Biology subjects

Cramard, J.

Publications and source records attributed to Cramard, J..

7 recordsLinked to original sources

bioRxiv2025

Loop extrusion creates rare, long-lived encounters underlying enhancer-promoter communication

Enhancers regulate transcription from distal genomic positions, but how their spatial encounters with promoters drive activation remains unclear. Using polymer simulations and high-resolution live-cell microscopy, we identify rare but long-lived chromatin encounters arising from cohesin-mediated loop extrusion. These events occur when cohesin loads near the midpoint between two loci and extrudes them through a defined spatial radius, producing encounter durations that exceed those of random collisions. We show that such encounters explain observed nonlinear relationships between contact probability and transcription, and accurately predict transcriptional changes upon perturbation of cohesin or its cofactors. Our findings support a time-gated model of distal enhancer-promoter communication in which only rare, long-lived and mostly extrusion-driven encounters are productive, offering a unifying framework for how chromosome dynamics control transcription in single cells.

molecular biology↗

bioRxiv2025

Enhancer control of promoter activity and variability via frequency modulation of clustered transcriptional bursts

Gene expression in mammalian cells is controlled by enhancers that are often dispersed across large cis-regulatory landscapes around a promoter. Yet how enhancers determine transcription of their target genes, and how this depends on their relative position inside a cis-regulatory landscape remains unclear. Here we use live-cell imaging to track the activity of a promoter under the control of the same enhancer, but inserted at different positions across a simplified regulatory landscape with minimal complexity. Combined with mathematical modeling, this reveals that RNA production from the promoter occurs in clusters of transcriptional bursts, with enhancer position controlling the frequency at which such clusters appear. This results in bursts being more frequent and occurring more uniformly across cells when the enhancer is genomically close to the promoter than when it is located at large genomic distance. Mathematical modeling further indicates that the enhancer modulates the promoters ability to transition from its basal transcriptional state to a regime where clusters of bursts become more frequent. Our results challenge existing models of mammalian promoter operation, and reveal that enhancer position within a cis-regulatory landscape critically controls the timing and variability of transcriptional output in single cells.

molecular biology↗

bioRxiv2022

Live-cell imaging and physical modeling reveal control of chromosome folding dynamics by cohesin and CTCF

Physical proximity between genomic sequences in mammalian chromosomes controls key biological processes such as transcriptional regulation and DNA repair. Yet it is currently unknown if chromosomal contacts are rare and stable or instead frequent and dynamic, and how they depend on the loop extrusion activity of cohesin or barriers such as CTCF. By imaging chromosomal locations at high spatial and temporal resolution over several hours in living cells, we show that sequences within topological associating domains (TADs) frequently come into physical proximity during the course of a cell cycle and remain close to each other only for a few minutes. Such contacts become nonetheless substantially longer and more frequent in the presence of convergent CTCF sites, resulting in a suppression of variability in chromosome folding in single cells across time. Supported by physical models of chromosome dynamics, our data additionally suggests that individual CTCF-anchored loops last around 10 minutes. The estimates of chromosomal contact dynamics in our study provide a novel quantitative framework to link chromosome structure to function and show that cohesin and CTCF stabilize otherwise highly dynamic chromosome structures to facilitate selected subsets of chromosomal interactions.

molecular biology↗

bioRxiv2021

Nonlinear control of transcription through enhancer-promoter interactions

Chromosome structure in mammals is thought to regulate transcription by modulating the three-dimensional interactions between enhancers and promoters, notably through CTCF-mediated interactions and topologically associating domains (TADs)1-4. However, how chromosome interactions are actually translated into transcriptional outputs remains unclear. To address this question we use a novel assay to position an enhancer at a large number of densely spaced chromosomal locations relative to a fixed promoter, and measure promoter output and interactions within a genomic region with minimal regulatory and structural complexity. Quantitative analysis of hundreds of cell lines reveal that the transcriptional effect of an enhancer depends on its contact probabilities with the promoter through a non-linear relationship. Mathematical modeling and validation against experimental data further provide evidence that nonlinearity arises from transient enhancer-promoter interactions being memorized into longer-lived promoter states in individual cells, thus uncoupling the temporal dynamics of interactions from those of transcription. This uncovers a potential mechanism for how enhancers control transcription across large genomic distances despite rarely meeting their target promoters, and for how TAD boundaries can block distal enhancers. We finally show that enhancer strength additionally determines not only absolute transcription levels, but also the sensitivity of a promoter to CTCF-mediated functional insulation. Our unbiased, systematic and quantitative measurements establish general principles for the context-dependent role of chromosome structure in long-range transcriptional regulation.

molecular biology↗

bioRxiv2020

Live-cell 3D single-molecule tracking reveals how NuRD modulates enhancer dynamics

Enhancer-promoter dynamics are crucial for the spatiotemporal control of gene expression, but it remains unclear whether these dynamics are controlled by chromatin regulators, such as the nucleosome remodelling and deacetylase (NuRD) complex. The NuRD complex binds to all active enhancers to modulate transcription and here we use Hi-C experiments to show that it blurs TAD boundaries and increases the proximity of intermediate-range (~1 Mb) genomic sequences and enhancer-promoter interactions. To understand whether NuRD alters the dynamics of 3D genome structure, we developed an approach to segment and extract key biophysical parameters from 3D single-molecule trajectories of the NuRD complex determined using live-cell imaging. Unexpectedly, this revealed that the intact NuRD complex decompacts chromatin structure and makes NuRD-bound enhancers move faster, increasing the overall volume of the nucleus that these key regulatory regions explore. Interestingly, we also uncovered a rare fast-diffusing state of NuRD bound enhancers that exhibits directed motion. The NuRD complex reduces the amount of time that enhancers remain in this fast-diffusing state, which we propose might otherwise re-organise enhancer-promoter proximity. Thus, we uncover an intimate connection between a chromatin remodeller and the spatial dynamics of the local regions of the genome to which it binds.

cell biology↗

bioRxiv2020

Differential regulation of lineage commitment in human and mouse primed pluripotent stem cells by NuRD

Differentiation of mammalian pluripotent cells involves large-scale changes in transcription and, among the molecules that orchestrate these changes, chromatin remodellers are essential to initiate, establish and maintain a new gene regulatory network. The NuRD complex is a highly conserved chromatin remodeller which fine-tunes gene expression in embryonic stem cells. While the function of NuRD in mouse pluripotent cells has been well defined, no study yet has defined NuRD function in human pluripotent cells. We investigated the structure and function of NuRD in human induced pluripotent stem cells (hiPSCs). Using immunoprecipitation followed by mass-spectrometry in hiPSCs and in naive or primed mouse pluripotent stem cells, we find that NuRD structure and biochemical interactors are generally conserved. Using RNA sequencing, we find that, whereas in mouse primed stem cells and in mouse naive ES cells, NuRD is required for an appropriate level of transcriptional response to differentiation signals, hiPSCs require NuRD to initiate these responses. This difference indicates that mouse and human cells interpret and respond to induction of differentiation differently. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=109 SRC="FIGDIR/small/935544v1_ufig1.gif" ALT="Figure 1"> View larger version (21K): org.highwire.dtl.DTLVardef@1f68187org.highwire.dtl.DTLVardef@b1e9f3org.highwire.dtl.DTLVardef@766e9forg.highwire.dtl.DTLVardef@158c57b_HPS_FORMAT_FIGEXP M_FIG NuRD acts like a conductor in an orchestra. A. In the presence of NuRD (pink blob figure, centre) differentiation occurs in an ordered fashion in both mouse (left) and human (right) ES cells. Gene expression changes in both cell types are tightly controlled with down-regulation of pluripotency genes and up-regulation of lineage appropriate genes. This is akin to a group of musicians producing musical notes in the right order and at the right amplitude to create a coherent piece of music. B. Loss of "the conductor" NuRD results in increased transcriptional noise in both systems, indicated here as a low-level blanket of sound in both systems. Consequences of MBD3/NuRD loss differs between human and mouse ES cells. In mouse ES cells, differentiation cues lead to some down-regulation of pluripotency genes and incomplete progression along a lineage appropriate pathway. This is like musicians who know that they should be making music but who lose their way without a conductors influence. In human iPS cells the background level of noise without NuRD results in a lack of order to gene expression changes in response to differentiation. The noise from these "musicians" would be truly awful. C_FIG

developmental biology↗

bioRxiv2018

MTA proteins act redundantly within the NuRD complex to ensure fidelity of ES cell lineage commitment

Multiprotein chromatin remodelling complexes show remarkable conservation of function amongst metazoans, even though components present in invertebrates are often present as multiple paralogous proteins in vertebrate complexes. In some cases these paralogues specify distinct biochemical and/or functional activities in vertebrate cells. Here we set out to define the biochemical and functional diversity encoded by one such group of proteins within the mammalian Nucleosome Remodelling and Deacetylation (NuRD) complex: Mta1, Mta2 and Mta3. We find that, in contrast to what has been described in somatic cells, MTA proteins are not mutually exclusive within ES cell NuRD and, despite subtle differences in chromatin binding and biochemical interactions, serve largely redundant functions. Nevertheless, ES cells lacking all three MTA proteins represent a complete NuRD null and are viable, allowing us to identify a previously undetected function for NuRD in maintaining differentiation trajectory during early stages of lineage commitment.

developmental biology↗