Sensitivity of cohesin\u2013chromatin association to high-salt treatment corroborates non-topological mode of loop extrusion

Arkadiy K Golov,Sergey V Razin,Anastasia V Golova,Alexey A Gavrilov

doi:10.1186/s13072-021-00411-w

Abstract

Cohesin is a key organizer of chromatin folding in eukaryotic cells. The two main activities of this ring-shaped protein complex are the maintenance of sister chromatid cohesion and the establishment of long-range DNA–DNA interactions through the process of loop extrusion. Although the basic principles of both cohesion and loop extrusion have been described, we still do not understand several crucial mechanistic details. One of such unresolved issues is the question of whether a cohesin ring topologically embraces DNA string(s) during loop extrusion. Here, we show that cohesin complexes residing on CTCF-occupied genomic sites in mammalian cells do not interact with DNA topologically. We assessed the stability of cohesin-dependent loops and cohesin association with chromatin in high-ionic-strength conditions in G1-synchronized HeLa cells. We found that increased salt concentration completely displaces cohesin from those genomic regions that correspond to CTCF-defined loop anchors. Unsurprisingly, CTCF-anchored cohesin loops also dissipate in these conditions. Because topologically engaged cohesin is considered to be salt resistant, our data corroborate a non-topological model of loop extrusion. We also propose a model of cohesin activity throughout the interphase, which essentially equates the termination of non-topological loop extrusion with topological loading of cohesin. This theoretical framework enables a parsimonious explanation of various seemingly contradictory experimental findings.

Highlights

Cohesin is a large nuclear protein complex involved in maintaining the structure and integrity of the genome in virtually all eukaryotic cells [1, 2]
To find out what proportion, if any, of cohesin complexes topologically entrap DNA molecules during the G1 phase in mammalian cells, we analyzed the possibility of extracting chromatin-bound cohesin with a high-salt solution
We synchronized HeLa cells in the G1 phase, lysed them in isotonic buffer, and incubated permeabilized cells on ice in either isotonic buffer or in a buffer containing 0.5 M NaCl. This relatively high concentration of salt should cause the extraction of most non-histone DNA-binding proteins, whereas topologically bound cohesin rings should remain associated with long chromosomal DNA molecules [12,13,14]

Summary

Introduction

Cohesin is a large nuclear protein complex involved in maintaining the structure and integrity of the genome in virtually all eukaryotic cells [1, 2]. To determine whether cohesin complexes mediating LE are bound to chromatin in a topological manner, we analyzed the salt-sensitivity of cohesin and CTCFanchored DNA loops in the G1 cell cycle phase. Results and discussion To find out what proportion, if any, of cohesin complexes topologically entrap DNA molecules during the G1 phase in mammalian cells, we analyzed the possibility of extracting chromatin-bound cohesin with a high-salt solution.

Results

Conclusion