eLife assessment

P-glycoprotein (Pgp) is a major ABC-transporter that couples ATP hydrolysis with the export of xenobiotics and drugs, influencing their pharmacokinetics. The authors provide convincing cryo-EM Pgp structures of drug complexes in previously unforeseen outward-facing conformations. These structures reveal important mechanistic insights that could be of broader impact, but because these transient-like states were captured by cysteine cross-linking the substrate to Pgp, support for the conclusions is incomplete, and further validation is required.

eLife assessment

This paper investigates the potential role of extracellular vesicles in providing extracellular matrix signals for migration of vascular smooth muscle cells. The findings could be useful for researchers interested in cell migration, but the evidence supporting the conclusions is presently incomplete.

eLife assessment

In this important study, the authors apply a residue-resolution protein coarse-grained model to investigate the differences in molecule dimensions and phase behaviour of three HP1 paralogs, HP1 paralog mixtures, and HP1/DNA mixtures. The simulations are well designed to investigate the impact of HP1 sequence on its phase behaviour. Solid evidence reveals that electrostatic interactions are a key determinant of HP1 paralog phase behaviour, thereby advancing our understanding of the molecular mechanisms driving the phase separation behaviour of HP1 paralogs.

eLife assessment

This valuable paper examines the Bithorax complex in several butterfly species, in which the complex is contiguous and not split, as it is in the well-studied fruit fly Drosophila. Based on genetic screens and genetic manipulations of a boundary element involved in segment-specific regulation of Ubx, the authors provide solid evidence for their conclusions, which could be further strengthened by additional data and analyses. The data presented are relevant for those interested in the evolution and function of Hox genes and of gene regulation in general.

eLife assessment

This study employs multiscale simulations to show that PIP2 lipids bind to the DIV S4-S5 linker within the inactivated state of a voltage-gated sodium channel, affecting the coupling of voltage sensors to the ion-conducting pore. The authors report the valuable finding that PIP2 prolongs inactivation by binding to the same site that binds the C-terminal tail during recovery from inactivation, suggesting binding to gating charges in the resting state that may impede activation. The coarse-grained and atomistic molecular dynamics simulations are solid, supporting the claims, although they do lack validation through experiment or simulation, such as studies of linker mutants to confirm PIP2 binding sites, a resting state Nav1.4 model to confirm voltage sensor binding, as well as tests using flexible linkers to probe interactions with PIP2 without backbone restraints.

eLife assessment

This important study advances our understanding of the biological significance of the DNA sequence adjacent to telomeres. The data presented convincingly demonstrates that subtelomeric repeats are non-essential and have a minimal, if any, role in maintaining telomere integrity of budding yeast. The work will be of interest to telomere community specifically and the genome integrity community more broadly.

eLife assessment:

This valuable study provides molecular-level insights into the functional mechanism of bacterial ice-nucleating proteins, detailing important electrostatic interactions in the domain architecture of multimeric assemblies. The evidence supporting the claims of the authors is solid, with results from protein engineering experiments, functional assays, and cryo-electron tomography, while the proposed structural model of protein self-assembly remains hypothetical. The work is of broad interest to researchers in the fields of protein structural biology, biochemistry, and biophysics, with implications in microbial ecology and atmospheric glaciation.

eLife assessment

ZMYM2 is a transcriptional corepressor but little was known about how it is recruited to chromatin. This important study reveals that ZMYM2 homes to distinct classes of retrotransposons bound by the TRIM28 and ChAHP complexes in human cells, which is broadly relevant for the field of transcriptional regulation. Much of the evidence supporting the claims of the authors is convincing. Since widespread ZMYM2-mediated control of transposon activity is not apparent in RNA-seq data, further experiments are needed to demonstrate a more general role beyond the retrotransposons analysed in this study.

eLife assessment

This study provides valuable findings showing the production of IL-22 from intestinal ILC3 during intermittent fasting promotes beigeing of white adipose tissue. However, the study is incomplete because of the lack of mechanistic insight. In particular, the authors should clearly show the mechanism by which IL-22-derived from ILC3 directly induces beigeing.

eLife assessment

Little is known about the role of the microbiome alterations in epithelial ovarian cancer. This important and rigorous study carefully examined the microbiome composition of 1001 samples from close to 200 ovarian cancer cases and controls, and presents compelling evidence that the fallopian tube microbiota are perturbed in ovarian cancer patients. These insights are expected to fuel further exploration into translational opportunities stemming from these findings.

eLife assessment

The presence or absence of a surrounding envelope, previously a clear distinguishing feature of different viruses, has been blurred by the recent recognition that many so-called 'nonenveloped' viruses are released from cells as quasi-enveloped virions cloaked in host cell membranes. This mechanism of viral egress allows for non-lytic infection, and has potentially important implications for pathogenesis. In this manuscript, Jassey and colleagues provide solid evidence that the protein deacetylase SIRT-1 is required for the non-lytic release of enteroviruses in extracellular vesicles.

eLife assessment

This study provides important information on the biogenesis of eccDNAs during spermatogenesis. The data presented are solid and supportive of the concussion that eccDNAs in spermatogenic cells are not derived from miotic recombination hotspots but rather represent oligonucleosomal DNA fragments from apoptotic male germ cells, whose ends are ligated through microhomology-mediated end-joining. This work is of interest to researchers working on germ cell biology and cancer biology.

eLife assessment

Despite the importance of T follicular helper cells (Tfh cells) in vaccine-induced humoral responses, it is still unclear which type of Tfh cells (Tfh1, Tfh2, and Tfh17) is critical for generating protective humoral immunity. By using the rhesus macaques model (most similar to human), the authors have addressed this potentially important question and obtained suggestive data that Tfh1 is critical. Although being suggestive, the evidence for the importance of Tfh1 is incomplete.

eLife assessment

This study presents a valuable finding on the function of the gene Cst7 in sex-divergent pathological changes in microglia in a mouse model of AB-driven Alzheimer's disease. The evidence supporting the claims of the authors is solid, although the study would be strengthened by validation of some of the key differentially expressed genes identified in RNA-sequencing experiments, and the inclusion of key controls and additional timepoints to address whether Cst7 drives disease progression or is simply upregulated as a result. The work will be of interest to neuroimmunologists and neuroscientists working on microglia and neurodegenerative disease.

eLife assessment

This study provides important insights into the structural biology and molecular mechanism of the sensory proteins ToxR/S that are associated with survival and virulence of the cholera pathogen. The structural studies are solid and supported by a series of biophysical experiments revealing a split, periplasmic protein binding interface for bile acid. The results are of interest to both protein biochemistry and pharmacology, potentially opening new routes for intervention in cholera disease.

eLife assessment

This study presents a valuable methodology with increased efficiency and precision of gene editing in human primary hematopoietic stem and progenitor cells. The evidence supporting the claims is solid, although a more extensive assessment of functional hematopoietic stem cell impacts and potential off-target effects would be helpful to further strengthen the conclusions. The work will be of interest to biologists studying hematopoietic stem and progenitor cells and gene targeting for potential clinical applications.

eLife assessment

This study used ChatGPT to assess certain linguistic characteristics (sentiment and politeness) of 500 peer reviews for 200 neuroscience papers published in Nature Communications. The vast majority of reviews were polite, but papers with female first authors received less polite reviews than papers with male first authors, whereas papers with a female senior author received more favourable reviews than papers with a male senior author. Overall, the study is an important contribution to work on gender bias, and the evidence for the potential utility of generative AI programs like ChatGPT in meta-research is solid.

eLife assessment

The work by O'Reilly and Delis is important to extend the synergy ideas using methods from signal processing and information theory to cluster muscles and task parameters, thereby advancing our understanding of the modular architecture of motor control. The method is innovative, and the findings are compelling from theoretical and practical perspectives. The work will be of broad interest to motor control and neural engineering researchers.

eLife assessment

Picard et al. propose a Facial Expression Pain Signature (FEPS) as a distinctive marker of pain processing in the brain. In this valuable study, they attempt to use functional magnetic resonance imaging (fMRI) data to predict facial expressions associated with painful heat stimulation. The crucial aspect of the claim is whether the proposed biomarker is distinctive, i.e., it is specific enough to distinguish facial expressions in response to pain from similar facial expressions of non-painful origin. The experimental setup does not control for that condition, so the evidence is incomplete to support their main claims.

eLife assessment

This important study in Drosophila identifies neuronal mechanisms influenced by altered activity during a critical period (CP) of larval locomotor circuit development. Increasing activity during the CP causes permanent network changes indicating that a setpoint of network excitability is determined during the CP. Most importantly, for excitability setpoint determination during the CP excitatory and inhibitory inputs are integrated such that the effect of CP hyperexcitation can be rescued by the stimulation of endogenous inhibitory inputs to the motoneurons. Solid experimental evidence supports their novel insight into how developing neural network excitability is tuned and how it can be entrained during the CP.

eLife assessment

In this manuscript, Jain and colleagues explore whether increasing adult neurogenesis is protective against status epilepticus and the development of spontaneous recurrent seizures (chronic epilepsy) in a mouse pilocarpine model of temporal lobe epilepsy. This is an important work that provides solid data, contradicting previous studies on suppressing chronic seizures by reduced neurogenesis. To ensure the validity and robustness of the results, there is a need for additional controls and validations.

eLife assessment

This study presents a valuable insight into a computational mechanism of pain perception. The evidence supporting the authors' claims is solid, although the inclusion of 1) more diverse candidate computational models, 2) more systematic analysis of the temporal regularity effects on the model fit, and 3) tests on clinical samples would have strengthened the study. The work will be of interest to pain researchers working on computational models and cognitive mechanisms of pain in a Bayesian framework.

eLife assessment

The findings described in this manuscript are important and have theoretical or practical implications beyond a single subfield. Both reviewers comment on the strength of the paper because of the use of multiple approaches (optical imaging, physiology, rapid freeze EM, modeling, double mutant analysis). The methods, data, and analyses provide convincing support for the distinct roles of the two proteins, Doc2a and Syt7 in regulating SV release.

eLife assessment

Gain-of-function mutations and amplifications of PPM1D are found across several human cancers and are associated with advanced tumor stage, worse prognosis, and increased lymph node metastasis. Unfortunately, clinical translation has so far not been possible due to the lack of PPM1D inhibitors with favorable pharmacokinetic properties. This useful study leveraged CRISPR/Cas9 screening to identify that SOD1 inhibition is synthetic lethal with PPM1D mutation in leukemia, although the mechanistic analyses were incomplete.

eLife assessment

This study presents compelling observations of the ability of Ym1 crystals to promote type 2 immunity in vivo, in mice, that advances our knowledge of how the abundantly expressed chitinase-like protein family may function, with new methodology, new crystal structures that are more rigorous than what is currently known and very convincing immunological data. The significance of the study is important, providing an exciting opportunity to mechanistically study the impact of protein crystals in vivo in relation to human disease. This work will be of broad interest to immunologists and researchers working on type 2 inflammatory disease, in the lung.

eLife assessment

This important work by Aballay et al. advances our understanding of how G protein-coupled receptors (GPCRs) regulate immunity and pathogen avoidance. The authors provide convincing evidence for the GPCR NPR-15 to mediate immunity by altering the activity of several key transcription factors. Nevertheless, both reviewers have raised questions to be addressed, at which time this work will be of broad interest to immunologists specifically, and biologists in general.

Evaluation statement (1 September 2023)

Flores-Aldama and colleagues set out to identify molecular determinants of fast inactivation in the TRPV6 ion channel, a mechanism not observed in the closely related TRPV5 channel. The work focuses on a helix-loop-helix (HLH) motif, located at the interface between several important regions for channel gating. Using molecular dynamics simulations and analysis of mutations, the authors identify pairs of amino acid residues in a structural triad formed by the HLH, S2-S3 linker, and transmembrane domains, which show different conformations in the available TRPV5 and TRPV6 cryo-EM structures. An important aspect of the study is that some of the structural hypotheses were derived from an evolutionary analysis of sequences from orthologues of both channels, demonstrating the value of this type of analysis.

Biophysics Colab considers this to be a convincing study and recommends it to scientists interested in the molecular determinants of ion channel gating.

(This evaluation by Biophysics Colab refers to the version of record for this work, which is linked to and has been revised from the original preprint following peer review.)

eLife assessment

This valuable manuscript investigates the role of a subpopulation of glutamatergic neurons in the suprammamillary nucleus that projects to the pre-optic hypothalamus area in active coping but not locomotor activity. It provides solid evidence from experiments using fibre photometry or photostimulation during threatening tasks that these neurons allow animals to produce flexible behaviours in response to stress. However, the evidence is incomplete in several ways, including validation and quantification of anatomical tracing data that serve as a basis for the behavioral testing, the use of statistics, sex as a biological variable, genotype differences between experimental and control groups in behavioral tests, limiting its broad interest to neuroscientists across sub-fields.

eLife assessment

This article provides important findings on how bacteria use small RNAs to regulate flagellar expression with implications for multiple fields. The data supporting the conclusions are convincing with a large amount of data that include results from phenotypic analyses, genomics approaches as well as in-vitro and in-vivo target identification and validation methods. This study on the varied effects of three sRNAs (UhpU, FliX and MotR) is of broad interest to RNA biochemists and microbiologists.

eLife assessment

This study based on the use of Cancer Drug Resistance Accelerator (CDRA) chip is valuable as a platform technology to assess chemoresistance mechanisms. The strength is convincing from the technological point of view. However, the use of a single cell line model and the lack of mechanistic insights could be improved.

eLife assessment

The findings in this study are important and have a practical implication for early cancer diagnosis. Furthermore, we found the ML approach and data analysis compelling. However, significant concerns regarding the quality of the source materials used for the analysis have been raised and need to be addressed.

eLife assessment

This article brings to bear a useful, extensive set of behavioral methods and neural data to report that activity in numerous cortical regions robustly covaries with the complexity of tone sequences encoded in memory. In its current state, the findings are solid but deserve further analysis to arrive at more convincing conclusions.

eLife assessment

This study reports a valuable finding on the discovery and evaluation of a potent small molecule inhibitor for CRM1 that may be important to treat extranodal NK/T cell lympohma (ENKTL). The evidence supporting the claims of the authors is solid that reflects an important finding for novel CRM1 inhibitors to treat ENKTL, although additional experimental evidence is needed. The work will be of interest to cancer biologists working on ENKTL.

eLife assessment

This paper measures the neural activity in reach-to-grasp and reach-only tasks using intrinsic optical imaging. The paper describes these in the relationship to the Intra-cortical micro stimulation maps of the same animals. The dataset is unique and potentially highly important. However, the claim of "clustered neural activity" is not tested against any quantifiable alternative hypothesis of non-clustered activity, and support for this idea is therefore currently incomplete.

eLife assessment

This study uncovers a unique feature of the nucleotide binding domain interface in human CFTR, offering valuable insights into the effects of different non-hydrolytic mutations on CFTR gating. While the evidence presented is solid, a more thorough examination of the non-hydrolytic mutants of zebrafish CFTR for comparison would strengthen the authors' claims. In the current form, more cautious interpretations of some of the data are needed. This study will be of interest to researchers in the fields of cystic fibrosis and proteins in the ATP Binding Cassette (ABC) transporter family.

eLife assessment

This paper will be of considerable interest to population geneticists and other scholars in the field of paleogenomics. The study provides an impressive dataset containing 200+ novel human ancient genome sequences and a very creative, robust, and novel approach for studying human migration across time using ancient DNA. The authors find that the population structure in Europe has been remarkably stable over time. The conclusions are well supported by the data and the methods used are thoughtful and rigorous.

eLife assessment

This important study examines the existence of a fear memory engram in acetylcholine neurons of the basal forebrain and seeks to link this to modulation of amygdala for fear expression. Using a combination of techniques including genetic access to cFos expressing neurons, in-vivo chemogenetics, and optical detection of acetylcholine (ACh), the authors present solid evidence that posteriorly-located amygdala projecting basal forebrain cholinergic neurons participate in cue-specific threat learning and memory. This paper will be of interest to those studying circuit-level mechanisms of learning and emotion regulation.

eLife assessment

This is an important study of the mechanism of how binding of the fatty acid myristic acid (MYR) inhibits the activity of the kinase c-Abl, a critical regulator of many cellular processes. While the general aspects of this regulation are known from structure determination and biochemical studies, the exact molecular mechanism and the nature of the allosteric inhibition were not known. The authors use MD simulation to close this gap and provide a detailed mechanistic description of the inhibitory mechanism, although some of the evidence remains incomplete.

eLife assessment

This valuable multicenter study provides solid evidence that the auditory noise emitted during online transcranial ultrasound stimulation (TUS) protocols can pose a considerable confound and is able to explain corticospinal excitability changes as measured with Motor Evoked Potentials (MEP). The findings may lay the ground for future studies optimizing protocols and control conditions to leverage transcranial ultrasound stimulation as a meaningful experimental and clinical tool. A clear strength of the study is the multitude of control conditions (i.e., control sites, acoustic masking, acoustic stimulation).

eLife assessment

This is an important study that improves gene models for the ferret genome and identifies neural progenitors that are comparable to those found in developing human brains. The data are convincing and clearly presented. Of particular interest to the field, the work identifies enriched expression of FOXJ1 in late truncated radial glia, strongly indicating that towards the end of neurogenesis, these cells likely give rise to ependymal cells. The work is of interest to anyone studying the development of the nervous system, especially colleagues studying the evolution of development.

eLife assessment

This is an important study that improves gene models for the ferret genome and identifies neural progenitors that are comparable to those found in developing human brains. The data are convincing and clearly presented. Of particular interest to the field, the work identifies enriched expression of FOXJ1 in late truncated radial glia, strongly indicating that towards the end of neurogenesis, these cells likely give rise to ependymal cells. The work is of interest to anyone studying the development of the nervous system, especially colleagues studying the evolution of development.

eLife assessment

This is a valuable investigation of the chromatin dynamics throughout the cell cycle by using fluorescence signals and patterns of GFP-PCNA and CY3-dUTP, which labels newly synthesized DNA. The authors report reduced chromatin mobility in S relative to G1 phase. The technology and methods used are solid. The data will be of interest to researchers working on chromatin dynamics.

eLife assessment

This is an important study that contributes to our understanding of the role of beta-arrestins in endosomal activation of the vasopressin type 2 receptors. While the use of a minigene as a tool is a weakness, the evidence is overall convincing and makes for significant findings whose theoretical and practical implications extend to other GPCRs.

eLife assessment

This manuscript describes a deep mutational scanning study of the kinase domain of the MET receptor tyrosine kinase. The study yields a valuable catalog of essentially all possible deleterious mutations in this portion of the receptor., with convincing evidence. The manuscript will be of interest to researchers working in the field of receptor tyrosine kinases.

eLife assessment

This study combines CLIP, RNA-seq, and splicing assays with manipulation of RBMX and its homologs RBMY and RBMXL2 to show that the RBMX family suppresses the recognition of cryptic splicing within long exons. The study is important in that it puts forward the intriguing claim that the RBMX family is responsible for the cryptic splice site repression in ultra-long exons. The methods, data, and analyses supporting the claims are solid, broadly supporting the claims, with some weaknesses. The generalization of the findings is somewhat overstated but could be strengthened by deeper statistical integration of the RNA-seq and CLIP datasets.

eLife assessment

This important manuscript focuses on the mechanisms by which food signals and food ingestion modulate animal foraging. The authors provide solid support for the interesting idea that chemosensory and interoceptive signals converge on transcriptional regulation of the TGF-beta ligand DAF-7 in a single pair of C. elegans chemosensory neurons to regulate behavior. Their studies implicate a conserved signaling molecule, ALK, in this regulation, suggesting a conserved link between food cues and the neuroendocrine control of foraging behavior.

eLife assessment

This important study provides convincing evidence of the criticality of estradiol - estrogen receptor-mediated upregulation of kisspeptin within neurons of the preoptic area to generate an ovulation-inducing luteinizing hormone surge. The use of in vivo CRIPSR-Cas9 is novel in this system and provides a road map for future studies in reproductive neuroendocrinology. This paper will be of interest to reproductive neuroscientists and endocrinologists.

eLife assessment

This study presents potentially useful findings describing how activity in the corticotropin-releasing hormone neurons in the paraventricular nucleus of the hypothalamus modulates sevoflurane anesthesia, as well as a phenomenon the authors term a "general anesthetic stress response". The technical approaches are solid and the data presented are largely clear. However, the primary conclusion, that the PVHCRH neurons are a mechanism of sevoflurane anesthesia, is inadequately supported.

eLife assessment

This paper explores how Notch activity acts together with homeodomain transcription Bsh factors to establish distinct cell fates (L4 vs L5) in the visual system of Drosophila. The findings are important and have theoretical or practical implications beyond a single subfield. The methods, data and analyses are solid, and broadly support the claims with only minor weaknesses.

eLife assessment

This paper, offering insights into the mechanisms of neuronal cell type diversification, provides important findings that have theoretical or practical implications beyond a single subfield. The data are compelling and provide evidence that features methods, data and analyses that are more rigorous than the current state-of-the-art.

eLife assessment

The presented study introduces a valuable non-AI computational method for segmenting noisy grayscale images, particularly highlighting its applicability in identifying immunostained potassium ion channel clusters. While the method's avoidance of AI training appeals to those lacking computational know-how and shows improved accuracy over basic threshold-based techniques, there are valid concerns regarding its performance in comparison to advanced methodologies. The evidence supporting the method's efficacy is solid but incomplete, necessitating comparisons to more advanced techniques and the provision of user-friendly computational tools for a comprehensive evaluation.

eLife assessment

This work is valuable in exploring the role of nutritional immunity in bacteria-mediated tumor therapeutics, involving proteomics and in vivo mouse model results which provide largely solid supporting evidence of the observational claims, but is incomplete when extrapolating the mechanisms of how manipulation of iron status can affect E. coli-mediated tumor therapy. This work is of interest to a broad audience including researchers in cancer biology, cell biology, and microbiology.

eLife assessment

This valuable manuscript systematically addresses the role of intracellular lipid transfer proteins on cellular lipid levels. The authors have provided solid evidence on the role of ORP9 and ORP11 in sphingolipid metabolism at the Golgi complex. The data is in general convincing, except for the claim that ORP9/11 might counter-exchange phosphatidylinositol 4-phosphate and phosphatidylserine, which is not fully supported by the data presented. This article will be of broad interest to cell biologists interested in lipid metabolism and membrane biology.

elife assessment:

This study presents valuable new findings on the role and mechanism of action of the poorly characterized cell surface protein TMEM263 in regulating mouse postnatal growth. The evidence supporting the whole-body growth and skeletal phenotypes, as well as the disruption of GH/insulin-like growth factor-1 (IGF-1) signaling seen in TMEM263 knockout mice, is convincing. However, additional data are needed to definitively conclude that the observed alteration of hepatic GH/IGF1 signaling is causative of the body growth and skeletal phenotypes.

eLife assessment

This is a confirmatory study providing useful findings bearing on the regulation of the inflammatory response of skeletal muscle macrophages by ultrasonic mechanostimulation. It provides solid data on its effect on promoting extracellular vesicle secretion and consolidates previous studies.

eLife assessment

This study presents valuable findings on the organization of respiratory chain complexes in mitochondria. It provides solid evidence that respiratory supercomplex formation in the fruit fly does not impact respiratory function, suggesting the role of these complexes is structural, rather than catalytic. However, whether the conclusions extend to other species requires further evidence. This manuscript will be of broad interest to the field of mitochondrial bioenergetics.

eLife assessment

This fundamental study substantially advances our physical understanding of the sharp increase and saturation of the viscosity of non-confluent tissues with increasing cell density. Through the analysis of a simplified model this study provides compelling evidence that polydispersity in cell size and the softness of cells together can lead to this phenomenon. The work will be of general interest to biologists and biophysicists working on development.

eLife assessment

This paper provides important insights into how asexual reproduction can arise in interspecific hybrids. The evidence supporting the conclusions is compelling, with rigorous molecular cytogenetic experiments showing the production of clonal gametes is common across hybrids between closely to moderately divergent sexual species. By highlighting the potential for asexuality to evolve in hybrids during a narrow window of species divergence, this work will be of broad interest to evolutionary biologists.

eLife assessment

The authors discuss an effect, "diffusive lensing", by which particles would accumulate in high-viscosity regions, for instance in the intracellular medium. To obtain these results, the authors rely on agent-based simulations using custom rules performed with the Ito stochastic calculus convention. The "lensing effect" discussed is a direct consequence of the choice of the Ito convention without spurious drift which has been discussed before and is likely to be inadequate for the intracellular medium, causing the presented results to likely have little relevance for biology.

eLife assessment

This important study combines cellular assays and biophysical methods to identify heparan sulfate (HS) as a regulator of apoptosis. The multidisciplinary approach for investigating HS•TRAIL interactions is highly compelling, but the central hypothesis could be further strengthened from additional investigations on downstream effects. This paper is very relevant to cancer biologists and biophysicists working on TRAIL-dependent apoptotic pathways.

eLife assessment

This manuscript aims to unravel the contribution of cholesterol to aquaporin-0 (AQP0) tetramer array formation within lens membranes. Convincing electron crystallography and molecular dynamics identify a specific cholesterol binding site of significance to protein clustering within lipid rafts. The important work advances our understanding of membrane biology and will be of broad interest to membrane transport biologists, biochemists, and structural biologists.

eLife assessment

This valuable work by Pulfer et al. advances our understanding of spatial-temporal cell dynamics both in vivo and in vitro. The authors provide convincing evidence for their innovative deep learning-based apoptosis detection system, ADeS, that utilizes the principle of activity recognition. Nevertheless, the work is incomplete due to the authors' claim that their system is valid for non-fluorescently labeled cells, without evidence supporting this notion. After revisions, this work will be of broad interest to cell biologists and neuroscientists.

eLife assessment

This study presents a valuable finding that serum androstenedione levels may provide a new biomarker for early detection and progression of glaucoma, although a single biomarker is unlikely to be singularly predictive due to the etiological heterogeneity of the disease. The strength of the evidence presented is solid, supported by multiple lines of evidence.

eLife assessment

This valuable study evaluates the effects of nifuroxazide on radiotherapy for the treatment of hepatocellular carcinoma. Solid evidence is provided to support the conclusion that nifuroxazide facilitates the downregulation of PD-L1 and may improve outcomes when combined with radiotherapy, though the inclusion of additional cell lines and animal models would have strengthened the study. This work will be of interest to cancer biologists and those working in immuno-oncology.

eLife assessment

These solid results demonstrate that arpin is expressed in the endothelium of blood vessels and its deficiency leads to leaky blood vessels in in vivo and in vitro models, although the work does not clarify the mechanistic connection between arpin and increased ROCK activity. The study adds some insights to our understanding of the complicated network of proteins that control this process, and it will be useful to individuals within this defined field of study.

eLife assessment

Studying several allergens in different mouse strains, the authors assess the role of IgM in airway inflammatory responses and show that IgM deficient mice have reduced airway hyperresponsiveness. Although the findings, based on experimental evidence from a wide range of immunological and other assays, including the expression of a protein that regulates actin in smooth cells, are interesting and useful, the study is incomplete as the data and analyses do not support their primary claims.

eLife assessment

This solid study presents valuable insights into the role of Cers1 on skeletal muscle function during aging, although further substantiation would help to fully establish the experimental assertions. It examines an unexplored aspect of muscle biology that is a relevant opening to future studies in this area of muscle research.

eLife assessment

Despite the importance of long-lived plasma cells (LLPCs), particularly in the vaccination field, their natures are still unclear. In this valuable manuscript, as a first step towards clarifying these natures, the authors used a solid genetic approach (time-stamping one) and successfully labelled only functional LLPCs. Although four groups have already published data by the same genetic approach, the authors' manuscript includes additional significant findings in the LLPC field.

eLife assessment

This manuscript presents a valuable approach to exploring CD4+ T-cell response in mice across stimuli and tissues through the analysis of their T-cell receptor repertoires. The authors use a transgenic mouse model, in which the possible diversity of the T-cell receptor repertoire is reduced, such that each of a diverse set of immune exposures elicits more detectably consistent T-cell responses across different individuals. However, whereas the proposed experimental system could be utilized to study convergent T-cell responses, the analyses done in this manuscript are incomplete and do not support the claims due to limitations in the statistical analyses and lack of data/code access.

eLife assessment

The authors' finding that PARG hydrolase removal of polyADP-ribose (PAR) protein adducts generated in response to the presence of unligated Okazaki fragments is important for S-phase progression is potentially valuable, but the evidence is incomplete, and identification of relevant PARylated PARG substrates in S-phase is needed to understand the role of PARylation and dePARylation in S-phase progression. Their observation that human ovarian cancer cells with low levels of PARG are more sensitive to a PARG inhibitor, presumably due to the accumulation of high levels of protein PARylation, suggests that low PARG protein levels could serve as a criterion to select ovarian cancer patients for treatment with a PARG inhibitor drug.

eLife assessment

This valuable study describes a new role of epithelial intercellular adhesion molecule 1 (ICAM-1) protein in controlling bile duct size. The effect is mediated via EBP-50 and subapical actomyosin to regulate size of bile canaliculi. These solid findings have theoretical and practical implications in hepatology and human disorders of bile ducts.

eLife assessment

This is a solid methods paper developing a machine learning based protocol differentiating normal and diseased myofibers. It emerges with and validates a potentially valuable approach to diifferentiate healthy and Duchenne muscle dystrophy myofibers.

eLife assessment

The studies are important to the field of hepatic steatosis and inflammation. The data provided are convincing that treatment with I3A mitigated Western diet (WD)-induced hepatic steatosis, inflammation and reversed WD-induced alterations in liver bile acids and free fatty acids in mice.

eLife assessment

This study provides a useful description of two evolutionary conserved motifs in CD4 that contribute to the CD4-mediated enhancement of T cell receptor (TCR) signaling, which are notably acting independently of the CD4-LCK interaction. The supporting evidence is solid and supported using a T cell hybridoma in vitro system, but remains incomplete without experiments with primary T cells or the use of in vivo models to assess the importance of the CD4 motifs in T-cell biology.

eLife assessment

This important work identifies a previously uncharacterized capacity for songbirds to recover vocal targets even without sensory experience. While the evidence supporting this claim is solid, with innovative experiments exploring vocal plasticity in deafened birds, additional behavioral controls and analyses are necessary to shore up the main claims. If improved, this work has the potential for broad relevance to the fields of vocal and motor learning.

eLife assessment

In this important study, the rabbit was used as a non-rodent mammalian model to show that DMRT1 has a testicular promoting function as it does in humans. The experiments are meticulous and compelling, and the arguments are clear and convincing. These results may explain the gonadal dysgenesis associated with mutations in human DMRT1 and highlight the need for mammalian models other than mice to better understand the process of gonadal sex determination in humans.

eLife assessment

This study presents Bactabolize, a valuable tool for the rapid genome-scale reconstruction of bacteria and the prediction of growth phenotypes. Using validated methodology, the tool relies on a reference pan-genome model to create strain-specific draft metabolic models, as demonstrated in this study using Klebsiella pneumoniae. While the evidence in this specific case is solid, validation across diverse bacterial species is yet to be confirmed.

eLife assessment

This is an important study showing how dendritic plateau potentials can enable neurons to perform reliable 'binary' computations in the face of realistic spike time jitter in cortical networks. The authors make a surprising and novel claim that dendritic plateau potentials perform equally well in short integration windows of only 10 ms and detail a biophysical mechanism for how this effect may occur. While the authors make many good arguments, and the general concept underlying the paper is sound, the evidence as of now is incomplete, with some unsupported statements that should be more thoroughly defended in the manuscript.

eLife assessment:

Chen and colleagues utilize an in situ explant model of the neural retina and retinal pigment epithelium (RPE), along with small molecule inhibition of key metabolic enzymes and targeted metabolomic analysis, to decipher key differences in metabolic pathways used by rods, cones, Muller glia, and the RPE. They conclude that rods are heavily reliant on oxidative metabolism, cones are heavily reliant on glycolysis, and multiple mechanisms exist to decouple glycolysis from oxidative metabolism in the retina. This study provides valuable metabolomic data and insights into the metabolic flexibility of different retinal cells. However, the evidence supporting the authors' claims is incomplete and would benefit from validating their evidence with animal models, comparing their data to previously published literature, performing C13 tracing experiments, and assessing mitochondrial function.

eLife assessment

This valuable study shows that distinct midbrain dopaminergic axons in the medial prefrontal cortex respond to aversive and rewarding stimuli and suggest that they are biased toward aversive processing. The use of innovative microprism based two-photon calcium imaging to study single axon heterogeneity is solid, although the experimental design could be optimized to distinguish aversive valence from stimulus salience and identity in this dopamine projection. This work will be of interest to neuroscientists working on neuromodulatory systems, cortical function and decision making.

eLife assessment

This work identifies cnidarian neuropeptides and pairs them to their GPCR, then shows that neuropeptide signaling systems have evolved and diversified independently in cnidarians and bilaterians. Neuropeptide-receptor partners were experimentally identified using established and widely used methodologies including single cell mapping, providing compelling evidence for the conclusions of the paper. This impressive accomplishment provides fundamental new insights into the evolution of neuropeptide signaling systems and will be of broad interest to neurobiologists and evolution of development researchers.

eLife assessment

The study by Chardon et al. is fundamental to advancing our understanding of presynaptic control of motor neuron output. Large-scale computer simulations were performed using well-established single motor neuron models to provide compelling evidence regarding the time-varying patterns of inputs that control motor neuron ensembles. The work will interest the community of motor control, motor unit physiology, neural engineering, and computational neuroscience.

eLife assessment

In this valuable study, Parrotta et al. showed that it is possible to modulate pain perception and heart rate by providing false heart rate (HR) acoustic feedback before administering electrical cutaneous shocks. The evidence supporting the claims of the authors is rather solid, although what they consider an interoceptive signal is not necessarily supported as such by the results. In this regard, including a larger number of trials per participant, increasing the sample size, and adding a measure of actual pain perception after its induction would have strengthened the study. Although mechanisms and some alternative explanations for this effect remain to be addressed, the work will nonetheless be of interest to neuroscientists working on predictions and perception, health psychologists, pain researchers, and placebo researchers.

eLife assessment

The study presents valuable insights into the regulation of astrocytes in the long-term potentiation of excitatory synapses onto inhibitory interneurons. However, reviewers identified concerns regarding originality and proper acknowledgment of replicated work, representation of interneuron diversity, and the robustness of certain conclusions. The strength of evidence provided is deemed incomplete, necessitating significant revisions for clarity, and accuracy, and to address highlighted concerns.

eLife assessment

This valuable study uses a combination of behavioral studies and computational modeling to argue that humans implicitly represent gravity as a Gaussian distribution (rather than as a deterministic vector). The experimental evidence is generally solid, with all experiments and model simulations being consistent with the proposed account. However, in the current version of the manuscript, the evidence for the main claim of the paper is incomplete: a number of alternative interpretations such as uncertainty about the presence of external lateral forces, or uncertainty about other object properties have not been considered sufficiently.

eLife assessment

The authors have updated a helpful resource for the neuroscience community regarding hippocampal cell types and their interactions, called the "hippocampome." The updates expand content, tools, and potential applications, and therefore the significance is important. The strong presentation makes the paper compelling.

eLife assessment

This valuable study analyzed a large cohort of Adolescent Idiopathic Scoliosis (AIS) patients and identified an association with a variant in COL11A1 (Pro1335Leu). Experimental testing of this potentially pathogenic variant suggests a link between Pax1, Col11a1, Mmp3, and estrogen signaling. While the connection between transcriptional regulation, the extracellular matrix, and estrogen signaling is compelling, the experimental evidence supporting this link is still incomplete and would benefit from a more direct and comprehensive approach. Strengthening the functional testing would help establish the pathogenicity of a collagen variant and the role of the proposed Pax1-Col11a1-Mmp3 signaling axis in AIS.

eLife assessment

This paper presents important new data related to the non-canonical SMC complex MukBEF in E. coli, which remains less well understood compared to more canonical SMC complexes in many other bacteria. The authors use a combination of Hi-C and ChIP-seq to demonstrate that MukBEF loads at multiple locations along the chromosome, preferentially in newly replicated DNA, with the MatP/matS system antagonizing MukBEF activity and localizing in terminus proximal regions. Most of the data to support these findings are compelling, with modest concerns raised about the effect sizes of the ChIP-seq data, which could be bolstered with some additional controls.

eLife assessment

This fundamental study provides compelling evidence to explain how chemical variations within a set of kinase inhibitors drive the selection of specific Erk2 conformations. Conformational selection plays a critical role in targeting medically relevant kinases such as Erk2 and the findings reported here open new avenues for designing small molecule inhibitors that block the active site while also steering the population of the enzyme into active or inactive conformations. Since protein dynamics and conformational ensembles are essential for enzyme function, this work will be of broad interest to those working in drug development, signal transduction, and enzymology.

eLife assessment

This is a valuable contribution to the electric fish community, and to studies of active sensing more generally, in that it provides evidence that a well-studied behavior (chirping) may serve in active sensing rather than communication. For the most part, the evidence is solid. In particular, the evidence showing increased chirping in more cluttered environments and the relationship between chirping and movement are convincing. Nevertheless, evidence to support the argument that chirps are mostly used for navigation rather than communication is incomplete.

eLife assessment

The authors investigate the mechanism of amyloid nucleation in a cellular system using novel ratiometric measurements, providing fundamental insight into the role of polyglutamine length and the sequence features of glutamine-rich regions in amyloid formation. The problem addressed by this study is very significant and the ability to assess nucleation in cells is of considerable value. The data, as presented and analyzed, are mostly convincing.

**Editors Assessment: **

While Bacterial Artificial Chromosomes libraries were once a key resource for building the human genome project over time they have been rendered relatively obsolete by long-read technologies. In the era of CRISPR-Cas systems pairing this data with one of the many guide-RNA libraries to find targets for manipulation with CRISPR tools is bringing back BACs advantages for genomics. With this in mind the authors have developed a BAC restriction map database containing the restriction maps for both uniquely placed and insert-sequenced BACs from 11 libraries covering the recognition sequences of available restriction enzymes. Alongside a set of Python functions to reconstruct the database and more easily access it (which were debugged and had improved documentation added during review). The presented data should be valuable for researchers simply using BACs, as well as those working with larger sections of the genome in terms of synthetic genes, large-scale editing, and mapping.

*This evaluation refers to version 1 of the preprint *

**Editors Assessment: **

This work presents a new standardized graphical approach for visualizing genetic associations across a wide range of allele frequencies. These proposed TrumpetPlots have a distinctive trumpet shape, hence the proposed name. With the majority of variants having low frequency and small effects, while a small number of variants have higher frequency and larger effects, this view can help to provide new and valuable insights into the genetic basis of traits and diseases, and also help prioritize efforts to discover new risk variants. The tool is provided as a novel R package and R Shiny application and to demonstrate its use the article illustrates the distribution of variant effect sizes across the allele frequency range for over 100 continuous traits available in the UK Biobank. After some problems in testing the package is now available and easy to deploy via CRAN.

*This assessment refers to version 1 of this preprint. *

eLife assessment

This is an important biophysical study combining native mass spectrometry with mutant cycles to estimate the thermodynamic components of lipid A binding to the ABC transporter MsbA. Binding energies for lipid-protein interactions are proposed, but the evidence is currently incomplete as they rely on a number of technical and theoretical assumptions, mostly related to multiple lipid binding sites in the protein.

eLife assessment

The manuscript represents an important contribution to an ongoing discussion about the substrate binding site and mechanism of the Bile Acid Sodium Symporter (BASS) family of transporters. Structural and biochemical analysis of a bacterial homolog, ASTBnm, in complex with its native substrate (not bile acids, but a vitamin A precursor, pantoate) show a new binding site that is consistent with classical proposals for elevator-type transport mechanisms. Molecular dynamics (MD) simulations highlight the improved stability for the substrate in the active site when ions are present, suggesting a binding order during the transport cycle. The structural studies, binding assays, and MD simulations are convincing.

eLife assessment

This important study combines a range of advanced ultrastructural imaging approaches to define the unusual endosomal system of African trypanosomes. Compelling images show that instead of a distinct set of compartments, the endosome of these protists comprises a continuous system of membranes with functionally distinct subdomains as defined by canonical markers of early, late and recycling endosomes. The findings suggest that the endocytic system of bloodstream stages has evolved to facilitate the extraordinarily high rates of membrane turnover needed to remove immune complexes and survive in the blood, which is of interest to anyone studying infectious diseases.

eLife assessment

This important study addresses mechanisms of feedback inhibition between planar cell polarity (PCP) protein complexes during convergent extension movements in Xenopus embryos. The authors propose an interesting model, in which non-canonical Wnt ligand stimulates transition of Dishevelled from its complex with Vangl to Frizzled, with essential roles of Vangl, Prickle and Ror in this process. The main functional observations supporting this model are convincing, but the immunoprecipitation data are incomplete and would benefit from additional clarification. With more rigorous approaches, this work will likely be of broad interest to cell and developmental biologists.

eLife assessment

In this manuscript by Xiong L et al., through TLR2, the authors have uncovered an important link between innate immune signaling and hair regeneration. The authors provide convincing evidence supporting the critical roles of TLR2 in sensing CEP levels in hair follicles, counteracting the action of BMP signaling, and facilitating the activation of HFSCs during the hair cycle and wound repair. Importantly, the authors also propose that decreased CEP production and TLR2 expression might be factors contributing to the decreased hair regeneration associated with age-related and obesity-related hair thinning and hair loss phenotypes.

eLife assessment

This work presents a fundamental meta-analysis on the causes of glucocorticoid variations in birds and mammals. It provides convincing evidence that an increase in metabolic rates increases glucocorticoid concentrations. The work will be of broad interest to animal physiologists.

eLife assessment

The biogenesis of outer membrane proteins in Gram-negative bacteria is still not fully understood, particularly substrate recognition and insertion by beta-assembly machinery. This work reports important results identifying a new amino acid sequence motif (i.e., "internal beta-signal") on outer membrane proteins recognized by this machinery. The authors carried out rigorous biochemical approaches, providing convincing evidence to support their conclusions.

eLife assessment

This important study advances our understanding of early Cambrian cnidarian paleoecology and suggests that the reconstructed ancestral feeding and respiration mechanisms predate jet-propelled swimming utilized by modern jellyfish. The work combines solid evidence of fluid and structural mechanics modeling, simulating for the first time the feeding and respiratory capacities in a microfossil (Quadrapyrgites), which in turn opens new possibilities using this approach for paleontological research. Assuming that the prior interpretations and assumptions concerning the modeled organism's soft part and skeletal anatomy are correct, the hypotheses that (1) the organism could alternately contract and expand the oral region and (2) such movement increased feeding efficiency seem plausible.

eLife assessment

This important study uses near full-length HIV-1 sequencing to examine proviral persistence in various tissues derived from three individuals who received antiretroviral therapy until time of death. Intact as well as defective HIV-1 proviruses are found at various anatomical sites including the central nervous system; the results are convincing and relevant for our understanding of latent viral reservoirs, especially in the brain.

eLife assessment

The study presented in this manuscript makes important contributions to our understanding of cell fate decisions, as well as to the role and effects of noise at various scales in gene regulatory networks involved in such fate decisions. The modelling approach and analyses provide solid support for the conclusion that distinct driving forces behind fate decisions can be distinguished by their noise profiles and reprogramming trajectories.

eLife assessment

This study examines the role of interaction between the PAS domain and the Cyclic Nucleotide-Binding Homology Domain (CNBHD) in voltage-dependent gating of EAG channels. The authors make the extraordinary claim that they have identified a hidden open state, thus providing a window for observing early conformational transitions associated with channel gating. Although the data are intriguing, the evidence supporting the conclusions is incomplete, and the experimental conditions used to propose the channel gating mechanism need to be revisited. With sufficiently strong experimental support, this work could become important for understanding the gating mechanisms of the KCNH family and would appeal to biophysicists interested in ion channels and physiologists interested in cancer biology.

eLife assessment

In this manuscript, the authors tested the hypothesis that Aβ42 toxicity arises from its proven affinity for γ-secretases. The authors provide useful findings, however, the results are incomplete and do not fit physiological conditions in the brain. The data will be of interest to all scientists working on neurodegenerative diseases.

eLife assessment

This useful manuscript presents a new therapeutic formulation and these solid findings have potential clinical significance as the efficacy of CK21 is relevant in various pancreatic cancer models.Further validation studies would help to strengthen the findings.

eLife assessment

The authors use a clever experimental design and approach to tackle an important set of questions in the field of decision-making. From this work, the authors have a number of intriguing results. However, questions remain regarding the extent to which a number of alternative models and interpretations, not considered in the paper, could account for the observed effects.

eLife assessment

This fundamental study provides compelling evidence for a new mechanism of host-microbe interaction, with indole, produced by proliferating bacteria in the C. elegans digestive system, signalling through the host via the transcription factor DAF-16 to induce the expression of genes controlling bacterial growth in the gut. The work is relevant to a wide audience as it invites deeper research into this mechanism, while also serving as a template for similar microbiome/host interactions in other systems.

eLife assessment

This important study reports biochemical and structural experiments that were carried out to determine the molecular basis of calcium-sensitive regulation of the guanine exchange factor Ric8A by the neuronal calcium sensor 1 (NCS-1). Structural and biochemical evidence for the NCS-1/Ric8A interface is convincing, but evidence for the full-length interactions is incomplete due to the low resolution of cryo-EM maps. This work will have important implications for scientists interested in G-protein signaling and molecular interactions that contribute to synapse function.

eLife assessment

This paper reports fundamental findings that substantially advance our understanding of a major research question - how hybridization events influence gene regulatory programs and how evolutionary pressures have shaped these programs in response to such events. The methods, data, and analyses are solid and broadly support the claims with only minor weaknesses. This convincing work uses appropriate and validated methodology in line with the current state-of-the-art.

eLife assessment

This is an important study that investigates the role of astrocytic Gi signaling in the anterior cingulate cortex in the modulation of extracellular L-lactate level and consequently impairment in flavor-place associates (PA) learning. However, whereas some of the behavioral observations and signaling mechanism data are compelling, the conclusions about effects on memory and the role of L-lactate are based on inadequate methods. With the addition of a few key experiments, this work will be of interest to neuroscientists studying learning and memory and the contribution of astrocytes to these processes.

eLife assessment

This important study delineates the molecular changes driving the progression from actinic keratosis (AK) to cutaneous squamous cell carcinoma (cSCC). Using state-of-the-art single-cell RNA profiling of 138,982 cells from 13 samples of six patients including AK, squamous cell carcinoma in situ (SCCIS), cSCC, and their matched normal tissues, thus covering comprehensive clinical courses of cSCC, the authors provide an invaluable data resource. This study identified several previously unreported and interesting candidate genes involved in different stages of the malignant progression of skin neoplasias, which have been validated in situ, and partially in vitro. Although data analysis needs improvement and comparison to other published data sets to fully support the claims and conclusions, these findings substantially advance our understanding of the molecular changes leading to skin cancer.

eLife assessment

Purines are native molecules that affect processes in the immune system, among others. The manuscript describes a valuable investigation of the mode of binding of purines, especially their carbohydrate moiety, to human receptors in cell culture and by computer-based modelling. Solid evidence is presented about the way purines interact with and activate two receptors.

eLife assessment

This study presents a valuable finding on an alternative method that combines multiple inhibitors to maximize on-target inhibition and minimize off-target inhibition. The evidence supporting the claims of the authors is solid, although a proper validation of the methodology could strengthen the paper. The work will be of interest to scientists working in the field of drug discovery, particularly in the field of kinase inhibitors.

eLife assessment

The authors developed a valuable mathematical model that describes the spatiotemporal dynamics of cells in the intestinal crypt. The proposed model makes an important contribution to the field, allowing a better understanding of the formation and response dynamics of the intestinal crypt through the effective evaluation of health, disease, and treatment conditions. The authors provided solid evidence of the validity of their model and their conclusions, but some minor claims are not properly justified in the current manuscript. This paper is meant for computational biologists and cancer researchers working on oncotherapies for the intestinal epithelium.

eLife assessment

This manuscript provides a useful qualitative analysis of hair extracts from mammals, with the goal of understanding how mosquitoes encode the chemicals in the odor. The data were collected using standard spectrometric approaches in the field of chemical ecology, but they suffer from artifacts associated with the sampling approach and analyses, and are limited in their replicates and sampling which may prevent inter-species comparisons at this point. Without additional analysis, the evidence supporting the claims currently remains unfortunately incomplete.

eLife assessment

This study presents a valuable comparison of the transcriptome and translatome between Drosophila neurons and glia. The data were collected and analyzed using validated methodology and the paper reports extensive variation in the translation efficiency of specific transcripts between neurons and glia. The authors argue that glia actively suppress the translation of neuronal transcripts by retaining ribosomes in their 5' leader regions. However, whereas the transcriptome and translatome analyses are convincing, the functional analyses are incomplete and would benefit from more rigorous validation to support the proposed model without over-interpretation.

eLife assessment

This study presents a useful resource for the gene expression profiles of different cell types in the parietal lobe of the cerebral cortex of prenatal macaques. The evidence supporting the claims of the authors is solid, although poor cell classification may be skewing interpretation of the results. This dataset will be of interest to developmental neurobiologists and could potentially be used for future comparative studies on early brain development.