Biology subjects

Gelleri, M.

Publications and source records attributed to Gelleri, M..

7 recordsLinked to original sources

bioRxiv2026

The nuclear actin cytoskeleton supports DNA double-strand break repair via VCP-mediated extraction of the KU70/80 complex from damaged chromatin

Double-strand breaks (DSBs) are critical lesions in genomic DNA, and their accurate repair is essential for maintaining genome stability. The nuclear actin cytoskeleton has been implicated in homology-directed repair (HDR) of DSBs. However, the underlying mechanism remains poorly understood. Here, we report that Myosin VI (Myo6), an actin-based motor protein, cooperates with F-actin in end resection and DSB mobilization. Our findings reveal that Myo6 directly interacts with both KU70 and the ubiquitin-dependent segregase VCP to facilitate the extraction of the KU70/80 complex from chromatin. This process is supported by F-actin, revealing an interplay between nuclear actin dynamics and the DSB repair machinery. By elucidating the function of Myo6 and its direct interactions with key repair factors, our study provides mechanistic insight into how repair mechanisms rely on nuclear actin to safeguard genome integrity.

cell biology↗

bioRxiv2026

BRD4, Mediator, and Pol II form heterogeneous condensates with distinct transcriptional and acetylation-dependent states

Transcriptional condensates at super-enhancers are thought to concentrate BRD4, Mediator, and RNA polymerase II (Pol II) to promote gene activation, yet their compositional organization and regulation remain poorly understood. We developed a high-throughput live-cell phenomics platform based on endogenous fluorescent tagging of BRD4, MED14 (Mediator), and POLR2A (Pol II) to systematically quantify transcriptional condensate states across >1,000 chemical perturbations. Contrary to prevailing models of largely co-occupied assemblies, we find compositionally heterogenous condensate populations. In particular, BRD4-only spots emerged as a prominent class that is depleted of Mediator and Pol II, enriched at chromatin, and resistant to transcription initiation inhibition. Mechanistically, compound screening coupled to mechanism-of-action analysis identifies histone acetylation as a dominant regulatory axis for BRD4-only spots: Bromodomain and Extra-Terminal motif (BET) and histone acetyltransferase inhibition selectively deplete BRD4-only condensates, while histone deacetylase inhibition expands them. Together, these findings support a model in which acetylation-dependent BRD4 condensates define a distinct chromatin-associated regulatory state that is separable from canonical transcriptionally engaged condensates. More broadly, our work establishes condensate composition as a quantitative phenotype and provides a scalable framework for systematically dissecting the regulation of condensates across perturbations, cell types, and disease contexts.

cell biology↗

bioRxiv2026

SUMO paralogues differentially affect phase separation and aggregation of intrinsically disordered proteins

SUMO, the small ubiquitin-like modifier, modulates interactions of folded proteins, but also affects the dynamics of biomolecular condensates. Moreover, SUMO can directly impinge on the biophysical properties of its targets, often increasing their solubility. TDP-43 is an RNA-binding protein containing an intrinsically disordered domain that participates in protein phase separation. Its aggregation, linked to neurodegenerative diseases, is counteracted by SUMOylation, but the underlying principles are poorly understood. We now reveal distinct mechanisms of how the two isoforms, SUMO1 and SUMO2, act on TDP-43. While SUMO1 inhibits phase separation by blocking TDP-43s self-assembly site, SUMO2 enhances phase separation via proximity-induced self-interaction involving its N-terminus and promotes a liquid-like state that prevents aggregation. The latter represents a general principle by which SUMO solubilises other targets. Our findings provide insight into how SUMOylation regulates transitions between soluble, liquid-like and aggregated states.

molecular biology↗

bioRxiv2024

The role of the synaptic vesicle protein SV2A in regulating mitochondrial morphology and autophagy.

The synaptic vesicle glycoprotein 2A (SV2A) is a transmembrane protein of synaptic vesicles. It is involved in key functions of neurons, focused on the regulation of neurotransmitter release. Here we report three novel findings suggesting a completely new role of SV2A. First, we demonstrate that SV2A is localized at the outer mitochondrial membrane (OMM) using confocal and super-resolution microscopy. Second, Inactivation of SV2A in our cell and animal models leads to fragmented mitochondria. In addition, SV2A also affects the basal autophagic flux as well as mitophagy. Third, using proteomics analysis we demonstrate that SV2A interacts with the fission factor DRP1 and the autophagy factor ATG9A. Using AlphaFold3 we provide a first glimpse of the molecular interaction between DRP1 and SV2A. Our findings demonstrate that SV2A is not only a vesicular protein but also a mitochondrial protein in the OMM with defined functions regulating mitochondrial morphology and autophagy.

molecular biology↗

bioRxiv2024

Space-time dynamics of genome replication studied with super-resolved microscopy

Genome replication requires duplication of the complete set of DNA sequences together with nucleosomes and epigenetic signatures. Notwithstanding profound knowledge on mechanistic details of DNA replication, major problems of genome replication have remained unresolved. In this perspective article, we consider the accessibility of replication machines to all DNA sequences in due course, the maintenance of functionally important positional and structural features of chromatid domains during replication, and the rapid transition of CTs into prophase chromosomes with two chromatids. We illustrate this problem with EdU pulse-labeling (10 min) and chase experiments (80 min) performed with mouse myeloblast cells. Following light optical serial sectioning of nuclei with 3D structured illumination microscopy (SIM), seven DNA intensity classes were distinguished as proxies for increasing DNA compaction. In nuclei of cells fixed immediately after the pulse-label, we observed a relative under-representation of EdU-labeled DNA in low DNA density classes, representing the active nuclear compartment (ANC), and an over-representation in high density classes representing the inactive nuclear compartment (INC). Cells fixed after the chase revealed an even more pronounced shift to high DNA intensity classes. This finding contrasts with previous studies of the transcriptional topography demonstrating an under-representation of epigenetic signatures for active chromatin and RNAPII in high DNA intensity classes and their over-representation in low density classes. We discuss these findings in the light of current models viewing CDs either as structural chromatin frameworks or as phase-separated droplets, as well as methodological limitations that currently prevent an integration of this contrasting evidence for the spatial nuclear topography of replication and transcription into a common framework of the dynamic nuclear architecture.

cell biology↗

bioRxiv2022

True-to-scale DNA-density maps correlate with major accessibility differences between active and inactive chromatin

Chromatin compaction differences may have a strong impact on accessibility of individual macromolecules and macromolecular assemblies to their DNA target sites. Estimates based on fluorescence microscopy with conventional resolution, however, suggested only modest compaction differences ([~]2-10x) between active and inactive nuclear compartments (ANC and INC). Here, we present maps of nuclear landscapes with true-to-scale DNA-densities, ranging from <5 Mbp/{micro}m3 to >300 Mbp/{micro}m3. Maps were generated from individual human and mouse cell nuclei with single-molecule localization microscopy at [~]20 nm lateral and [~]100 nm axial resolution and supplemented by electron spectroscopic imaging. Microinjection of fluorescent nanobeads with sizes corresponding to macromolecular assemblies for transcription and replication into nuclei of living cells, demonstrated their localization and movements within the ANC and exclusion from the INC.

cell biology↗

bioRxiv2018

A conformational sensor based on genetic code expansion reveals an autocatalytic component in EGFR activation

Epidermal growth factor receptor (EGFR) activation by growth factors (GFs) relies on dimerization and allosteric activation of its intrinsic kinase activity, resulting in trans-phosphorylation of tyrosines on its C-terminal tail. While structural and biochemical studies identified this EGF-induced allosteric activation, imaging collective EGFR activation in cells and molecular dynamics simulations pointed at additional catalytic EGFR activation mechanisms. To gain more insight in EGFR activation mechanisms in living cells, we developed a Forster Resonance Energy Transfer (FRET) based conformational EGFR indicator (CONEGI) using genetic code expansion that reports on conformational transitions in the EGFR activation loop. Comparing conformational transitions, self-association and auto-phosphorylation of CONEGI and its Y845F mutant revealed that Y845 phosphorylation induces a catalytically active conformation in EGFR monomers. This conformational transition depends on EGFR kinase activity and auto-phosphorylation on its C-terminal tail, generating a looped causality that leads to autocatalytic amplification of EGFR phosphorylation at low EGF dose.

molecular biology↗