Make or brake it: On the roles of the cohesin acetylation cycle

Abstract

Human chromosomes are centimeters in length, but are structured to fit into a micrometer- scale nucleus. Key to this organization are structural maintenance of chromosomes complexes, which includes the cohesin complex. Cohesin is discovered for its essential role in holding together the sister chromatids from S-phase until mitosis. By tethering these two DNA molecules in trans, cohesin ensures faithful chromosome segregation. Cohesin also brings together DNA elements in cis through the formation of chromatin loops. Such loops form throughout interphase to provide structure to chromosomes. Cohesin thus shapes the 3D genome through the formation of loops and by holding together the sister chromatids. How cohesin performs these two distinct functions is only partially understood. In this thesis we set out to further understand the molecular mechanisms underlying these two processes, with a focus on the role of the cohesin acetylation cycle.

The acetylation of cohesin’s SMC3 subunit is a dynamic process that involves the acetyl transferases ESCO1 and ESCO2, and the deacetylase HDAC8. Earlier work showed that the acetylation of cohesin is important to establish and maintain sister chromatid cohesion by protecting cohesive cohesin against WAPL-mediated DNA release. In this thesis we show that the cohesin acetylation cycle also plays a role in 3D genome organization. ESCO1 restricts the length of chromatin loops, and of architectural stripes emanating from CTCF sites. HDAC8 conversely promotes the extension of such loops and stripes. This role in controlling loop length turns out to be distinct from the canonical role of cohesin acetylation that protects against WAPL-mediated DNA release. We reveal that acetylation rather controls cohesin’s interaction with PDS5A to restrict chromatin loop length. Our data supports a model in which this PDS5A-bound state acts as a brake that enables the pausing and restart of loop enlargement.

Cohesin complexes are present in two variants which differ in their stromal antigen (SA) subunit. Earlier work showed that these two variants, cohesinSA1 and cohesinSA2, have different acetylation levels, which has been proposed to be due to preferential acetylation of cohesinSA1. Using unbiased genetic screens and immunoprecipitation experiments we reveal that the ability of these complexes to become acetylated in fact is similar. We find that cohesinSA2 is rather preferentially deacetylated by HDAC8. This unexpected role for HDAC8 in controlling the acetylation levels of cohesinSA1 and cohesinSA2 highlights the importance of a dynamic cohesin acetylation cycle, and suggests that not only the acetylation of cohesin is tightly regulated, but also its deacetylation.

Our finding that the cohesin acetylation cycle controls the 3D genome in a WAPL-independent way inspired us to investigate whether a similar mechanism might be at play in sister chromatid cohesion. We find that acetyl transferases ESCO1 and ESCO2 indeed control sister chromatid cohesion beyond the protection against WAPL, but whether this is likewise dependent on controlling PDS5 binding remains to be further investigated. We also find that HDAC8 has a dual role in cohesion establishment. HDAC8 antagonizes the establishment reaction, and also promotes cohesion by recycling cohesin complexes to enable de novo SMC3 acetylation.