Quantitative multimodal pulse-and-label time-resolved chromatin maps

Introduction

Chromatin packages the eukaryotic genome in a highly dynamic fashion, with dramatic structural changes during every cell cycle. At the same time, chromatin provides remarkable stability for transcriptional regulation and genome organization, for example in maintaining gene expression programs and lineage identity during complex organismal development. A mechanism to propagate information through ‘disruptive’ transitions in the cell cycle, namely DNA replication and mitosis, is key in maintaining heritable, so-called ‘epigenetic’ chromatin states.

Motivation

Proteins that build and interact with chromatin, foremost histones, are much more than static architectural components. This motivates the development of time-resolved quantitative assays in the living cell, which will allow capturing dynamic features of chromatin states over timescales from minutes to days.

Methodology

Building on a synthetic biology toolbox, a quantitative multimodal pulse-and-label strategy will be developed. This approach will involve:

1. Following protein populations in both time and subcellular/genomic space.
2. Capturing dynamic protein-protein interaction networks.
3. Creating chromatin maps.

Techniques

The project will utilize state-of-the-art quantitative biochemical, imaging, genomics, and proteomics readouts, including single-cell readouts. These data will feed into mathematical models to:

• Describe the dynamics and potential heterogeneity/stochasticity of the system under study.
• Predict its response to perturbations.
• Guide mechanistic hypotheses.

Objectives

The project will systematically decipher mechanisms for propagating epigenetic chromatin states, starting with the fundamental rules of histone inheritance through replication and mitosis. Additional levels of complexity introduced through:

• Chromatin remodeling activities.
• Nucleosome turnover.
• Histone exchange.

will be integrated. Finally, the dynamics underlying developmental chromatin state transitions, including asymmetric cell fate decisions, will be resolved.