bluuebunny/crossref_metadata_2025 huggingface dataset:
Miniature linear and split-belt treadmills reveal mechanisms of adaptive motor control in walkingDrosophila
Authors: Pratt Brandon G. , Lee Su-Yee J. , Chou Grant M. , Tuthill John C.
Published: February 2024

To navigate complex environments, walking animals must detect and overcome unexpected perturbations. One technical challenge when investigating adaptive locomotion is measuring behavioral responses to precise perturbations during naturalistic walking; another is that manipulating neural activity in sensorimotor circuits often reduces spontaneous locomotion. To overcome these obstacles, we introduce miniature treadmill systems for coercing locomotion and tracking 3D kinematics of walkingDrosophila. By systematically comparing walking in three experimental setups, we show that flies compelled to walk on the linear treadmill have similar stepping kinematics to freely walking flies, while kinematics of tethered walking flies are subtly different. Genetically silencing mechanosensory neurons alters step kinematics of flies walking on the linear treadmill across all speeds, while inter-leg coordination remains intact. We also found that flies can maintain a forward heading on a split-belt treadmill by adapting the step distance of their middle legs. Overall, these new insights demonstrate the utility of miniature treadmills for studying insect locomotion.
RinRK1 Promotes Accumulation of Nod Factor Receptors in Nanodomain-like structures at the tip of infected root hairs
Authors: Zhou Ning , Li Xiaolin , Zheng Zhiqiong , Liu Jing , Downie J. Allan , Xie Fang
Published: February 2024

Legume-rhizobia root-nodule symbioses involve the recognition of rhizobial Nod factor (NF) signals by NF receptors, triggering both nodule organogenesis and rhizobial infection.RinRK1is induced by NF signaling and is essential for infection thread (IT) formation inLotus japonicus. However, the precise mechanism underlying this process remains unknown. Here, we show that RinRK1 interacts with the extracellular domains of NF receptors NFR1 and NFR5 to promote their accumulation at root hair tips in response to rhizobia or NFs. Furthermore, Flotillin 1 (Flot1), a nanodomain-organizing protein, associates with the kinase domains of NFR1, NFR5 and RinRK1. RinRK1 promotes the interactions between Flot1 and NFR1 or NFR5 and both RinRK1 and Flot1 are necessary for the accumulation of NFR1 and NFR5 at root hair tips upon NF stimulation. Our study show that RinRK1 and Flot1 play a crucial role in NF receptor complex assembly within localized plasma membrane signaling centers to promote symbiotic infection.
High-confidence calling of normal epithelial cells allows identification of a novel stem-like cell state in the colorectal cancer microenvironment
Authors: Wei Tzu-Ting , Blanc Eric , Peidli Stefan , Bischoff Philip , Trinks Alexandra , Horst David , Sers Christine , Blüthgen Nils , Beule Dieter , Morkel Markus , Obermayer Benedikt
Published: February 2024

Single-cell analyses can be confounded by assigning unrelated groups of cells to common developmental trajectories. For instance, cancer cells and admixed normal epithelial cells could potentially adopt similar cell states thus complicating analyses of their developmental potential. Here, we develop and benchmark CCISM (for Cancer Cell Identification using Somatic Mutations) to exploit genomic single nucleotide variants for the disambiguation of cancer cells from genomically normal non-cancer epithelial cells in single-cell data. In colorectal cancer datasets, we find that our method and others based on gene expression or allelic imbalances identify overlapping sets of cancer versus normal epithelial cells, depending on molecular characteristics of individual cancers. Further, we define consensus cell identities of normal and cancer epithelial cells with higher transcriptome cluster homogeneity than those derived using existing tools. Using the consensus identities, we identify significant shifts of cell state distributions in genomically normal epithelial cells developing in the cancer microenvironment, with immature states increased at the expense of terminal differentiation throughout the colon, and a novel stem-like cell state arising in the left colon. Trajectory analyses show that the new cell state extends the pseudo-time range of normal colon stem-like cells in a cancer context. We identify cancer-associated fibroblasts as sources of WNT and BMP ligands potentially contributing to increased plasticity of stem cells in the cancer microenvironment. Our analyses advocate careful interpretation of cell heterogeneity and plasticity in the cancer context and the consideration of genomic information in addition to gene expression data when possible.

Novelty and Impact

Single-cell analyses have become standard to assess cell heterogeneity and developmental hierarchies in cancer tissues. However, these datasets are complex and contain cancer and non-cancer lineage cells. Here, we develop and systematically benchmark tools to distinguish between cancer and non-cancer single-cell transcriptomes, based on gene expression or different levels of genomic information. We provide strategies to combine results of different tools into consensus calls tailored to the biology and genetic characteristics of the individual cancer.
Role of PLK1 in the epigenetic maintenance of centromeres
Authors: Conti Duccio , Verza Arianna Esposito , Pesenti Marion E. , Cmentowski Verena , Vetter Ingrid R. , Pan Dongqing , Musacchio Andrea
Published: February 2024

The centromere, a chromosome locus defined by the histone H3-like protein CENP-A, seeds the kinetochore to bind microtubules during cell division. Centromere maintenance requires CENP-A to be actively replenished by dedicated protein machinery in the early G1 cell-cycle phase, compensating for its two-fold dilution following DNA replication. Cyclin-dependent kinases (CDKs) limit CENP-A deposition to once per cell cycle and function as negative regulators outside early G1. Antithetically, Polo-like kinase 1 (PLK1) promotes CENP-A deposition in early G1, but the molecular details are still unknown. We reveal a phosphorylation network that recruits PLK1 to the deposition machinery to control a conformational switch required for licensing the CENP-A deposition reaction. Our findings solve the long-standing question of how PLK1 contributes to the epigenetic maintenance of centromeres.

One-Sentence Summary

PLK1 licenses epigenetic maintenance of centromeres by regulating a conformational switch on the MIS18α protein.
Memory load influences our preparedness to act on visual representations in working memory without affecting their accessibility
Authors: Nasrawi Rose , Mautner-Rohde Mika , van Ede Freek
Published: February 2024

It is well established that when we hold more content in working memory, we are slower to act upon specific content when it becomes relevant for behavior. Here, we asked whether slower responses with higher working-memory load can be accounted for by slower access to sensory representations held in working memory (according to a serial internal search), or by a lower preparedness to act upon them. To address this, we designed a visual-motor working-memory task in which participants memorized the orientation of two or four colored bars, of which one was cued for reproduction. We independently tracked the selection of visual (cued item location) and motor (relevant manual action) information from the EEG time-frequency signal, and compared their latencies between our memory-load conditions. We confirm slower memory-guided behavior with higher working-memory load and show that this is associated with delayed motor selection, but find no evidence for a concomitant delay in the latency of visual selection. Moreover, we show that variability in decision times within each memory-load condition is associated with corresponding changes in the latency of motor, but not visual selection. These results reveal how memory load affects our preparedness to act on, without necessarily changing the ability to access, sensory representations in working memory. This posits action readiness is a key factor that shapes the speed of memory-guided behavior and that underlies delayed responses with higher working-memory load.
Impacts of a prolonged marine heatwave and chronic local human disturbance on juvenile coral assemblages
Authors: Tietjen Kristina L. , Perks Nelson F. , O’Brien Niallan C. , Baum Julia K.
Published: February 2024

Coral reefs are threatened by climate change and chronic local human disturbances. Although some laboratory studies have investigated the effects of combined stressors, such as nutrient enrichment and heat stress, on growth and survival of early life stage corals,in situstudies remain limited. To assess the influence of multiple stressors on juvenile corals, we quantified densities of corals ≤5 cm at 18 forereef sites with different exposure levels to underlying chronic local human disturbance before, during, and after the 2015-2016 El Niño. This marine heatwave caused prolonged heat stress and devastating losses of coral cover at our study site, Kiritimati, in the central equatorial Pacific Ocean. In total, we enumerated 7732 juvenile corals from 13 different families. Over 80% of corals were from four families: 70% from Agariciidae, Merulinidae, or Poritidae, which all have stress-tolerant life history strategies, and 11% from Acroporidae which has a competitive life-history strategy. Both local disturbance and heat stress were significantly negatively related to juvenile coral densities. Prior to the heatwave, juvenile densities were on average 72% lower at the most disturbed sites (7.2 ± 1.9 m2) compared to the least disturbed ones (15.3 ± 3.8 m2). Overall, juvenile corals had a lower bleaching prevalence and lower mortality during the heatwave when compared to their adult counterparts. Still, the heatwave resulted in the loss of half (50%) of all juvenile corals, with competitive and weedy life history strategy corals undergoing greater declines than stress-tolerant ones. Although juvenile coral densities increased slightly in the year following the heatwave, the effect was statistically non-significant. Our results highlight the influence of local chronic anthropogenic and climate change-driven marine heatwaves on juvenile coral densities.
Mammalian Cell Models of the Distal Convoluted Tubule: A Systematic Review of Cell Lines, Culture Condition and Gene Expression
Authors: Zhong Chutong , Sun Zhen , Grillo Alessandra , Walsh Stephen B , Siew Keith
Published: February 2024

This review examines the crucial role of human cellular models in renal physiology research, with a specific focus on the distal convoluted tubule (DCT). It aims to provide a comprehensive summary of the origins, culture practices, and genetic studies associated with commonly employed DCT cell models. To achieve this, a systematic literature review was performed on Europe PMC, employing a Boolean search strategy. A total of 6,559 articles were initially screened, resulting in 301 articles on human-origin cell models, 69 on murine models, and 29 on canine models being included in the final analysis. Notably, the review identified two studies that introduced novel immortalised human DCT cell lines developed from primary DCT cells. This paper provides a detailed account of the lineage of each cell model, their prevalent culture conditions, and the frequency and nature of gene transfections—both wild type and mutant—conducted in these models. A significant observation from the analysis was the inconsistent reporting of methodological details across studies, which compromises the reproducibility of the research. Additionally, there was a considerable variation in culture conditions and transfection methods used across different studies. The review also highlights that HEK293 family and murine DCT cell lines do not serve as accurate models for DCT cells, pointing out the frequent need for transfecting DCT-specific genes to simulate DCT functionality adequately.

NEW & NOTEWORTHY

This review is the first review to detail the current state of play with cell models of the distal convoluted tubule, including endogenous protein expression, culture conditions etc. We expect this review to be of great utility to established and early career researchers who are interested in nephrological or hypertensive patho/physiology. We have highlighted some important gaps in reporting the established cell models, we think that this kind of review is important for open science.
Evaluation of Bayesian Linear Regression Derived Gene Set Test Methods
Authors: Bai Zhonghao , Gholipourshahraki Tahereh , Shrestha Merina , Hjelholt Astrid , Kjølby Mads , Rohde Palle Duun , Sørensen Peter
Published: February 2024

Gene set tests can pinpoint genes and biological pathways that exert small to moderate effects on complex diseases like Type 2 Diabetes (T2D). By aggregating genetic markers based on biological information, these tests can enhance the statistical power needed to detect genetic associations. Our goal was to develop a gene set test utilizing Bayesian Linear Regression (BLR) models, which account for both linkage disequilibrium (LD) and the complex genetic architectures intrinsic to diseases, thereby increasing the detection power of genetic associations. Through a series of simulation studies, we demonstrated how the efficacy of BLR derived gene set tests is influenced by several factors, including the proportion of causal markers, the size of gene sets, the percentage of genetic variance explained by the gene set, and the genetic architecture of the traits. Comparing our method with other approaches, such as the gold standard MAGMA (Multi-marker Analysis of Genomic Annotation) approach, our BLR gene set test showed superior performance. This suggests that our BLR-based approach could more accurately identify genes and biological pathways underlying complex diseases.
Structural Determinants for Activity of the Antidepressant Vortioxetine at Human and Rodent 5-HT3receptors
Authors: López-Sánchez Uriel , Munro Lachlan Jake , Ladefoged Lucy Kate , Pedersen Anders Juel , Nielsen Christian C. , Lyngby Signe M. , Baud Delphine , Lummis Sarah C. R. , Bang-Andersen Benny , Schiøtt Birgit , Chipot Christophe , Schoehn Guy , Neyton Jacques , Dehez Francois , Nury Hugues , Kristensen Anders S.
Published: February 2024

Vortioxetine (VTX) is a recent antidepressant that targets a variety of serotonin receptors. We investigate the drug’s molecular mechanism of operation at serotonin 5-HT3receptors (5-HT3R), which features two mysterious properties: VTX acts differently on rodent and human 5-HT3R; VTX appears to suppress any subsequent response to agonists. Using a combination of cryo-EM, electrophysiology, and molecular dynamics, we show that VTX stabilizes a resting inhibited state of the mouse 5-HT3R and an agonist bound-like state of the human 5-HT3R, in line with the functional profile of the drug. We report four human 5-HT3R structures and show that the human receptor transmembrane domain is intrinsically fragile. We also explain the lack of recovery after VTX administration via a membrane partition mechanism.
Design, optimization, and inference of biphasic decay of infectious virus particles
Authors: Seurat Jérémy , Gerbino Krista R. , Meyer Justin R. , Borin Joshua M. , Weitz Joshua S.
Published: February 2024

Virus population dynamics are driven by counter-balancing forces of production and loss. Whereas viral production arises from complex interactions with susceptible hosts, the loss of infectious virus particles is often approximated as a first-order kinetic process. As such, experimental protocols to measure infectious virus loss are not typically designed to identify non-exponential decay processes. Here, we propose methods to evaluate if an experimental design is adequate to identify multiphasic virus particle decay and to optimize the sampling times of decay experiments, accounting for uncertainties in viral kinetics. First, we evaluate synthetic scenarios of biphasic decays, with varying decay rates and initial proportions of subpopulations. We show that robust inference of multiphasic decay is more likely when the faster decaying subpopulation predominates insofar as early samples are taken to resolve the faster decay rate. Moreover, design optimization involving non-equal spacing between observations increases the precision of estimation while reducing the number of samples. We then apply these methods to infer multiple decay rates associated with the decay of bacteriophage (‘phage’) ΦD9, an evolved isolate derived from phage Φ21. A pilot experiment confirmed that ΦD9 decay is multiphasic, but was unable to resolve the rate or proportion of the fast decaying subpopulation(s). We then applied a Fisher information matrix-based design optimization method to propose nonequally spaced sampling times. Using this strategy, we were able to robustly estimate multiple decay rates and the size of the respective subpopulations. Notably, we conclude that the vast majority (94%) of the phage ΦD9 population decays at a rate 16-fold higher than the slow decaying population. Altogether, these results provide both a rationale and a practical approach to quantitatively estimate heterogeneity in viral decay.
C. elegansdisplay antipathy behavior towards food after contemporaneous integration of nutritional needs and dietary lipid availability
Authors: Stuhr Nicole L. , Ramos Carmen M. , Turner Chris D. , Soukas Alexander A. , Curran Sean P.
Published: February 2024

Organisms utilize sophisticated neurocircuitry to select optimal food sources within their environment.Methylobacteriumis a lifespan-promoting bacterial diet forC. elegansthat drives faster development and longevity, however after ingestion,C. elegansconsistently choose any other food option available. A screen for genetic regulators of the avoidance behavior towardMethylobacteriumidentified the AWB and AWC sensory neurons and theodr-1guanylate cyclase expressed exclusively in those four ciliated neurons as mediators of the antipathy response. Metabolic profiling of theMethylobacteriumdiet reveals a macromolecular profile enriched in saturated fats and here we show thatC. eleganssense and integrate signals related to the type of ingested lipids that subsequently cues food-related behaviors. Moreover, disruption of endogenous lipid metabolism modifies the intensity of antipathy towardMethylobacteriumwhich suggests that the current state of lipid homeostasis influences food preference. Enhanced expression of the sphingolipid degradation enzyme Saposin/spp-9enhances antipathy behaviors and activation of the sphingosine rheostat and more specifically modulation of the bioactive lipid mediator sphingosine-1-phosphate (S1P) acts as a signal to promote avoidance ofMethylobacterium. Taken together, our work reveals thatC. elegansmodify food choices contemporaneously based on the availability of dietary lipids and the ability to metabolize dietary lipids.

HIGHLIGHTS

Uncover new molecular mechanisms underlying the decision matrix an animal uses to choose what foods to eat.

Define the molecular mechanisms underlying an antipathy behavioral response toward foods after initial ingestion that contemporaneously integrates dietary needs with nutritional profile.

ODR-1 signaling from AWB and AWC ciliated neurons of theC. elegansnervous system mediate the antipathy response to diet.

Manipulation of sphingosine-1-phosphate (S1P) of the sphingosine rheostat controls the intensity of the antipathy behavioral response.

Modulating antipathy behaviors can impact the magnitude of the lifespan-promoting effects of longevity diets.
Park7deletion leads to age- and sex-specific transcriptome changes involving NRF2-CYP1B1 axis in mouse midbrain astrocytes
Authors: Helgueta Sergio , Heurtaux Tony , Sciortino Alessia , Gui Yujuan , Ohnmacht Jochen , Mencke Pauline , Boussaad Ibrahim , Halder Rashi , Garcia Pierre , Krüger Rejko , Mittelbronn Michel , Buttini Manuel , Sauter Thomas , Sinkkonen Lasse
Published: February 2024

Abstractdispe

Loss-of-function mutations inPARK7, encoding for DJ-1, can lead to early onset Parkinson’s disease (PD). In mice,Park7deletion leads to dopaminergic deficits during aging, and increased sensitivity to oxidative stress. However, the severity of the reported phenotypes varies. To understand the early molecular changes upon loss of DJ-1, we performed transcriptomic profiling of midbrain sections from young mice. Interestingly, while at 3 months the transcriptomes of both male and female mice were unchanged compared to their wildtype littermates, an extensive deregulation was observed specifically in 8-month-old males. The affected genes are involved in processes such as focal adhesion, extracellular matrix interaction, and epithelial-to-mesenchymal transition (EMT), and enriched for primary target genes of Nuclear factor erythroid 2-related factor 2 (NRF2). Consistently, the antioxidant response was altered specifically in the midbrain of male DJ-1 deficient mice. Many of the misregulated genes are known target genes of estrogen and retinoic acid signaling and show sex-specific expression in wildtype mice. Depletion of DJ-1 or NRF2 in male, but not female primary astrocytes recapitulated many of thein vivochanges, including downregulation of cytochrome P450 family 1 subfamily B member 1 (CYP1B1), an enzyme involved in estrogen and retinoic acid metabolism. Interestingly, knock-down of CYP1B1 led to gene expression changes in focal adhesion and EMT in primary male astrocytes. Finally, male iPSC-derived astrocytes with loss of function mutation in thePARK7gene also showed changes in the EMT pathway and NRF2 target genes. Taken together, our data indicate that loss ofPark7leads to sex-specific gene expression changes specifically in males through astrocytic alterations in the NRF2-CYP1B1 axis. These findings suggest higher sensitivity of males to loss of DJ-1 and might help to better understand variation in the reportedPark7−/−phenotypes.
Gene Expression Reactomes Across Species Do Not Correlate with Gene Structural Similarity
Authors: Chen Wei , Chai Yi , Cui Nancy Y. , Gopinath Ashwin , Zhang David Yu
Published: February 2024

Mammalian animal model species for preclinical drug testing share a large number of genes with similar names and/or protein primary structures, but gene expression patterns in different tissues and in response to different drugs have not been previously systematically characterized. Here, we experimentally measure the gene expression responses (reactomes) of each gene in the mouse and rat genomes in 10 different organs to 3 different drugs. Surprisingly, we observe that gene reactomes across species generally do not correlate with structural similarity. Thus, we propose an alternative functional cross-species gene mapping approach, based on organ-specific gene expression response to drug dosing.
BET Degraders Reveal BRD4 Disruption of 7SK and P-TEFb is Critical for Effective Reactivation of Latent HIV in CD4+ T-cells
Authors: Turner Anne-Marie W. , Bashore Frances M. , Falcinelli Shane D. , Fox Joshua A. , Keller Alana L. , Fenton Anthony D. , Geyer Renee F. , Allard Brigitte , Kirchherr Jennifer L. , Archin Nancie M. , James Lindsey I. , Margolis David M.
Published: February 2024

HIV cure strategies that aim to induce viral reactivation for immune clearance leverage latency reversal agents to modulate host pathways which directly or indirectly facilitate viral reactivation. Inhibition of BET (bromo and extra-terminal domain) family member BRD4 reverses HIV latency, but enthusiasm for the use of BET inhibitors in HIV cure studies is tempered by concerns over inhibition of other BET family members and dose-limiting toxicities in oncology trials. Here we evaluated the potential for bivalent chemical degraders targeted to the BET family as alternative latency reversal agents. We observed that despite highly potent and selective BRD4 degradation in primary CD4+ T-cells from ART-suppressed donors, BRD4 degraders failed to induce latency reversal as compared to BET inhibitors. Further, BRD4 degraders failed to mimic previously observed synergistic HIV reactivation between BET inhibitors and an activator of the non-canonical NF-κB pathway. Mechanistic investigation of this discrepancy revealed that latency reversal by BET inhibitors is not related to the abatement of competition between Tat and BRD4 for P-TEFb, but rather the ability of BRD4 to disrupt 7SK and increase the levels of free P-TEFb. This activity is dependent on the shift of BRD4 from chromatin-bound to soluble and retargeting of P-TEFb to chromatin which is dependent on intact BRD4 but independent of the bromodomains.
An uncommon phosphorylation mode regulates the activity and protein-interactions of N-acetylglucosamine kinase
Authors: Celik Arif , Beyer Ida , Fiedler Dorothea
Published: February 2024

While the function of protein phosphorylation in eukaryotic cell signaling is well established, the role of a closely related modification, protein pyrophosphorylation, is just starting to surface. A recent study has identified several targets of endogenous protein pyrophosphorylation in mammalian cell lines, including N-acetylglucosamine kinase (NAGK). Here, a detailed functional analysis of NAGK phosphorylation and pyrophosphorylation on serine 76 (S76) has been conducted. This analysis was enabled by using amber codon suppression to obtain phosphorylated pS76-NAGK, which was subsequently converted to site-specifically pyrophosphorylated NAGK (ppS76-NAGK) with a phosphorimidazolide regent. A significant reduction in GlcNAc kinase activity was observed upon phosphorylation, and near-complete inactivation upon pyrophosphorylation. The formation of ppS76-NAGK proceededviaan ATP-dependent autocatalytic process, and once formed, ppS76-NAGK displayed notable stability towards dephosphorylation in mammalian cell lysates. Proteomic examination unveiled a distinct set of protein-protein interactions for ppS76-NAGK, suggesting an alternative function, independent of its kinase activity. Overall, a significant regulatory role of pyrophosphorylation on NAGK activity was uncovered, providing a strong incentive to investigate the influence of this unusual phosphorylation mode on other kinases.
Epigenetic Control of Adaptive or Homeostatic Splicing of Alternative Exons andProlactinGene Expression During Interval-Training Activities of Pituitary Cells
Authors: Liu Ling , Nguyen Hai , Das Urmi , Ogunsola Samuel , Yu Jiankun , Lei Lei , Kung Matthew , Pejhan Shervin , Rastegar Mojgan , Xie Jiuyong
Published: February 2024

Interval-training activities induce adaptive cellular changes without altering their fundamental identity, but the precise underlying molecular mechanisms are not fully understood. In this study, we demonstrate that interval-training depolarization (ITD) of pituitary cells triggers distinct adaptive or homeostatic splicing responses of alternative exons. This occurs while preserving the steady-state expression of theProlactinand other hormone genes. The nature of these splicing responses depends on the exon’s DNA methylation status, the methyl-C-binding protein MeCP2 and its associated CA-rich motif-binding hnRNP L. Interestingly, the steady expression of theProlactingene is also reliant on MeCP2, whose disruption during ITD leads to exacerbated overexpression and multi-exon aberrant splicing of the hormone gene transcripts, similar to the observed hyperprolactinemia or activity-dependent aberrant splicing in Rett Syndrome. Therefore, depending on how many times cells are stimulated, exons may exhibit different splicing responses to cell activities. During the ITD, epigenetic control is crucial for both adaptive or homeostatic splicing and the steady expression of theProlactinhormone gene. Disruption in this regulation may have significant implications for the development of progressive diseases.
Asynchronous abundance fluctuations can drive giant genotype frequency fluctuations
Authors: Ascensao Joao A , Lok Kristen , Hallatschek Oskar
Published: February 2024

Large stochastic population abundance fluctuations are ubiquitous across the tree of life1–7, impacting the predictability of population dynamics and influencing eco-evolutionary outcomes. It has generally been thought that these large abundance fluctuations do not strongly impact evolution, as the relative frequencies of alleles in the population will be unaffected if the abundance of all alleles fluctuate in unison. However, we argue that large abundance fluctuations can lead to significant genotype frequency fluctuations if different genotypes within a population experience these fluctuations asynchronously. By serially diluting mixtures of two closely relatedE. colistrains, we show that such asynchrony can occur, leading to giant frequency fluctuations that far exceed expectations from models of genetic drift. We develop a flexible, effective model that explains the abundance fluctuations as arising from correlated offspring numbers between individuals, and the large frequency fluctuations result from even slight decoupling in offspring numbers between genotypes. This model accurately describes the observed abundance and frequency fluctuation scaling behaviors. Our findings suggest chaotic dynamics underpin these giant fluctuations, causing initially similar trajectories to diverge exponentially; subtle environmental changes can be magnified, leading to batch correlations in identical growth conditions. Furthermore, we present evidence that such decoupling noise is also present in mixed-genotypeS. cerevisiaepopulations. We demonstrate that such decoupling noise can strongly influence evolutionary outcomes, in a manner distinct from genetic drift. Given the generic nature of asynchronous fluctuations, we anticipate that they are widespread in biological populations, significantly affecting evolutionary and ecological dynamics.
New Dual Inducible Cellular Model to Investigate Temporal Control of Oncogenic Cooperating Genes
Authors: Kent Matthew R. , Jay Amanda N. , Kendall Genevieve C.
Published: February 2024

The study of cooperating genes in cancer can lead to mechanistic understanding and identifying potential therapeutic targets. To facilitate these types of studies, we developed a new dual-inducible system utilizing the tetracycline- and cumate-inducible systems driving HES3 and the PAX3::FOXO1 fusion-oncogene, respectively, as cooperating genes from fusion-positive rhabdomyosarcoma. With this new model, we can independently induce expression of either HES3 or PAX3::FOXO1, as well as simultaneously induce expression of both genes. This new model will allow us to further investigate the cooperation between HES3 and PAX3::FOXO1 including the temporal requirements for genetic cooperation. This dual-inducible model can be adapted for any cooperating genes, allowing for independent, simultaneous, or temporally controlled gene expression.
CACTI: An in-silicodrug-target prediction tool through the integration of chemogenomic data and clustering analysis
Authors: Godinez-Macias Karla P. , Winzeler Elizabeth A.
Published: February 2024

It is well-accepted that knowledge of a small molecule’s target can accelerate optimization. Although chemogenomic databases are helpful resources for predicting or finding compound interaction partners, they tend to be limited and poorly annotated. Furthermore, unlike genes, compound identifiers are often not standardized, and many synonyms may exist, especially in the biological literature, making batch analysis of compounds difficult. Here, we constructed an open-source annotation and target prediction tool that explores some of the largest chemical and biological databases, mining these for both common name, synonyms, and structurally similar molecules. We used this Chemical Analysis and Clustering for Target Identification (CACTI) tool to analyze the Pathogen Box collection, an open-source set of 400 drug-like compounds active against a variety of microbial pathogens. Our analysis resulted in 4,315 new synonyms, 35,963 pieces of new information and target prediction hints for 58 members.

Scientific Contribution

With the employment of this tool, a comprehensive report with known evidence, close analogs and drug-target prediction can be obtained for large-scale chemical libraries that will facilitate their evaluation and future target validation and optimization efforts.
Topographical changes in extracellular matrix during skin fibrosis and recovery can be evaluated using automated image analysis algorithms
Authors: Wyetzner Rachel H. , Segal Ella X. , Jussila Anna R. , Atit Radhika P.
Published: February 2024

Skin fibrosis is characterized by fibroblast activation and intradermal fat loss, resulting in excess deposition and remodeling of dermal extracellular matrix (ECM). The topography of the dominant ECM proteins, such as collagens, can indicate skin stiffness and remains understudied in evaluating fibrotic skin. Here, we adapted two different unbiased image analysis algorithms to define collagen topography and alignment in a genetically inducible and reversible Wnt activation fibrosis model. We demonstrated that Wnt activated fibrotic skin has altered collagen fiber characteristics and a loss of collagen alignment, which were restored in the reversible model. This study highlights how unbiased algorithms can be used to analyze ECM topography, providing novel avenues to evaluate fibrotic skin onset, recovery, and treatment.
KSR1 mediates small-cell lung carcinoma tumor initiation and cisplatin resistance
Authors: Chatterjee Deepan , Svoboda Robert A. , Huisman Dianna H. , Drapkin Benjamin J , Vieira Heidi M. , Rao Chaitra , Askew James W. , Fisher Kurt W. , Lewis Robert E.
Published: February 2024

Small-cell lung cancer (SCLC) has a dismal five-year survival rate of less than 7%, with limited advances in first line treatment over the past four decades. Tumor-initiating cells (TICs) contribute to resistance and relapse, a major impediment to SCLC treatment. Here, we identify Kinase Suppressor of Ras 1 (KSR1), a molecular scaffold for the Raf/MEK/ERK signaling cascade, as a critical regulator of SCLC TIC formation and tumor initiationin vivo. We further show that KSR1 mediates cisplatin resistance in SCLC. While 50-70% of control cells show resistance after 6-week exposure to cisplatin, CRISPR/Cas9-mediated KSR1 knockout prevents resistance in >90% of SCLC cells in ASCL1, NeuroD1, and POU2F3 subtypes. KSR1 KO significantly enhances the ability of cisplatin to decrease SCLC TICs viain vitroextreme limiting dilution analysis (ELDA), indicating that KSR1 disruption enhances the cisplatin toxicity of cells responsible for therapeutic resistance and tumor initiation. The ability of KSR1 disruption to prevent cisplatin resistant in H82 tumor xenograft formation supports this conclusion. Previous studies indicate that ERK activation inhibits SCLC tumor growth and development. We observe a minimal effect of pharmacological ERK inhibition on cisplatin resistance and no impact on TIC formation viain vitroELDA. However, mutational analysis of the KSR1 DEF domain, which mediates interaction with ERK, suggests that ERK interaction with KSR1 is essential for KSR1-driven cisplatin resistance. These findings reveal KSR1 as a key regulatory protein in SCLC biology and a potential therapeutic target across multiple SCLC subtypes.

Statement of Implication

Genetic manipulation of the molecular scaffold KSR1 in small-cell lung cancer cells reveals its contribution to cisplatin resistance and tumor initiation.
GhCPK28 negatively regulates the immune response by phosphorylating GhTIFY3b
Authors: Gao Shengqi , Shao Wukui , Wu Jiawen , Zhao Zhun , Hu Wenran , Shao Panxia , Hua Jian , Zou Baohong , Huang Quansheng
Published: February 2024

The soil-borne fungusVerticillium dahliae(V. dahliae) seriously inhibits cotton (Gossypium hirsutum) growth and productivity. The immune system of cotton against this pathogen is largely unknown. Here we investigated the involvement of Ca2+-dependent protein kinases (CPKs) in this immunity. One CPK coding geneGhCPK28had reduced expression after infection byV. dahliaein cotton. Knocking down of theGhCPK28by virus induced gene silencing led to enhanced resistance toV. dahliaewhich is accompanied by a higher ROS accumulation and systemic acquired response. GhCPK28 was found to phosphorylate a TIFY family protein GhTIFY3b and reduce its stability. SilencingGhTIFY3bincreased the susceptibility of cotton toV. dahliae. Together, this study indicate that GhCPK28 is a negative regulator of defense againstV. dahliaeinfection and the GhTIFY3b might be a target protein of its activity. It sheds light on the immune system againstV. dahliaeand provides candidate genes for improving resistance against Verticillium wilt in cotton.
PRENATAL STRESS MODIFIES SPATIAL COGNITION IN MICE: EFFECTS OF AGE AND SEX
Authors: McCarthy M , Buck TG , Sodhi MS
Published: February 2024

Prenatal stress is linked with neuropathology of the cortical-hippocampal circuit, due to abnormal brain development that leads to long term neurological deficits in offspring, in both rodents and in humans with psychiatric disorders. Conflicting reports exist of the effects of prenatal stress in C57BL/6J mice, which is an inbred strain that is the most frequently used for neurobehavioral research. We now present comprehensive analyses of the effects of prenatal stress on spatial cognition and related behaviors in this inbred strain, in males and females at young adult and aged adult stages. The prenatal stress exposure was conducted by exposing pregnant mice to restraint stress three times daily from day 7 to day 18 of gestation. We tested the effects of prenatal stress on cognitive behavior using a battery of behavioral assays including the open-field test, the light-dark box, Morris water maze spatial acquisition and spatial reversal testing, in addition to assays of social interaction and social memory. We compared the behavioral phenotype of male and female offspring in young adult (10-20 weeks) and older adult ages (60- 70 weeks). Data reveal that male but not female mice had reduced body weight throughout the lifespan after exposure to prenatal stress. Young prenatally stressed adult females showed greater organization of tracking behavior in spatial acquisition tests than other groups. Prenatal stress improved reversal learning in aged females relative to non-stressed aged females. We detected decreased extinction of the spatial acquisition memory in prenatally stressed young adult mice of both sexes. Overall results indicate that prenatal stress may protect females from detrimental effects of age on function of the hippocampus. Our data support the finding that C57BL/6J are a relatively resilient strain of mouse that may be useful for investigating the differences between resilience and susceptibility to stress. Genomic differences between the C57BL/6J strain relative to Swiss Webster strain, which is more susceptible to prenatal stress are discussed. Future studies of prenatal stress in the C57BL/6J mouse strain should focus on the identification of molecular pathways that underlie resilience to prenatal stress, to identify novel targets for drug development.
Synthetic type III-E CRISPR-Cas effectors for programmable RNA-targeting
Authors: Brogan Daniel J. , Benetta Elena Dalla , Wang Tianqi , Lin Calvin P. , Chen Fangying , Li Harry , Lin Claire , Komives Elizabeth A. , Akbari Omar S.
Published: February 2024

The recent discovery of the type III-E CRISPR-Cas effector class has reshaped our fundamental understanding of CRISPR-Cas evolution and classification. Type III-E effectors are composed of several Cas7-like domains and a single Cas11-like domain naturally fused together to create a single polypeptide capable of programmably targeting and degrading RNA. Here we identify a novel composition of a type III-E-like effector composed of Cas7-like and a Cas1-like domain, that can be engineered into an active chimeric RNA-targeting Cas effector and presents a new function of Cas1 in RNA-targeting. Furthermore, we demonstrate a unique modularity of type III-E effectors by methodically substituting domains between orthogonal type III-E proteins to engineer compact synthetic Cas effectors. We refine our methods to generate several compact effectors for programmable RNA-targeting in mammalian cells. Cas7-S represents a new understanding of type III-E architecture and modularity, and provides a platform for engineering genome engineering technologies from the blueprint of nature.
Exploring the repository ofde novodesigned bifunctional antimicrobial peptides through deep learning
Authors: Dong Ruihan , Liu Rongrong , Liu Ziyu , Liu Yangang , Zhao Gaomei , Li Honglei , Hou Shiyuan , Ma Xiaohan , Kang Huarui , Liu Jing , Guo Fei , Zhao Ping , Wang Junping , Wang Cheng , Wu Xingan , Ye Sheng , Zhu Cheng
Published: February 2024

Antimicrobial peptides (AMPs) are attractive candidates to combat antibiotic resistance for their capability to target bio-membranes and restrict a wide range of pathogens. It is a daunting challenge to discover novel AMPs due to their sparse distributions in a vast peptide universe, especially for peptides that demonstrate potencies for both bacterial membranes and viral envelopes. Here we establish ade novoAMP design framework by bridging a deep generative module and a graph-encoding activity regressor. The generative module learns hidden ‘grammars’ of AMP features and produces candidates sequentially pass antimicrobial predictor and antiviral classifiers. We discover 16 bifunctional AMPs and experimentally validated their abilities to inhibit a spectrum of pathogensin vitroand in animal models. Notably, P076 is a highly potent bactericide with the minimal inhibitory concentration of 0.21 μM against multidrug-resistantA. baumannii, while P002 broadly inhibits five enveloped viruses. Our study provides feasible means to uncover sequences that simultaneously encode antimicrobial and antiviral activities, thus bolstering the function spectra of AMPs to combat a wide range of drug-resistant infections.
Deep Learning Behavioral Phenotyping System in the Diagnosis of Parkinson’s Disease with Drosophila melanogaster
Authors: Dong Keyi , Burch April , Huang Kang
Published: February 2024

Drosophila Melanogasteris widely used as animal models for Parkinson’s disease (PD) research. Because of the complexity of MoCap and quantitative assessment amongDrosophila Melanogaster, however, there is a technical issue that identify PD symptoms within drosophila based on objective spontaneous behavioral characteristics. Here, we developed a deep learning framework generated from kinematic features of body posture and motion between wildtype and SNCAE46Kmutant drosophila genetically modeled □-Syn, supporting clustering and classification of PD individuals. We record locomotor activity in a 3D-printed trap, and utilize the pre-analysis pose estimation software DeepLabCut (DLC) to calculate and generate numerical data representing the motion speed, tremor frequency, and limb motion ofDrosophila Melanogaster. By plugging these data as the input, the diagnosis result (1/0) representing PD or WT as the output. Our result provides a toolbox which would be valuable in the investigation of PD progressing and pharmacotherapeutic drug development.
Decoding Stability and Epistasis in Human Myoglobin by Deep Mutational Scanning and Codon-level Machine Learning
Authors: Küng Christoph , Protsenko Olena , Vanella Rosario , Nash Michael A.
Published: February 2024

Understanding the linkage between protein sequence and phenotypic expression level is crucial in biotechnology. Machine learning algorithms trained with deep mutational scanning (DMS) data have significant potential to improve this understanding and accelerate protein engineering campaigns. However, most machine learning (ML) approaches in this domain do not directly address effects of synonymous codons or positional epistasis on predicted expression levels. Here we used yeast surface display, deep mutational scanning, and next-generation DNA sequencing to quantify the expression fitness landscape of human myoglobin and train ML models to predict epistasis of double codon mutants. When fed with near comprehensive single mutant DMS data, our algorithm computed expression fitness values for double codon mutants using ML-predicted epistasis as an intermediate parameter. We next deployed this predictive model to screen > 3·106unseen double codon mutantsin silicoand experimentally tested highly ranked candidate sequences, finding 14 of 16 with significantly enhanced expression levels. Our experimental DMS dataset combined with codon level epistasis-based ML constitutes an effective method for bootstrapping fitness predictions of high order mutational variants using experimental data from variants of lower order.
Exploring the conformational ensembles of protein-protein complex with transformer-based generative model
Authors: Wang Jianmin , Wang Xun , Chu Yanyi , Li Chunyan , Li Xue , Meng Xiangyu , Fang Yitian , No Kyoung Tai , Mao Jiashun , Zeng Xiangxiang
Published: February 2024

Protein-protein interactions are the basis of many protein functions, and understanding the contact and conformational changes of protein-protein interactions is crucial for linking protein structure to biological function. Although difficult to detect experimentally, molecular dynamics (MD) simulations are widely used to study the conformational ensembles and dynamics of protein-protein complexes, but there are significant limitations in sampling efficiency and computational costs. In this study, a generative neural network was trained on protein-protein complex conformations obtained from molecular simulations to directly generate novel conformations with physical realism. We demonstrated the use of a deep learning model based on the transformer architecture to explore the conformational ensembles of protein-protein complexes through MD simulations. The results showed that the learned latent space can be used to generate unsampled conformations of protein-protein complexes for obtaining new conformations complementing pre-existing ones, which can be used as an exploratory tool for the analysis and enhancement of molecular simulations of protein-protein complexes.
Rapid spread of a vertically transmitted symbiont induces drastic shifts in butterfly sex ratio
Authors: Miyata Mai , Nomura Masashi , Kageyama Daisuke
Published: February 2024

BRIEF SUMMARY

Sex ratio dynamics constitutes a pivotal subject in evolutionary biology1. Under conditions of evolutionary equilibrium, the male-to-female ratio tends to be approximately 1:1; however, this equilibrium is susceptible to distortion by selfish genetic elements exemplified by driving sex chromosomes and cytoplasmic elements2,3. While previous studies have substantiated instances of these genetic elements distorting the sex ratio, studies specifically tracking the process with which these distorters spread within populations, leading to a transition from balanced parity to a skewed, female-biased state, are notably lacking. Herein, we present compelling substantiation regarding the rapid spread of the cytoplasmic endosymbiontWolbachiawithin a localized population of the pierid butterflyEurema hecabe(Figure 1A). This resulted in a shift in the sex ratio from near parity to an exceedingly skewed state overwhelmingly biased toward females, reaching 94.4% within a remarkably brief period of 4 years.