Exploring Mammalian Genome within Phase-Separated Nuclear Bodies: Experimental Methods and Implications for Gene Expression

Abstract

The importance of genome organization at the supranucleosomal scale in the control of gene expression is increasingly recognized today. In mammals, Topologically Associating Domains (TADs) and the active/inactive chromosomal compartments are two of the main nuclear structures that contribute to this organization level. However, recent works reviewed here indicate that, at specific loci, chromatin interactions with nuclear bodies could also be crucial to regulate genome functions, in particular transcription. They moreover suggest that these nuclear bodies are membrane-less organelles dynamically self-assembled and disassembled through mechanisms of phase separation. We have recently developed a novel genome-wide experimental method, High-salt Recovered Sequences sequencing (HRS-seq), which allows the identification of chromatin regions associated with large ribonucleoprotein (RNP) complexes and nuclear bodies. We argue that the physical nature of such RNP complexes and nuclear bodies appears to be central in their ability to promote efficient interactions between distant genomic regions. The development of novel experimental approaches, including our HRS-seq method, is opening new avenues to understand how self-assembly of phase-separated nuclear bodies possibly contributes to mammalian genome organization and gene expression.

The regulatory landscape of the Dlx gene system in branchial arches: Shared characteristics among Dlx bigene clusters and evolution

“…The intergenic leading enhancers (I12a, I56i, and I37-2) and the distal leading enhancers (URE2, eDlx#18, eDlx#19, and mU4-Tad3) are physically located relatively close by CTCF-cohesin loop (Figure 4). Upon activation, transcription factors (TFs) accumulate on the leading enhancers and could form small seeds of phase-separated molecular condensates consisting of TFs and co-activators (Hnisz et al, 2017, Shin et al, 2018, Lesne et al, 2019.…”

Suppression of liquid-liquid phase separation by 1,6-hexanediol partially compromises the 3D genome organization in living cells

“…To investigate the role played by LLPS in spatial genome organization, we used human HeLa cells that were treated with 1,6-HD. This aliphatic alcohol is predicted to disrupt weak hydrophobic interactions, both in vitro and in vivo, disassembling LLPS-dependent macromolecular condensates (Kroschwald et al 2017;Alberti et al 2019;Lesne et al 2019).…”

“…Molecular condensates depended purely on phase separation driven by electrostatic interactions are expected to be unaffected by 1,6-HD treatment (Lesne et al 2019). Nevertheless, 1,6-HD is the only available tool to test in vivo the contribution of LLPS in the assembly of macromolecular assemblies at this moment (Kroschwald et al 2017).…”

Three-dimensional genome rewiring during the development of antibody-secreting cells

“…Subsequent Hi-C studies with greater resolution further divided the compartments into six different sub-compartments with distinct histone marks and epigenetic properties [15]. It is therefore speculated that the compartmentalisation is mediated by the similar physical properties of different chromatin states, possibly via phase separation [27][28][29][30][31] (Figure 1B). In support of this, several studies have demonstrated HP1α and other heterochromatic H3K9me3 recognition complexes are able to form phase-separated liquid droplets, containing other macromolecules, such as DNA [29,32,33].…”