Biology subjects

Nitzan, A.

Publications and source records attributed to Nitzan, A..

6 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

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

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

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↗

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↗

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↗