Illuminating radial genome organization in the nucleus

Introduction

In exciting preliminary experiments leveraging our GPSeq method, we discovered that the genome of mammalian cells in interphase folds into a steep radial gradient of guanine and cytosine (GC) density, which seems to persist at the level of individual mitotic chromosomes. However, we still lack a fundamental understanding of how this higher-order 3D genome architecture is established and what its functional implications are.

Hypothesis

Here, I go beyond the state-of-the-art and propose that the observed steep radial GC-gradient is a universal design principle of the radial arrangement of the genome in the nucleus—which I call the radiality principle—that provides a biophysical framework for spatially orchestrating key nuclear processes, beyond gene expression regulation.

Objectives

To test this hypothesis, in this project I pursue five objectives:

1. Development of GP-C: First, we develop a novel approach (GP-C) for high-resolution single-cell 3D genome reconstructions to study whether the radial GC-gradient is indeed a universal property of nuclei across different species and cell types.

2. Monitoring Genome Radiality: Next, we apply GP-C together with RNA-seq to monitor genome radiality and concurrent gene expression changes as cells undergo karyotype rewiring or significant epigenetic perturbations.

3. Probing Mitotic Chromosomes: In parallel, we develop innovative approaches to probe the internal structure of mitotic chromosomes and model how genome radiality is reorganized as cells traverse mitosis.

4. Expanding Findings on DNA-RNA Contact Hubs: We then expand our preliminary finding of large-scale DNA-RNA contact hubs that seem to shape cell-type specific radiality landscapes by opposing the radial GC-gradient.

5. Profiling Nuclear Proteins: Finally, we pioneer methods for profiling nuclear proteins radially and apply them to test the bold hypothesis that the radiality principle provides a blueprint for organizing numerous nuclear processes.

Conclusion

This project aims at conclusively addressing long-standing questions in the field of 3D genome biology and proposes novel mechanisms of nuclear function regulation.