Biology subjects

Venit, T.

Publications and source records attributed to Venit, T..

8 recordsLinked to original sources

bioRxiv2026

Nuclear Myosin VI stabilises Ku-associated DNA ends during non-homologous end joining

DNA double-strand breaks (DSBs) require rapid signalling and physical stabilisation of broken DNA ends to preserve genome integrity. Here, we identify myosin VI (MVI) as an ATM-regulated component of the DSB response. DNA damage induces rapid nuclear accumulation and nanoscale reorganisation of MVI across multiple cell models, in an ATM-dependent manner. Pharmacological or genetic perturbation of MVI attenuates {gamma}H2AX signalling and disrupts Ku80 organisation, while DNA damage persists. This leads to increased sensitivity to cisplatin and bleomycin. Super-resolution imaging reveals spatial association of MVI with Ku80-containing repair structures, implicating MVI in non-homologous end joining (NHEJ). In a minimal reconstituted system, MVI and actin enhance the proximity of Ku70/80-bound DNA ends. Together, our findings identify MVI as a regulator of DSB repair that links ATM signalling to Ku-associated DNA-end stabilisation and suggest that targeting MVI may sensitise tumour cells to genotoxic therapy.

cancer biology↗

bioRxiv2025

Nuclear Myosin 1 links genomic architecture to adipose tissue remodeling, metabolic inflammation and obesity in mice

During adipogenesis, a metabolic shift from oxidative phosphorylation (OXPHOS) to aerobic glycolysis enables preadipocytes to meet the biosynthetic and energetic demands of differentiation. Nuclear myosin 1 (NM1), a chromatin-associated actomyosin motor that regulates transcription and chromatin accessibility, is essential for maintaining OXPHOS. Here, we identify NM1 as a key regulator of adipocyte differentiation and adipose tissue homeostasis. Integration of ATAC-seq and RNA-seq in NM1-deficient mouse embryonic fibroblasts revealed coordinated dysregulation of adipogenic genes (Insig1, Lipg, Fat1) and altered enhancer accessibility near Klf6, Foxo3, Smad5, and Gata4. NM1 knockout mesenchymal stem cells showed impaired adipogenic differentiation despite adipocyte hypertrophy. In vivo, NM1-deficient mice developed age-dependent visceral obesity with transcriptional reprogramming in white adipose tissue (WAT), including downregulation of adipogenesis and mitochondrial pathways, and activation of IFNG-, IL33-, and TNF-driven inflammation. Cross-species analysis revealed overlap with MYO1C-centered regulatory modules in human adipose tissue, implicating NM1/MYO1C in conserved chromatin-based control of adipose biology

cell biology↗

bioRxiv2024

Nuclear actin-dependent Meg3 expression suppresses metabolic genes by affecting the chromatin architecture at sites of elevated H3K27 acetylation levels

Three-dimensional organization of the eukaryotic genome is directly affected by the nuclear {beta}-actin pool that regulates enhancer function by affecting H3K27 acetylation levels. This actin-based mechanism, in turn, influences enhancer-dependent transcriptional regulation and plays a crucial role in driving gene expression changes observed upon compartment-switching. Using a combination of bulk RNA-seq and qPCR analyses performed on total RNA from WT mouse embryonic fibroblasts (MEFs), {beta}-actin heterozygous (HET) MEFs, and {beta}-actin KO MEFs, in this study we demonstrate that expression of several lncRNAs is directly affected by {beta}-actin depletion. Among these lncRNAs, Meg3 expression increases in a {beta}-actin dosage-dependent manner. Using ChIRP-seq, ChIRP-MS and f-RIP-qPCR, we show that {beta}-actin depletion leads to alterations in Meg3 genomic association. It also leads to Meg3 enrichment at or close to gene regulatory sites including enhancers and promoters concomitantly with increased H3K27 acetylation levels. At these sites, specific Meg3 association with H3K27 acetylation leads to loss of promoter-enhancer interactions as revealed by the Activity by Contact (ABC) model that builds on RNA-seq, H3K27acetylation ChIP-seq, ATAC-seq and HiC-seq obtained in WT and {beta}-actin KO MEFs. Results from metabolomics experiments in WT, HET and {beta}-actin KO MEFs show these mechanisms contribute to the repression of genes involved in metabolic biosynthetic pathways for chondroitin, heparan, dermatan sulfate, and phospholipases, hence impacting their synthesis. We propose that at sites of actin-dependent increase in H3K27acetylation levels Meg3 interferes with promoter-enhancer interactions, potentially impairing local genome organization (or DNA looping) and negatively regulating gene expression.

molecular biology↗

bioRxiv2023

Assessing nanobody interaction with SARS-CoV-2 Nsp9

The interaction between SARS-CoV-2 non-structural protein Nsp9 and the nanobody 2NSP90 was investigated by NMR spectroscopy using the paramagnetic perturbation methodology PENELOP (Paramagnetic Equilibrium vs Nonequilibrium magnetization Enhancement or LOss Perturbation). The Nsp9 monomer is an essential component of the replication and transcription complex (RTC) that reproduces the viral gRNA for subsequent propagation. Therefore preventing Nsp9 recruitment in RTC would represent an efficient antiviral strategy that could be applied to different coronaviruses, given the Nsp9 relative invariance. The NMR results were consistent with a previous characterization suggesting a 4:4 Nsp9-to-nanobody stoichiometry with the occurrence of two epitope pairs on each of the Nsp9 units that establish the inter-dimer contacts of Nsp9 tetramer. The oligomerization state of Nsp9 was also analyzed by molecular dynamics simulations and both dimers and tetramers resulted plausible. However a different distribution of the mapped epitopes on the tetramer surface with respect to the former 4:4 complex could also be possible, as well as different stoichiometries of the Nsp9-nanobody assemblies such as the 2:2 stoichiometry suggested by the recent crystal structure of the Nsp9 complex with 2NSP23 (PDB ID: 8dqu), a nanobody exhibiting essentially the same affinity as 2NSP90. The experimental NMR evidence, however, ruled out the occurrence in liquid state of the relevant Nsp9 conformational change observed in the same crystal structure.

biophysics↗

bioRxiv2023

Nanobodies against SARS-CoV-2 non-structural protein Nsp9 inhibit viral replication by targeting innate immunity

Nanobodies are emerging as critical tools for drug design. Several have been recently created to serve as inhibitors of SARS-Cov-2 entry in the host cell by targeting surface-exposed Spike protein. However, due to the high frequency of mutations that affect Spike, these nanobodies may not target it to their full potential and as a consequence, inhibition of viral entry may not be efficient. Here we have established a pipeline that instead targets highly conserved viral proteins that are made only after viral entry into the host cell when the SARS-Cov-2 RNA-based genome is translated. As proof of principle, we designed nanobodies against the SARS-CoV-2 non-structural protein Nsp9, required for viral genome replication. To find out if this strategy efficiently blocks viral replication, one of these anti-Nsp9 nanobodies, 2NSP23, previously characterized using immunoassays and NMR spectroscopy for epitope mapping, was encapsulated into lipid nanoparticles (LNP) as mRNA. We show that this nanobody, hereby referred to as LNP-mRNA- 2NSP23, is internalized and translated in HEK293 cells. We next infected HEK293-ACE2 cells with multiple SARS-CoV-2 variants and subjected them to LNP-mRNA-2NSP23 treatment. Analysis of total RNA isolated from infected cells treated or untreated with LNP-mRNA-2NSP23 using qPCR and RNA deep sequencing shows that the LNP-mRNA-2NSP23 nanobody protects HEK293-ACE2 cells and suppresses replication of several SARS-CoV-2 variants. These observations indicate that following translation, the nanobody 2NSP23 inhibits viral replication by targeting Nsp9 in living cells. We speculate that LNP-mRNA-2NSP23 may be translated into an innovative technology to generate novel antiviral drugs highly efficient across coronaviruses.

cell biology↗

bioRxiv2023

Nuclear Myosin 1 regulates platelet activation and immune response in mice

Cellular differentiation involves complex events associated with changes in cellular shape, function and proliferative capacity. This process is regulated by specific expression of multiple genes, which guide the cell through differentiation but also ensure proper function of terminal cell types. Over the last decade, the role of cellular metabolism in maintaining stem cells pluripotency and differentiation has been getting more attention due to a direct link between metabolic state and differentiation potential of cells. Since Nuclear Myosin 1 (NM1) deletion leads to a switch from oxidative phosphorylation to aerobic glycolysis and tumorigenesis in mice, here we asked if NM1 also contributes to cell differentiation. Indeed, analysis of metabolomic profiles and cytokine and chemokine levels upon NM1 depletion support a role in hematopoiesis. In an NM1 KO mouse model results from total blood cell count and bleeding assay show decreased erythropoiesis and thrombopoiesis as well as failure in hemostasis. Next, we explored the role of NM1 during the differentiation of hematopoietic progenitor stem cells to terminal blood cells by transcriptionally profiling bone marrow, spleen and peripheral blood from NM1 KO mice. We found that in bone marrow, NM1 deletion leads to overexpression of genes associated with glycolysis-dependent platelet activation and suppression of the innate immune system. These expression patterns are preserved and become more complex in spleen and peripheral blood. The study, therefore, provides insights into the underlying mechanisms of hematopoietic differentiation and activation of specific blood cell types and suggests NM1 as a potential therapeutic target for blood-related disorders.

cell biology↗

bioRxiv2022

Regulation of oxidative phosphorylation by Nuclear myosin 1 protects cells from metabolic reprogramming and tumorigenesis in mice

Metabolic reprogramming is one of the hallmarks of tumorigenesis. Using a combination of multi-omics, here we show that nuclear myosin 1 (NM1) serves as a key regulator of cellular metabolism. As part of the nutrient-sensing PI3K/Akt/mTOR pathway, NM1 forms a positive feedback loop with mTOR and directly affects mitochondrial oxidative phosphorylation (OXPHOS) via transcriptional regulation of mitochondrial transcription factors TFAM and PGC1. NM1 depletion leads to suppression of PI3K/Akt/mTOR pathway, underdevelopment of mitochondria inner cristae, and redistribution of mitochondria within the cell, which is associated with reduced expression of OXPHOS genes, decreased mitochondrial DNA copy number and deregulated mitochondrial dynamics. This leads to metabolic reprogramming of NM1 KO cells from OXPHOS to aerobic glycolysis and with a metabolomic profile typical for cancer cells, namely, increased amino acid-, fatty acid-, and sugar metabolism, and increased glucose uptake, lactate production, and intracellular acidity. We show that NM1 KO cells form solid tumors in a nude mouse model even though they have suppressed the PI3K/Akt/mTOR signaling pathway suggesting that the metabolic switch towards aerobic glycolysis provides a sufficient signal for carcinogenesis. We suggest that NM1 plays a key role as a tumor suppressor and that NM1 depletion may contribute to the Warburg effect at the early onset of tumorigenesis.

cell biology↗

bioRxiv2020

Nuclear myosin VI regulates the spatial organization of mammalian transcription initiation.

During transcription, RNA Polymerase II (RNAPII) is spatially organised within the nucleus into clusters that correlate with transcription activity. While this is a hallmark of genome regulation in mammalian cells, the mechanisms concerning the assembly, organisation and stability which underpin the function these transcription factories remain unknown. Here, we have used combination of single molecule imaging and genomic approaches to explore the role of nuclear myosin VI in the nanoscale organisation of RNAPII. We reveal that myosin VI acts as the molecular anchor that holds RNAPII into transcription factories. Perturbation of myosin VI leads to the disruption of RNAPII localisation, changes in chromatin organisation and subsequently a decrease in gene expression. Overall, we uncover the fundamental role of myosin VI in the spatial regulation of gene expression during the rapid response to changes in the cellular environment.

cell biology↗