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Tracking live-cell single-molecule dynamics enables measurements of heterochromatin-associated protein-protein interactions

ABSTRACT: Tracking live-cell single-molecule dynamics enables measurements of heterochromatin-associated protein-protein interactions

PROVIDER: PRJNA1097604 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR28592483_1.fastq.gz	Fastqsanger.gz
	SRR28592483_2.fastq.gz	Fastqsanger.gz
	SRR28592484_1.fastq.gz	Fastqsanger.gz
	SRR28592484_2.fastq.gz	Fastqsanger.gz
	SRR28592485_1.fastq.gz	Fastqsanger.gz

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Tracking live-cell single-molecule dynamics enables measurements of heterochromatin-associated protein-protein interactions

Project description:Visualizing and measuring molecular-scale interactions in living cells represents a major challenge, but recent advances in microscopy are bringing us closer to achieving this goal. Single-molecule super-resolution microscopy enables high-resolution and sensitive imaging of the positions and movement of molecules in living cells. HP1 proteins are important regulators of gene expression because they selectively bind and recognize H3K9 methylated (H3K9me) histones to form heterochromatin-associated protein complexes that silence gene expression. Here, we extended live-cell single-molecule tracking studies in fission yeast to determine how HP1 proteins interact with their binding partners in the nucleus. We measured how genetic perturbations that affect H3K9me alter the diffusive properties of HP1 proteins and each of their binding partners based on which we inferred their most likely interaction sites. Our results indicate that H3K9me promotes specific complex formation between HP1 proteins and their interactors in a spatially restricted manner, while attenuating their ability to form off-chromatin complexes. As opposed to being an inert platform or scaffold to direct HP1 binding, our studies propose a novel function for H3K9me as an active participant in enhancing HP1-associated complex formation in living cells.

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

Project description: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.

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