Mouse Heterochromatin Adopts Digital Compaction States without Showing Hallmarks of HP1-Driven Liquid-Liquid Phase Separation

Fabian Erdel,Anne Rademacher,Rifka Vlijm,Jana Tünnermann,Lukas Frank,Robin Weinmann,Elisabeth Schweigert,Klaus Yserentant,Johan Hummert,Caroline Bauer,Sabrina Schumacher,Ahmad Al Alwash,Christophe Normand,Dirk-Peter Herten,Johann Engelhardt,Karsten Rippe

doi:10.1016/j.molcel.2020.02.005

Abstract

SummaryThe formation of silenced and condensed heterochromatin foci involves enrichment of heterochromatin protein 1 (HP1). HP1 can bridge chromatin segments and form liquid droplets, but the biophysical principles underlying heterochromatin compartmentalization in the cell nucleus are elusive. Here, we assess mechanistically relevant features of pericentric heterochromatin compaction in mouse fibroblasts. We find that (1) HP1 has only a weak capacity to form liquid droplets in living cells; (2) the size, global accessibility, and compaction of heterochromatin foci are independent of HP1; (3) heterochromatin foci lack a separated liquid HP1 pool; and (4) heterochromatin compaction can toggle between two “digital” states depending on the presence of a strong transcriptional activator. These findings indicate that heterochromatin foci resemble collapsed polymer globules that are percolated with the same nucleoplasmic liquid as the surrounding euchromatin, which has implications for our understanding of chromatin compartmentalization and its functional consequences.

Highlights

Cells partition their genome into distinct chromatin domains with specific functions
Mouse HP1a and GFP-HP1a Form Droplets in the Presence of DNA In Vitro It has recently been reported that Drosophila HP1a and human HP1a can form liquid droplets in vitro, which for human HP1a is promoted by phosphorylation, addition of DNA, or removal of salt (Larson et al, 2017; Strom et al, 2017; Wang et al, 2019; Zhang et al, 2019)
To test the ability of mouse heterochromatin protein 1 (HP1) to form droplets in vitro, we expressed and purified recombinant mouse HP1a and GFP-HP1a (Figure S1A) and mixed both proteins with a concentrated solution of fragmented salmon sperm DNA. Both HP1a and GFP-HP1a formed droplets (Figure 1A) as well as more irregular structures, which might correspond to assemblies of coagulated droplets (Figure S1B)

Summary

Introduction

Cells partition their genome into distinct chromatin domains with specific functions. A prominent example is that of the dense heterochromatin foci at silenced pericentric satellite repeats, which are called chromocenters because of their intense DAPI staining (Probst and Almouzni, 2008). Chromocenters contain elevated levels of DNA methylation, repressive histone modifications like trimethylation of histone H3 at lysine 9 (H3K9me3), and a specific set of proteins, including HP1, that can bind to H3K9me via its chromodomain (Bannister et al, 2001). The repressive heterochromatin state can spread to genomic sequences in proximity to pericentric repeats, leading to a phenomenon called position effect variegation (Elgin and Reuter, 2013). Because the accurate position and size of heterochromatin domains is critical for proper cell function (Fodor et al, 2010), it is crucial to understand how chromatin partitioning is faithfully accomplished

Methods

Results

Discussion

Conclusion