Biology subjects

Zaidel-Bar, R.

Publications and source records attributed to Zaidel-Bar, R..

14 recordsLinked to original sources

bioRxiv2026

Conserved tyrosine Y16 acts as a phosphorylation switch controlling Ena/VASP recruitment to sites of actin assembly

Ena/VASP proteins are powerful actin polymerases that accumulate at sites of active actin assembly, including lamellipodial and microspike tips, focal adhesions and pathogen surfaces, where they drive processive filament elongation. However, the mechanisms controlling their recruitment remain elusive. Here, we used cells lacking all Ena/VASP-family members to systematically test phosphomimetic and nonphosphorylatable substitutions at conserved tyrosine and serine residues implicated in subcellular targeting. Combining cell-based reconstitution with biochemical assays, we identify phosphorylation of Y16 in vertebrate Ena/VASP proteins as the decisive inhibitory modification controlling subcellular Ena/VASP recruitment, in contrast to previously proposed roles for Y39 and S157. Corresponding phosphomimetic substitutions in the C. elegans orthologue UNC-34 phenocopied uncoordinated locomotion of UNC-34 null worms, supporting the evolutionary conservation of this regulatory mechanism. Thus, our findings identify a conserved phosphorylation-sensitive switch at the EVH1-FP4 ligand interface that links EVH1-dependent targeting to dynamic control of actin assembly in adhesion, migration and pathogenesis.

cell biology↗

bioRxiv2026

Capping protein regulates the balance of assembly among diverse actin networks in C. elegans zygotes

Actin cytoskeleton networks exhibit specialized architectural properties for specific cellular tasks, as determined by the actin-binding proteins (ABPs) associated with each network. Proper allocation of a limiting pool of actin monomers also helps shape the assembly of different F-actin networks. The ABP capping protein (CP) modulates F-actin network architecture through regulation of actin filament length by capping filament barbed ends. Using a combination of in vitro biochemistry and quantitative live-cell imaging, we characterize CP as a major regulator of inter-network competition between filopodia and mini-comets, two F-actin networks in the one-cell C. elegans embryo (zygote). We establish that this regulation is facilitated in part by competition for binding barbed ends between CP and the F-actin elongator formin CYK-1. Together, these results reveal a role for CP in determining F-actin network architecture and dynamics, regulating the coordination between actin assembly factors to assemble and maintain different dynamic F-actin networks, and allocation of G-actin between competing cortical F-actin networks. Summary for table of contentsCells assemble diverse actin cytoskeleton networks within a common cytoplasm for essential cellular processes. Yde et al. establish a role for Capping Protein, a regulator of actin filament length, in coordinating the balanced assembly of distinct actin networks in the C. elegans zygote.

cell biology↗

bioRxiv2026

Scavenger Cells Failure to Maintain Systemic RNA Homeostasis Causes Epigenetically Inherited Germline Tumors

Temporary disruptions to epigenetic mechanisms can misroute development and permanently alter cell fate. In particular, it was recently shown that transient loss of Polycomb silencing in flies irreversibly reprograms cells toward cancer (Parreno et al. 2024). Whether somatic dysfunction in parents can create multi-generational heritable susceptibility to tumorigenesis is unknown. In eutelic organisms like Caenorhabditis elegans, adult somatic cells no longer divide, precluding somatic cancer, yet tumors can still form in the continuously dividing germline. Here, we show that disruption of coelomocytes, somatic scavenger cells, just in C. elegans mothers, provokes transgenerationally heritable germline tumorigenesis that persists for multiple generations in genetically wild-type descendants. We found that when the coelomocytes phagocytic activity dysfunctions, it impairs clear out of RNA from body fluids, and thus disrupts systemic RNA homeostasis, allowing excess somatic RNAs to access the germline, and leading to widespread transcriptional and small RNA dysregulation and transgenerational loss of germline identity. Converging lines of evidence point towards small RNAs being the heritable agents carrying the pathological information. Together, these findings highlight mechanisms which maintain systemic RNA homeostasis as an important protective barrier against heritable tumorigenesis.

genetics↗

bioRxiv2025

A switch in collagen expression regulates cessation of distal tip cell migration in C. elegans

Development of the C. elegans gonad requires precise regulation of cell migration. The distal tip cell (DTC) guides the elongation of the gonad into its final U-shaped structure before halting in adulthood. How cessation of elongation is regulated remains unknown. Here, we analyze an RNA-seq data set isolated from stage-specific DTCs to uncover the temporal gene expression dynamics underlying this process. Collagens emerged as the most enriched gene family during the transition from migratory larval stages to adulthood. We identify distinct temporal waves of collagen expression, culminating in a core adult-specific module that coincides with migration cessation. Functional analysis by RNAi depletion revealed that many collagens upregulated in adulthood are required for timely migration arrest, while others affect gonad shape. Perturbation of collagen remodeling enzymes phenocopied these effects. Our findings uncover a stage-specific collagen program in the DTC and suggest that terminal migration arrest is actively reinforced by matrix remodeling.

developmental biology↗

bioRxiv2025

Stage-specific transcriptomics of a leader cell reveals cell machineries driving collective invasion

Collective cell invasion underlies organ development, epithelial repair, and cancer metastasis. "Leader cells" remodel extracellular matrix, sense guidance cues, reorganize their cytoskeleton, and coordinate follower cells, but the molecular programs enabling these functions remain unclear. Here, we present a stage-specific transcriptomic dataset of the C. elegans gonadal leader cell, the distal tip cell (DTC), which invades basement membrane and guides germ cells to form U-shaped gonadal arms. Comparing invasive larval-stage DTCs with non-invasive adult-stage DTCs defines the molecular signature of an actively invading leader cell in vivo. Our dataset recapitulates known regulators of gonad morphogenesis and reveals numerous uncharacterized genes with potential roles in leader cell activity. As proof of concept, we identify vesicular trafficking proteins as enriched in invading DTCs, and demonstrate their importance for gonad development using endogenous tagging and DTC-specific RNAi. We also catalog diverse DTC-specific knockdown phenotypes. This resource establishes a molecular framework for leader cell activity and a platform to investigate conserved mechanisms of invasive migration.

developmental biology↗

bioRxiv2025

Twinfilin is a potent uncapper of actin capping protein and modulates actomyosin contractility in the C. elegans spermatheca

The actin cytoskeleton is dynamically remodelled by conserved regulators to control cellular and tissue mechanics. While the functions of these proteins are well studied, how they drive tissue-specific contractility remains unclear. Twinfilin, an actin uncapper and depolymerase, has not previously been linked to tissue contractility. Here, we show that the sole twinfilin ortholog in C. elegans, TWF-2, regulates actomyosin contractility in the spermatheca. TWF-2 localizes to the spermathecal cortex via interactions with -spectrin (SPC-1) and {beta}-spectrin (UNC-70). In vitro, TWF-2 promotes barbed-end depolymerization and rapidly removes CAP-1 from actin filaments. In vivo, embryonic lethality caused by CAP-1 depletion is partially rescued by simultaneous loss of TWF-2. Similarly, loss of the contractility regulator SPV-1 leads to elevated F-actin and phosphorylated myosin, causing hypercontractility. Notably, removing TWF-2 suppresses this hypercontractility by reducing F-actin levels-- without affecting myosin or its phosphorylation--highlighting a specific role in F-actin regulation. Together, these findings show that TWF-2 modulates actin dynamics in a tissue-specific manner. This work provides the first in vivo evidence that twinfilin regulates contractility, and reveals how its interactions with capping protein and spectrins help maintain balanced actomyosin levels in the spermatheca.

cell biology↗

bioRxiv2024

AI-assisted imaging screening reveals mechano-molecular tissue organizers and network of signaling hubs.

Cadherin-mediated adhesions are crucial mechanical and signaling hubs that connect cells within a tissue and probe the mechanics of the surrounding environment. They constitute a physical link between the actin cytoskeleton of neighboring cells, providing the mechanical coordination needed for morphogenetic processes, tissue homeostasis, collective migration, and regeneration. Disruptions in adhesion mechanisms are closely linked to the breakdown of epithelial structure and the emergence of disease-related traits characteristic of cancer progression. The cadhesome network comprises over 170 structural and regulatory proteins involved in cadherin-mediated adhesion. While this network is essential for coordinating tissue responses to mechanical stress, its complexity has historically limited our understanding of how individual components contribute to force transmission and tissue homeostasis. Recent technological advances offer tools to investigate large molecular networks in cellular function and pathology (functional omics). Leveraging these advances, we developed an experimental and analytical platform combining high-throughput gene silencing, imaging, and artificial intelligence (AI) to systematically profile each role of each protein in tissue formation, mechanical stability, and response to induced tension. Using EpH4 cells as an epithelial tissue model, we performed systematic silencing in triplicate, capturing a range of tissue phenotypes under baseline and tension-inducing conditions. Machine learning methods were used to analyze complex imaging data, quantify tissue ruptures, characterize junctional organization, and measure tension states of the tissue. By incorporating machine learning algorithms, we automated image feature extraction, clustering, and classification, enabling an unprecedented quantitative evaluation of tissue mechanics at scale. Our machine learning models allowed us to identify significant patterns, including protein-specific responses to tension and their roles in tissue-level mechanical integrity. Finally, we constructed a protein interaction network detailing the roles of each protein, their physical interactions, and known links to cancer. The network analysis revealed three prominent mechanotransductive and signaling subnetworks centered around E-cadherin, EGFR, and RAC1. Our study provides a foundational framework for investigating mechanosensing proteins and it offers a scalable blueprint for discovering potential therapeutic targets in diseases like cancer, where tissue mechanics play a crucial role. TeaserAI-aided screening identifies key regulators of epithelial tissue mechanics, uncovering potential therapeutic targets in cancer.

cell biology↗

bioRxiv2024

Genetically encoded biosensor for fluorescence lifetime imaging of PTEN dynamics in the intact brain

The phosphatase and tensin homolog (PTEN) is a vital signaling protein which maintains an inhibitory brake that is critical for cellular metabolism, proliferation, and growth. The importance of PTEN signaling is evident from the broad spectrum of human pathologies associated with its loss of function. Moreover, loss or gain of PTEN function in animal models leads to aberrant cellular morphology, function, and metabolic regulation. However, despite the important role of PTEN signaling, there is currently no method to dynamically monitor its activity with cellular specificity within intact biological systems. Here, we describe the development of a novel PTEN biosensor, optimized for two-photon fluorescence lifetime imaging microscopy (2pFLIM). This biosensor is designed to measure PTEN activity within intact cells, tissues, and organisms. Our approach is based on monitoring FRET-dependent changes in PTEN conformation, which serves as a proxy for the activity state in living cells. We identify a point mutation that allow us to express this biosensor with minimal interference to endogenous PTEN signaling and cellular function. We demonstrate the utility of imaging PTEN signaling in cell lines, developing C. elegans, and in the living mouse brain. To complement this approach, we developed a red-shifted PTEN sensor variant that permits simultaneous imaging with GFP-based sensors. Finally, we use in vivo PTEN imaging in the mouse brain to identify cell-type specific dynamics of PTEN activity in excitatory and inhibitory cortical cells. In summary, our approach enables dynamic imaging of PTEN activity in vivo with unprecedented spatial and temporal resolution.

neuroscience↗

bioRxiv2024

A Tunable and Druggable Mechanism to Delay Forgetting of Olfactory Memories in C. elegans

The poet W.B Yeats wrote that "All that is personal soon rots, it must be packed in ice or salt". Here we show that in Caenorhabditis elegans nematodes, simple animals with just 302 neurons, memories are preserved on ice and in lithium salt. C. elegans nematodes can form associative memories, which are typically forgotten quickly. We discovered that when placed on ice, worms delay forgetting of specific olfactory memories by at least 8-fold. Delayed forgetting was canceled completely when the worms were gradually adapted to low temperatures, owing to a genetically-encoded program that turns acclimated worms cold-tolerant. RNA-seq, mutant analyses, and pharmacological assays revealed that regulation of membrane properties switches cold-induced delayed forgetting ON and OFF, and, remarkably, that lithium delays forgetting only in cold-sensitive but not cold-tolerant worms. We found that downregulation of the diacylglycerol pathway in the AWC sensory neurons is essential for lithium-mediated delayed forgetting, and using neuronal activity recordings located the memory trace to the downstream AIY interneurons. We suggest that the awesome genetic tractability of C. elegans might be harnessed to study the effects of lithium and cold temperatures on the brain, why it influences psychiatric disorders, and even more fundamentally how memory is stored and lost.

neuroscience↗

bioRxiv2023

Chloride intracellular channel (CLIC) proteins function as fusogens

Chloride Intracellular Channel (CLIC) family members uniquely transition between soluble and membrane-associated conformations. Despite decades of extensive functional and structural studies, CLICs function as ion channels remains debated, rendering our understanding of their physiological role incomplete. Here, we expose a novel function of CLIC5 as a fusogen. We demonstrate that purified CLIC5 directly interacts with the membrane and induces fusion, as reflected by increased liposomal diameter and lipid and content mixing between liposomes. Moreover, we show that this activity is facilitated by acidic pH, a known trigger for CLICs transition to a membrane-associated conformation, and that increased exposure of the hydrophobic inter-domain interface is crucial for this process. Finally, mutation of a conserved hydrophobic interfacial residue diminishes the fusogenic activity of CLIC5 in vitro and impairs excretory canal extension in C. elegans in vivo. Together, our results unravel the long-sought physiological role of these enigmatic proteins.

biochemistry↗

bioRxiv2021

Actin capping protein regulates actomyosin contractility to maintain germline architecture in C. elegans

Actin dynamics play an important role in the morphogenesis of cells and tissues, yet the control of actin filament growth takes place at the molecular level. A challenge in the field is to link the molecular function of actin regulators with their physiological function. Here, we report the in vivo role of the actin capping protein CAP-1 in the C. elegans germline. We show that CAP-1 is associated with actomyosin structures in the cortex and rachis, where it keeps the level of contractility in check. A 60% reduction in the level of CAP-1 leads to a 2-fold increase in F-actin and non-muscle myosin II and only a 30% increase in Arp2/3. CAP-1 depletion leads to severe structural defects in the syncytial germline and oocytes, which can be rescued by reducing myosin activity. Thus, we uncover a physiological role for actin capping protein in maintaining C. elegans fertility by regulating the level of actomyosin contractility.

cell biology↗

bioRxiv2021

Probe the effect of clustering on EphA2 receptor signaling efficiency by subcellular control of ligand-receptor mobility

Clustering of ligand:receptor complexes on the cell membrane is widely presumed to have functional consequences for subsequent signal transduction. However, it is experimentally challenging to selectively manipulate receptor clustering without altering other biochemical aspects of the cellular system. Here, we develop a microfabrication strategy to produce substrates displaying mobile and immobile ligands that are separated by roughly one micron and thus experience an identical cytoplasmic signaling state, enabling precision comparison of downstream signaling reactions. Applying this approach to characterize the ephrinA1:EphA2 signaling system reveals that EphA2 clustering enhances receptor phosphorylation. Single molecule imaging clearly resolves increased molecular binding dwell time at EphA2 clusters for both Grb2:SOS and NCK:NWASP signaling modules. This type of intracellular comparison enables a substantially higher degree of quantitative analysis than is possible when comparisons must be made between different cells and essentially eliminates the effects of cellular response to ligand manipulation.

biophysics↗

bioRxiv2019

Germ Granules Functions are Memorized by Transgenerationally Inherited Small RNAs

In C. elegans nematodes, components of liquid-like germ granules were shown to be required for transgenerational small RNA inheritance. Surprisingly, we show here that mutants with defective germ granules (pptr-1, meg-3/4, pgl-1) can nevertheless inherit potent small RNA-based silencing responses, but some of the mutants lose this ability after many generations of homozygosity. Animals mutated in pptr-1, which is required for stabilization of P granules in the early embryo, display extremely strong heritable RNAi responses, which last for tens of generations, long after the responses in wild type animals peter out. The phenotype of mutants defective in the core germ granules proteins MEG-3 and MEG-4, depends on the genotype of the ancestors: Mutants that derive from maternal lineages that had functional MEG-3 and MEG-4 proteins exhibit enhanced RNAi inheritance for multiple generations. While functional ancestral meg-3/4 alleles correct, and even potentiates the ability of mutant descendants to inherit RNAi, defects in germ granules functions can be memorized as well; Wild type descendants that derive from lineages of mutants show impaired RNAi inheritance for many (>16) generations, although their germ granules are intact. Importantly, while P granules are maternally deposited, wild type progeny derived from meg-3/4 male mutants also show reduced RNAi inheritance. Unlike germ granules, small RNAs are inherited also from the sperm. Moreover, we find that the transgenerational effects that depend on the ancestral germ granules require the argonaute protein HRDE-1, which carries heritable small RNAs in the germline. Indeed, small RNA sequencing reveals imbalanced levels of many endogenous small RNAs in germ granules mutants. Strikingly, we find that hrde-1;meg-3/4 triple mutants inherit RNAi, although hrde-1 was previously thought to be essential for heritable silencing. We propose that germ granules sort and shape the RNA pool, and that small RNA inheritance memorizes this activity for multiple generations.

genetics↗

bioRxiv2018

The RhoGAP SPV-1 regulates calcium signaling to control the contractility of the C. elegans spermatheca during embryo transits.

Contractility of the non-muscle and smooth muscle cells that comprise biological tubing is regulated by the Rho-ROCK and calcium signaling pathways. Although many molecular details about these signaling pathways are known, less is known about how they are coordinated spatiotemporally in biological tubes. The spermatheca of the C. elegans reproductive system enables study of the signaling pathways regulating actomyosin contractility in live adult animals. The RhoGAP SPV-1 was previously identified as a negative regulator of RHO-1/Rho and spermathecal contractility. Here, we uncover a role for SPV-1 as a key regulator of calcium signaling. spv-1 mutants expressing the calcium indicator GCaMP in the spermatheca exhibit premature calcium release, elevated calcium levels, and disrupted spatial regulation of calcium signaling during spermathecal contraction. Although RHO-1 is required for spermathecal contractility, RHO-1 does not play a significant role in regulating calcium. In contrast, activation of CDC-42 recapitulates many aspects of spv-1 mutant calcium signaling. Depletion of cdc-42 by RNAi does not suppress the premature or elevated calcium signal seen in spv-1 mutants, suggesting other targets remain to be identified. Our results suggest SPV-1 works through both the Rho-ROCK and calcium signaling pathways to coordinate cellular contractility.\n\nHighlight SummaryThrough in vivo imaging of the calcium sensor GCaMP, we show that the RhoGAP SPV-1 is a key regulator of calcium signaling in the C. elegans spermatheca. Our data suggests SPV-1 acts at least partially through the small GTPase CDC-42 to modulate calcium signaling, while also acting on RHO-1 to modulate Rho-ROCK signaling. This places SPV-1 as a central regulator of cellular contractility.

cell biology↗