Biology subjects

Wallace, A.

Publications and source records attributed to Wallace, A..

16 recordsLinked to original sources

bioRxiv2026

Sex-Specific Regulation of Endothelial-to-Mesenchymal Transition (EndMT) During Atherosclerosis

Sex differences in atherosclerotic plaque biology underlie clinically distinct manifestations of acute coronary syndromes, yet the molecular mechanisms driving these differences remain incompletely understood. Endothelial-to-mesenchymal transition (EndMT) is increasingly recognized as a contributor to plaque remodeling, but whether EndMT is regulated in a sex-specific and stage-dependent manner across atherosclerosis has not been systematically examined. Here, we integrated bulk RNA sequencing with single-cell transcriptomic analyses in endothelial cells and human atherosclerotic plaques to define sex-specific EndMT regulation across healthy and advanced-stage disease. Using trajectory analyses in endothelial cells undergoing EndMT, we observed pronounced sex-specific regulation in healthy endothelium, with females exhibiting stronger early EndMT activation, whereas endothelial cells derived from atherosclerotic plaques displayed markedly attenuated sex differences with disease progression. Consistent sex-divergent pseudotime trajectories were observed in human carotid plaque endothelial single-cell RNA-seq data, with females showing greater EndMT activation at earlier stages and males at later stages. Together, these findings support a stage-dependent model of sex-specific EndMT regulation, indicating that the functional consequences of EndMT are highly context dependent and may differ across early and late disease stages. Integration of these datasets prioritized high-confidence sex-specific EndMT regulators, including COL4A1, PECAM1, CD151, JAG1, FN1, NEDD9, PODXL, MAFB, PROCR, and CDH13, providing a mechanistic framework to explain clinically observed sex differences in plaque biology and to guide targeted functional interrogation.

genomics↗

bioRxiv2025

Metabolic and Redox Pathway Dysregulation in HIV-Associated Coronary Endothelial Dysfunction

BackgroundPeople with HIV (PWH), even with sustained viral suppression on antiretroviral therapy (ART), remain at increased risk for cardiovascular disease. Coronary endothelial dysfunction, a sensitive marker of early vascular injury and a potential target for intervention is common in this population, but its biological basis remains unknown. MethodsWe performed a cross-sectional study combining in vivo coronary MRI and high-throughput serum proteomics to investigate mechanisms of coronary endothelial dysfunction in treated HIV. Forty-five virally suppressed PWH and twenty-nine age- and sex-matched healthy controls underwent coronary MRI during isometric handgrip exercise to quantify coronary endothelial function, defined as the percentage change in coronary cross-sectional area (%CSA) from rest to stress. An increase in coronary CSA <2% indicated endothelial dysfunction. Parallel serum proteomic profiling was performed using the SomaScan 7K platform, and differential protein expression between groups was analyzed using linear modeling (LIMMA). ResultsCoronary endothelial dysfunction was more prevalent in PWH with suppressed viral load compared to controls (67% vs 10%, p<0.001). Pathway analysis of differentially expressed proteins between participants with and without endothelial dysfunction highlighted significant dysregulation of glutathione dependent detoxification, oxidative metabolism and fatty acid {beta}-oxidation pathways in individuals with endothelial dysfunction (adjusted p value <0.05). ConclusionsEndothelial dysfunction in PWH on ART is associated with metabolic and redox imbalance. These findings highlight glutathione and fatty acid oxidation related pathways as potential therapeutic targets for reducing cardiovascular risk in this patient population.

immunology↗

bioRxiv2025

Measuring partner synchrony during salsa dancing and its relationship to changes in psychosocial domains

BackgroundDance is a promising rehabilitation adjunct but understanding the mechanisms through which dance improves physical and psychosocial well-being is necessary for implementation. This study investigated the feasibility of a measure of one possible mechanism: partner synchrony. Secondary objectives were to examine the relationships between partner synchrony, instructor ratings, and changes in psychosocial variables. MethodsParticipants wore an Inertial Measurement Unit (IMU) sensor during a salsa class that was video recorded. Partner synchrony was quantified with correlations of x, y, and z-axis acceleration between dancing pairs. Psychosocial variables were measured pre- and post-class. Salsa instructors rated partner synchrony from video recordings. Feasibility parameters included percentage of participants with data collected and sensor comfort. Pre-post changes were analyzed with Wilcoxon ranked tests and relationships were analyzed with Spearman correlations. ResultsData was collected for 23/24 (96%) participants and 17/24 (81%) reported the sensor was comfortable. All psychosocial variables improved from pre- to post-class. Partner synchrony was significantly associated with instructor ratings and change in positive and negative affect. ConclusionsAn IMU-based measure of dance partner synchrony is feasible, associated with ratings by dance instructors and related to changes in mood. The partner synchrony measure can be used to advance our understanding of dance as a therapeutic tool after its reliability is investigated.

neuroscience↗

bioRxiv2025

Predicting specificity of TCR-pMHC interactions using machine learning and biophysical models

Understanding the mechanism of T-cell activation and T-cell receptor (TCR) discrimination of MHC-presented epitope peptides (pMHCs) remains an open problem. Machine learning (ML)-based prediction of TCR specificity has gained considerable recent attention. However, the capacity of current models to generalize to peptides unseen during training is currently unknown. Here, we use a proprietary cancer-patient data set that profiles TCR binding to novel regions of peptide space to show that peptide generalization remains an unsolved problem. Specifically, we show that while ML methods have demonstrable utility in predicting TCR specificity for known peptides, they fail to generalize to novel peptides. We also show that physics-based methods utilizing classical energy functions outperform ML methods when predicting TCR binding to novel peptides but underperform them on known peptides. In light of these observations, we develop a new ML model that leverages general knowledge acquired by protein foundation models to achieve better or comparable performance than either ML or biophysical methods on both in- and out-of-distribution TCR-pMHC specificity prediction. We furthermore analyze model performance as a function of distance of TCR sequence specificity between the training and test sets to quantitatively characterize the generalization potential of any given TCR-pMHC model. Our analysis sheds light on the status of modeling TCR-pMHC interactions and suggests new paths forward for continued method development and data acquisition.

immunology↗

bioRxiv2024

Antibody-Fab and -Fc features promote Mycobacterium tuberculosis restriction.

Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), is a leading cause of death by an infectious disease globally, with no efficacious vaccine. Antibodies are implicated in Mtb control, but the mechanisms of antibody action remain poorly understood. We assembled a library of TB monoclonal antibodies (mAb) and screened for the ability to restrict Mtb in mice, identifying protective antibodies targeting known and novel antigens. To dissect the mechanism of mAb-mediated Mtb restriction, we optimized a protective lipoarabinomannan-specific mAb through Fc-swapping. In vivo analysis of these Fc-variants revealed a critical role for Fc-effector function in Mtb restriction. Restrictive Fc-variants altered distribution of Mtb across innate immune cells. Single-cell transcriptomics highlighted distinctly activated molecular circuitry within innate immune cell subpopulations, highlighting early activation of neutrophils as a key signature of mAb-mediated Mtb restriction. Therefore, improved antibody-mediated restriction of Mtb is associated with reorganization of the tissue-level immune response to infection and depends on the collaboration of antibody Fab and Fc.

immunology↗

bioRxiv2024

Comprehensive identification of coronary artery disease-associated variants regulating vascular smooth muscle cell gene expression

Coronary artery disease (CAD) is a complex disorder with genetic and environmental influences. Genome-wide association studies (GWAS) have identified over 300 genomic loci associated with disease risk. However, identifying the functional variants within these loci has been limited in large part due to linkage disequilibrium. This represents a critical step in understanding the molecular mechanisms underlying disease risk. To identify and prioritize candidate causal CAD-associated variants, we performed lentivirus-based massively parallel reporter assays (lentiMPRAs) in primary vascular smooth muscle cells (SMCs), which play significant roles in atherosclerosis, the underlying cause of CAD. We tested 25892 CAD-associated variants for their allele-specific enhancer activity in quiescent and proliferative SMCs, modeling healthy and disease conditions. We identified 122 candidate variants showing significant enhancer activity and differences in reporter gene expression between risk and non-risk alleles. We also identified 23 variants showing condition-biased allelic imbalance and 41 variants showing sex-biased allelic imbalance. We further functionally characterized 25892 variants by performing CUT&RUN assays to identify variants in enhancer and promoter regions of SMCs. By integrating the results of these experiments, we identified a credible set of 49 CAD-associated variants in functionally relevant regions. Furthermore, by comparing these identified variants with our previously obtained expression quantitative trait loci (eQTL) data, 27 of these 49 variants were associated with SMC gene expression levels. Finally, we performed CRISPRi experiments on 8 variants comprising 9 variant-gene pairs, rs35976034 (MAP1S), rs4888409 (CFDP1), rs73193808 (MAP3K7CL), rs67631072 (INPP5B/FHL3), rs1651285 (SNHG18), rs17293632 (SMAD3), rs2238792 (ARVCF), rs4627080 (NRIP3), to confirm their regulatory potential of nearby gene expression. Taken together, our results comprehensively fine-map the causal variants that confer increased risk of CAD through their effects on vascular smooth muscle cells.

genomics↗

bioRxiv2024

Extracellular vesicles-based point-of-care testing for the diagnosis and monitoring of Alzheimer's disease

Alzheimers disease (AD) is a debilitating condition that affects millions of people worldwide. One promising strategy for detecting and monitoring AD early on is using extracellular vesicles (EVs)-based point-of-care testing; however, diagnosing AD using EVs poses a challenge due to the low abundance of EV-biomarkers. Here, we present a fully integrated organic electrochemical transistor (OECT) that enables high accuracy, speed, and convenience in the detection of EVs from AD patients. We incorporated self-aligned acoustoelectric enhancement of EVs on a chip that rapidly propels, enriches, and specifically binds EVs to the OECT detection area. With our enhancement of pre-concentration, we increased the sensitivity to a limit of detection of 500 EV particles/L and reduced the required detection time to just two minutes. We also tested the sensor on an AD mouse model to monitor AD progression, examined mouse A{beta} EVs at different time courses, and compared them with intraneuronal A{beta} cumulation using MRI. This innovative technology has the potential to diagnose Alzheimers and other neurodegenerative diseases accurately and quickly, enabling monitoring of disease progression and treatment response.

bioengineering↗

bioRxiv2024

The evolutionary dynamics of alternative splicing during primate neuronal differentiation

Alternative splicing (AS) is emerging as an important regulatory process for complex biological processes such as neuronal differentiation. To uncover the functional consequences of AS during neuronal differentiation we performed a comparative transcriptomic analysis using human, rhesus, chimpanzee and orangutan pluripotent stem cells. Transcriptomic studies commonly involve the identification and quantification of alternative processing events, but the need for predicting the functional consequences of changes to the relative inclusion of alternative events remains largely unaddressed. Many tools exist for the former task, albeit often limited to rudimentary event types. Few tools exist for the latter task; each with significant limitations. To address these issues we developed junctionCounts, which captures both simple and complex pairwise AS events and quantifies them with straightforward exon-exon and exon-intron junction reads in RNA-seq data, performing competently among similar tools in terms of sensitivity, false discovery and quantification accuracy. Its partner utility, cdsInsertion identifies transcript coding sequence information, including the presence of premature termination codons, gathered via in silico translation from annotated start codons. It then couples transcript-level information to AS events to predict functional effects, i.e. nonsense-mediated decay (NMD). We used junctionCounts and related tools to discover both conserved and species-specific splicing dynamics as well as regulation of NMD during differentiation. Our work demonstrates this tools capacity to robustly characterize AS and bridge the gap of predicting its potential effect on mRNA isoform fate. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/581203v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@17b2be4org.highwire.dtl.DTLVardef@17e8992org.highwire.dtl.DTLVardef@79e47forg.highwire.dtl.DTLVardef@888161_HPS_FORMAT_FIGEXP M_FIG C_FIG junctionCounts is an alternative splicing analysis tool that identifies both simple and complex splicing events from a gene annotation and then measures their percent spliced-in from mapped RNA-seq junction reads.

molecular biology↗

bioRxiv2023

Absolute Quantification of Microbiota in Shotgun Sequencing Using Host Cells or Spike-Ins

BackgroundAn ongoing challenge for DNA sequencing of samples containing microorganisms is the ability to meaningfully compare different samples and to connect the results back to clinically relevant disease states. The reads of DNA sequence from each sample do not, in and of themselves, give sufficient information to calculate the absolute abundances of each observed organism. Using relative abundances alone is insufficient to determine whether absolute abundances have increased or decreased in the organisms of interest from one sample to the next. This is a well-studied problem in 16S sequencing, but solutions in shotgun sequencing are lacking. Here we show how spike-ins can be used in shotgun sequencing to calculate absolute abundances of organisms present. We also propose the use of the host cells already in the sample as an alternative calculation method. Mammalian host cells are typically of sufficient size that they can be easily and cheaply counted prior to sequencing by a variety of methods and combining this with sequencing data provides sufficient information to calculate the absolute abundances of microbial organisms. ResultsMicrobial abundances in the samples calculated via this method were consistent with manufacturer-stated values of microbial communities, with qPCR, and with our method tested against itself with regard the spike-in and host-cell based options. R2 values on the log10 scale in these tests ranged from 0.85 to 0.98, and the log10-RMSE ranged from 0.1 to 0.7. ConclusionsThe proposed method can consistently calculate absolute microbial abundances to within an order of magnitude. Both versions of the method, where spike-ins are added to the samples, or where host cells in the sample are counted, are viable. Calculating absolute abundances allows for direct comparisons to be made between different samples. If disease-thresholds have been identified, absolute abundances can quantify disease states.

microbiology↗

bioRxiv2022

Discovery of prevalent, clinically actionable tumor neoepitopes via integrated biochemical and cell-based platforms

Strategies for maximizing the potency and specificity of cancer immunotherapies have sparked efforts to identify recurrent epitopes presented in the context of defined tumor-associated neoantigens. Discovering these "neoepitopes" can be difficult owing to the limited number of peptides that arise from a single point mutation, a low number of copies presented on the cell surface, and variable binding specificity of the human leukocyte antigen (HLA) class I complex. Due to these limitations, many discovery efforts focus on identifying neoepitopes from a small number of cancer neoantigens in the context of few HLA alleles. Here we describe a systematic workflow to characterize binding and presentation of neoepitopes derived from 47 shared cancer neoantigens in the context of 15 HLA alleles. Through the development of a high-throughput neoepitope-HLA binding assay, we surveyed 24,149 candidate neoepitope-HLA combinations resulting in 587 stable complexes. These data were supplemented by computational prediction that identified an additional 257 neoepitope-HLA pairs, resulting in a total of 844 unique combinations. We used these results to build sensitive targeted mass spectrometry assays to validate neoepitope presentation on a panel of HLA-I monoallelic cell lines engineered to express neoantigens of interest as a single polypeptide. Altogether, our analyses detected 84 unique neoepitope-HLA pairs derived from 37 shared cancer neoantigens and presented across 12 HLA alleles. We subsequently identified multiple TCRs which specifically recognized two of these neoantigen-HLA combinations. Finally, these novel TCRs were utilized to elicit a T cell response suggesting that these neoepitopes are likely to be immunogenic. Together these data represent a validated, extensive resource of therapeutically relevant neoepitopes and the HLA context in which they can be targeted.

cancer biology↗

bioRxiv2022

Fc-engineered antibodies leverage neutrophils to drive control of Mycobacterium tuberculosis

Novel vaccination and therapeutic strategies are urgently needed to mitigate the tuberculosis (TB) epidemic. While extensive efforts have focused on potentiating cell-mediated immunity to control Mycobacterium tuberculosis (Mtb) infection, less effort has been invested in exploiting the humoral immune system to combat Mtb. Emerging data point to a role for antibodies in microbial control of Mtb, however the precise mechanism(s) of this control remain incompletely understood. Here we took an antibody Fc-engineering approach to determine whether Fc-modifications could improve the ability of antibodies to restrict Mtb, and to define Fc-mediated mechanism(s) antibodies leverage for this restriction. Using an antibody specific to the capsular polysaccharide -glucan, we engineer a panel of Fc variants to augment or dampen select antibody effector functions, rationally building antibodies with enhanced capacity to promote Mtb restriction in a human whole blood model of infection. Surprisingly, restrictive Fc-engineered antibodies drive Mtb control in a neutrophil, not monocyte, dependent manner. Using single cell RNA sequencing, we show that restrictive antibodies promote neutrophil survival and expression of cell intrinsic antimicrobial programs. These data provide a roadmap for exploiting Fc-engineered antibodies as a novel class of TB therapeutics able to harness the protective functions of neutrophils to achieve disease control.

immunology↗

bioRxiv2021

Origins and timing of emerging lesions in advanced renal cell carcinoma

PurposeRenal cell carcinoma (RCC) with venous tumor thrombus (VTT) arising from the primary tumor occurs in 4-10% of cases and is associated with advanced disease. RCC with VTT and distant metastasis represents a unique clinical entity, and provides opportunities to examine the origins and relative timing of tumor lesion emergence and to identify molecular correlates with disease state. Experimental DesignWe performed genomic and evolutionary analyses on 16 RCC patients with VTT, with eight also having metastases, using multi-region exome and RNA sequencing. ResultsNo genomic alterations were specifically associated with the VTT or metastasis lesions; each tumor had multiple hallmark driver alterations, consistent with advanced disease state. We found that 21% (3/14) of clear-cell RCC cases could be assigned a previously defined "evolutionary subtype". Somatic mutation signatures were largely consistent with previously established RCC signatures, and showed low heterogeneity across regions of each tumor. Mismatch repair and homologous recombination ("BRCA-ness") deficiency signatures consistently co-occurred across most tumors, suggesting a pervasive role for intracellular DNA damage in RCC and the potential for related treatment strategies. Phylogenetic timing analysis of metastatic cases suggested that in most tumors, metastases branched from the primary tumor prior to formation of VTT and in some cases before diversification of the primary tumor. Both VTT and the earliest metastases were predicted to emerge many years prior to diagnosis. Transcriptional landscape analysis identified key differences distinguishing each lesion type from primary tumor: VTT upregulated TNF signaling and associated inflammatory pathways, whereas metastases upregulated MTOR signaling. ConclusionsOur results provide a map of how RCC tumors can evolve, with metastatic clones typically emerging early in RCC development and taking hold via MTOR signaling, and later formation of VTT via local inflammatory processes. Statement of Translational RelevanceRenal cell carcinoma (RCC) is a deadly and relatively common malignancy, which often presents as or progresses to metastatic disease. We used multi-region sequencing of RCC patients with venous tumor thrombus (VTT) and metastasis to ask how and when new lesions arise from the primary tumor, and what genomic factors contribute to their spread. Phylogenetic analysis of patients with VTT and co-presenting metastases suggested that in most cases, the VTT and metastases derive from distinct tumor clones. Moreover, metastatic clones often appear many years prior to diagnosis. We found that local TNF inflammation may contribute to VTT formation, whereas MTOR signaling is associated with metastases. Our study sheds light on the relationship of VTT and metastases, suggests therapeutic and biomarker strategies for RCC, and points to the need for early detection studies in RCC to better understand when metastases emerge and to identify at-risk patients.

cancer biology↗

bioRxiv2021

Indication-specific tumor evolution and its impact on neoantigen targeting and biomarkers for individualized cancer immunotherapies

Individualized neoantigen specific immunotherapy (iNeST) requires robustly expressed clonal neoantigens for efficacy, but tumor mutational heterogeneity, loss of neoantigen expression, and variable tissue sampling present challenges. To characterize these potential obstacles, we combined multi-region sequencing (MR-seq) analysis of five untreated, synchronously sampled metastatic solid tumors with re-analysis of published MR-seq data from 103 patients. Branching evolution in colorectal cancer and renal cell carcinoma led to fewer clonal neoantigens and to clade-specific neoantigens (those shared across a subset of tumor regions but not fully clonal), with the latter not being readily distinguishable in single tumor samples. Prioritizing mutations with higher purity- and ploidy-adjusted variant allele frequency enriched for globally clonal neoantigens (those found in all tumor regions), whereas estimated cancer cell fraction derived from clustering-based tools, surprisingly, did not. Neoantigen quality was associated with loss of neoantigen expression in the bladder cancer case, and HLA-allele loss was observed in the renal and non-small cell lung cancer cases. Our results show that indication type, multi-lesion sampling, neoantigen expression, and HLA allele retention are important factors for iNeST targeting and patient selection.

cancer biology↗

bioRxiv2020

CaBagE: a Cas9-based Background Elimination strategy for targeted, long-read DNA sequencing

A substantial fraction of the human genome is difficult to interrogate with short-read DNA sequencing technologies due to paralogy, complex haplotype structures, or tandem repeats. Long-read sequencing technologies, such as Oxford Nanopores MinION, enable direct measurement of complex loci without introducing many of the biases inherent to short-read methods, though they suffer from relatively lower throughput. This limitation has motivated recent efforts to develop amplification-free strategies to target and enrich loci of interest for subsequent sequencing with long reads. Here, we present CaBagE, a novel method for target enrichment that is efficient and useful for sequencing large, structurally complex targets. The CaBagE method leverages the stable binding of Cas9 to its DNA target to protect desired fragments from digestion with exonuclease. Enriched DNA fragments are then sequenced with Oxford Nanopores MinION long-read sequencing technology. Enrichment with CaBagE resulted in up to 416X coverage of target loci when tested on five genomic targets ranging from 4-20kb in length using healthy donor DNA. Four cancer gene targets were enriched in a single reaction and multiplexed on a single MinION flow cell. We further demonstrate the utility of CaBagE in two ALS patients with C9orf72 short tandem repeat expansions to produce genotype estimates commensurate with genotypes derived from repeat-primed PCR for each individual. With CaBagE there is a physical enrichment of on-target DNA in a given sample prior to sequencing. This feature allows adaptability across sequencing platforms and potential use as an enrichment strategy for applications beyond sequencing. CaBagE is a rapid enrichment method that can illuminate regions of the hidden genome underlying human disease.

genetics↗

bioRxiv2020

Oral administration of a single anti-CfaE nanobody provides broadly cross-protective immunity against major pathogenic Enterotoxigenic Escherichia coli strains

Enterotoxigenic Escherichia coli (ETEC) is estimated to cause approximately 380,000 deaths annually during sporadic or epidemic outbreaks worldwide. There is currently no vaccine licensed to prevent ETEC. Development of prophylaxis against ETEC is challenging due to the vast heterogeneity of the ETEC strains. The discovery of nanobodies has emerged as a successful new biologics in treating mucosal infectious disease as nanobodies can recognize conserved epitopes on hypervariable pathogens. In this study, we performed large screens using immunized llamas and a naive nanobody yeast display library against adhesins of colonization factors. Cross-protective nanobodies were selected with in vitro activities inhibiting mannose-resistant hemagglutination (MRHA) against all eleven major pathogenic ETEC strains. Oral administration of nanobodies led to significant reduction of bacterial colonization in animals challenged with multiple ETEC strains. Structural analysis revealed novel conserved epitopes as critical structural features for pan-ETEC vaccine design. Two of the lead nanobodies, 2R215 and 1D7, were further engineered as trimer or fused with human IgA Fc-fragments as fusionbodies. Oral administration of the trimers or fusionbodies protected mice from infection at a much lower dose compared to the monomeric format. Importantly, fusionbodies prevented infection as a pre-treatment when administrated 2 hours before ETEC challenge to the animals. Together, our study provides the first proof of concept that oral administration of a single nanobody could confer broad protection against major pathogenic ETEC strains. Technological advances in large-scale manufacturing of biological proteins in plants and microorganisms will make nanobody-based immunotherapy a potent and cost-effective prophylaxis or treatment for ETEC.

microbiology↗

bioRxiv2019

RhoGAP RGA-8 supports morphogenesis in C. elegans by polarizing epithelia through CDC-42

CDC-42 regulation of non-muscle myosin/NMY-2 is required for polarity maintenance in the one-cell embryo of C. elegans. CDC-42 and NMY-2 regulate polarity throughout embryogenesis, but their contribution to later events of morphogenesis are less understood. We have shown that epidermal enclosure requires the GTPase CED-10/Rac1 and WAVE/Scar complex, its effector, to promote protrusions that drive enclosure through the branch actin regulator Arp2/3. Our analysis here of RGA-8, a homolog of SH3BP1/Rich1/ARHGAP17/Nadrin, with BAR and RhoGAP motifs, suggests it regulates CDC-42, so that NMY-2 promotes two events of epidermal morphogenesis: ventral enclosure and elongation. Genetic and molecular data suggest RGA-8 regulates CDC-42, and the CDC-42 effectors WSP-1 and MRCK-1, in parallel to F-BAR proteins TOCA-1 and TOCA-2. The RGA-8-CDC-42-WSP-1 pathway enriches myosin in migrating epidermal cells during ventral enclosure. We propose TOCA proteins and RGA-8 use BAR domains to localize and regenerate CDC-42 activity, thus regulating F-actin levels, through the branched actin regulator WSP-1, and myosin polarity through the myosin kinase MRCK-1. Regulated CDC-42 thus polarizes epithelia, to control cell migrations and cell shape changes of embryonic morphogenesis. SummaryRGA-8, a protein with membrane binding and actin regulatory motifs, promotes embryonic morphogenesis by localizing active CDC-42 in developing epithelia, thus controlling actin and actin motors during cell movements.

developmental biology↗