However, it is unclear, both in the text and the caption, if the trace shown in Figure 2 panel C was collected under tension. If it was, please include the applied pressure value in either the text or caption. Additionally, within this section the applied pressure to the patch is frequently unclear. One way to clear this up would be to 1- add the applied pressure to each trace or to 2- add the applied pressure for each patch to the figure caption. -In Panel E: can you comment on the conductance of the channels in the three traces? Why do you see channels that are approximately 1/2 the size of the first trace in the second two traces?

B) In the section 'TcMscS gene targeting by CRISPR-Cas9' the authors utilized CRISPR to KO TcMscS to determine its function, based on the immunofluorescence and qPCR TcMscS has been successfully knocked out. In lines 251-264, the authors complemented the KO with an overexpression vector in an attempt to confirm the role of TcMscS. In this section, it is very unclear what strains C1 and C2 are and how they are different from one another. Neither of these constructs successfully restores the growth rate. The authors can clarify the differences between the two constructs or they can remove this section from the manuscript, particularly Figure 5 supplement 3. The manuscript is strong and compelling without this panel.

Since publication, the FDA has rescinded its authorisation of bamlanivimab (LY-CoV555), due to its lack of efficacy against circulating variants of concern, particularly B.1.351 (South African), as a result of E484K substitution A,B. Eli Lilly are now pursuing the use of their combination therapy of bamlanivimab with etesevimab (LY-CoV016).

The antibody cocktail REGN-CoV2 showed sustained efficacy against tested variant strains and thus remains a viable treatment option. However, a mutational library scan by Starr et al. revealed that a single amino acid change (E406W) is all that is required for a future variant to escape this therapy C.

Circulating variants highlight the limited efficacy of monoclonal antibodies to an evolving virus, particularly in those which are restricted to the RBD. A diverse panel of monoclonal antibodies, which bind subdominant epitopes may be a more sustainable approach.

A – Wang, P et al. Antibody resistance of SARS-CoV-2 variants B.1.351 and B.1.1.7. 2021. Nature. https://doi.org/10.1038/s41586-021-03398-2

B – Starr, T.N.et al. Complete map of SARS-CoV-2 RBD mutations that escape the monoclonal LY-CoV555 and its cocktail with LY-CoV016. 2021. Cell Reports Medicine. https://doi.org/10.1016/j.xcrm.2021.100255

C – Starr, T.N.et al. Prospective mapping of viral mutations that escape antibodies used to treat COVID-19. 2021. Science. https://doi.org/10.1126/science.abf9302

Kreier, F. (2021). ‘Unprecedented achievement’: Who received the first billion COVID vaccinations? Nature. https://doi.org/10.1038/d41586-021-01136-2