The spatial organization of gene regulation in embryonic development.

Introduction

Master transcription factors (TFs) cooperate with chromatin in regulating genomic activity in animal development and stress response. Local clustering of TFs into dense, sub-micrometer size condensates is emerging as a key feature of transcriptional regulation. This includes intranuclear condensates formed by pioneer TFs during development, transient clusters of RNA Polymerase II complex, and nuclear stress bodies formed during a heat shock.

Background

Despite the ubiquity of these assemblies, we know little about the biophysical mechanism of their formation or physiological function. The last years have seen the development of new frameworks to study such transient assemblies, including various types of phase transitions and tools to probe and modulate them. However, this work has been limited mainly to cell culture and in vitro experiments that failed to incorporate the vital role of chromatin in organizing gene regulation.

Proposed Strategy

I propose a cross-disciplinary strategy consisting of novel in vitro assays and functional studies in a nematode, Caenorhabditis elegans, to study the spatial organization of transcription in embryonic development and stress response. The specific aims are:

1. Understand the physiological relevance of transcriptional condensates in animal development and stress response.
2. Determine the molecular composition and regulators of transcriptional condensates.
3. Dissect the mechanism of transcriptional condensate formation using a novel chromatin carpet assay.

Methodology

We will use state-of-the-art microscopy-based tools to investigate the formation and function of nuclear condensates in developing animal embryos. Additionally, we will develop innovative assays to probe the condensation of TFs on the surface of purified native chromatin.

Expected Outcomes

The obtained results will provide unprecedented insight into the composition, assembly mechanism, and physiological relevance of biomolecular condensates formed by the transcriptional apparatus during differentiation and stress response.