Biology subjects

Varamogianni-Mamatsi, V.

Publications and source records attributed to Varamogianni-Mamatsi, V..

5 recordsLinked to original sources

bioRxiv2024

Synergistic and antagonistic activities of IRF8 and FOS enhancer pairs during an immune cell fate switch

Cell-fate instructive genes tend to be regulated by large clusters of enhancers. Whether and how individual enhancers within such clusters cooperate in regulating gene expression is poorly understood. We have previously developed a computational method, SEGCOND, that identifies hubs consisting of enhancer clusters and their target genes, termed Putative Transcriptional Condensates (PTCs). Using SEGCOND, we identified PTCs in a CEBPA-induced B-cell to macrophage transdifferentiation system. We found them to be enriched for highly expressed, lineage-restricted genes and to associate with BRD4, a component of transcriptional condensates. Here we performed single and combinatorial deletions of enhancers within two active PTCs after transdifferentiation is induced, harboring IRF8 and FOS. Two enhancers within the IRF8 PTC were found to form a backup mechanism when combined, safeguarding IRF8 expression and transdifferentiation kinetics. Unexpectedly, two individual enhancers within the FOS PTC antagonize each other at Day 1 of transdifferentiation, delaying the conversion of B-cells to macrophages and reducing FOS expression, but cooperate to increase FOS levels in Day 7 induced cells. Our results reveal differentiation stage-specific, complex interactions, between individual enhancers within a cluster.

genomics↗

bioRxiv2024

Two distinct chromatin modules regulate proinflammatory gene expression

Various mechanisms have been proposed to explain gene activation and co-regulation, including enhancer-promoter interactions via chromatin looping and the enrichment of transcription factors into hubs or condensates. However, these conclusions often stem from analyses of individual loci, and genome-wide studies exploring mechanistic differences with coupled gene expression are lacking. In this study, we dissected the proinflammatory gene expression program induced by TNF in primary human endothelial cells using NGS- and imaging-based techniques. Our findings, enabled by our novel RWireX approach for single-cell ATAC-seq analysis, revealed two distinct regulatory chromatin modules: autonomous links of co-accessibility (ACs) between separated sites, and domains of contiguous co-accessibility (DCs) with increased local transcription factor binding. Genes in ACs and DCs exhibited different transcriptional bursting kinetics, highlighting the existence of two structurally and functionally distinct regulatory chromatin modules in the proinflammatory response. These findings provide a novel mechanistic framework for understanding how cells achieve rapid and precise gene expression control. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=71 SRC="FIGDIR/small/606159v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1f5f7e8org.highwire.dtl.DTLVardef@166b4d7org.highwire.dtl.DTLVardef@1ea9206org.highwire.dtl.DTLVardef@118d534_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LITwo distinct, non-mutually exclusive chromatin modules, ACs and DCs, that regulate proinflammatory gene expression were identified based on deep scATAC-seq. C_LIO_LIACs represent long-range genomic interactions with regulation occurring more by transcription burst frequency. C_LIO_LIDCs are regions of increased local transcription factor binding that can modulate transcription burst size. C_LIO_LIThe AC/DC model integrates sequencing-based evidence for chromatin looping with microscopy observations of transcription factor hubs/condensates into a unified model. C_LIO_LIOur findings provide a novel framework for understanding how cells achieve rapid and precise gene expression control. C_LI

genomics↗

bioRxiv2024

Senescent cells cluster CTCF on nuclear speckles to sustain their splicing program

Senescence --the endpoint of replicative lifespan for normal cells-- is established via a complex sequence of molecular events. One such event is the dramatic reorganization of CTCF into senescence-induced clusters (SICCs). However, the molecular determinants, genomic consequences, and functional purpose of SICCs remained unknown. Here, we combine functional assays, super-resolution imaging, and 3D genomics with computational modelling to dissect SICC emergence. We establish that the competition between CTCF-bound and non-bound loci dictates clustering propensity. Upon senescence entry, cells repurpose SRRM2 --a key component of nuclear speckles-- and BANF1 --a molecular glue for chromosomes-- to cluster CTCF and rewire genome architecture. This CTCF-centric reorganization in reference to nuclear speckles functionally sustains the senescence splicing program, as SICC disruption fully reverts alternative splicing patterns. We therefore uncover a new paradigm, whereby cells translate changes in nuclear biochemistry into architectural changes directing splicing choices so as to commit to the fate of senescence. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/603680v1_ufig1.gif" ALT="Figure 1"> View larger version (85K): org.highwire.dtl.DTLVardef@cfb561org.highwire.dtl.DTLVardef@8cfdf3org.highwire.dtl.DTLVardef@7485d5org.highwire.dtl.DTLVardef@1fe9051_HPS_FORMAT_FIGEXP M_FIG C_FIG HIGHLIGHTSO_LIHMGB2-bound loci compete with CTCF-bound ones for nuclear speckle association C_LIO_LISenescent cells repurpose SRRM2 and BANF1 to cluster CTCF on speckles C_LIO_LIBANF1 is essential, but not sufficient for CTCF clustering C_LIO_LIThe SRRM2 RNA-binding domain directs CTCF clustering C_LIO_LISICCs rewire chromatin positioning to sustain the senescence splicing program C_LI

cell biology↗

bioRxiv2023

Increasingly efficient chromatin binding of cohesin and CTCF supports chromatin architecture formation during zebrafish embryogenesis

The three-dimensional folding of chromosomes is essential for nuclear functions such as DNA replication and gene regulation. The emergence of chromatin architecture is thus an important process during embryogenesis. To shed light on the molecular and kinetic underpinnings of chromatin architecture formation, we characterized biophysical properties of cohesin and CTCF binding to chromatin and their changes upon cofactor depletion using single-molecule imaging in live developing zebrafish embryos. We found that chromatin-bound fractions of both cohesin and CTCF increased significantly between the 1000-cell and shield stages, which we could explain through changes in both their association and dissociation rates. Moreover, increasing binding of cohesin restricted chromatin motion, potentially via loop extrusion, and showed distinct stage-dependent nuclear distribution. Polymer simulations with experimentally derived parameters recapitulated the experimentally observed gradual emergence of chromatin architecture. Our findings suggest a kinetic framework of chromatin architecture formation during zebrafish embryogenesis.

biophysics↗

bioRxiv2023

Rapid and synchronous chemical induction of replicative-like senescence via a small molecule inhibitor

Cellular senescence is now acknowledged as a key contributor to organismal ageing and late-life disease. Although popular, the study of senescence in vitro can be complicated by the prolonged and asynchronous timing of cells committing to it and its paracrine effects. To address these issues, we repurposed the small molecule inhibitor inflachromene (ICM) to induce senescence to human primary cells. Within six days of treatment with ICM, senescence hallmarks, including the nuclear eviction of HMGB1 and -B2, are uniformly induced across IMR90 cell populations. By generating and comparing various high throughput datasets from ICM-induced and replicative senescence, we uncovered significant similarity of the two states. Notably though, ICM suppresses the proinflammatory secretome associated with senescence, thus alleviating most paracrine effects. In summary, ICM induces a senescence-like phenotype rapidly and synchronously thereby allowing the study of its core regulatory program without any confounding heterogeneity.

genomics↗