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Dryad

DNA-stimulated liquid-liquid phase separation by eukaryotic topoisomerase II modulates catalytic function

Cite this dataset

Berger, James et al. (2022). DNA-stimulated liquid-liquid phase separation by eukaryotic topoisomerase II modulates catalytic function [Dataset]. Dryad. https://doi.org/10.5061/dryad.z08kprrgc

Abstract

Type II topoisomerases modulate chromosome supercoiling, condensation, and catenation by moving one double-stranded DNA segment through a transient break in a second duplex. How DNA strands are chosen and selectively passed to yield appropriate topological outcomes – e.g., decatenation vs. catenation – is poorly understood. Here we show that at physiological enzyme concentrations, eukaryotic type IIA topoisomerases (topo IIs) readily coalesce into condensed bodies. DNA stimulates condensation and fluidizes these assemblies to impart liquid-like behavior. Condensation induces both budding yeast and human topo IIs to switch from DNA unlinking to active DNA catenation, and depends on an unstructured C-terminal region, the loss of which leads to high levels of knotting and reduced catenation. Our findings establish that local protein concentration and phase separation can regulate how topo II creates or dissolves DNA links, behaviors that can account for the varied roles of the enzyme in supporting transcription, replication, and chromosome compaction.

Methods

Microscopy data was collected on an LSM 800 Confocal microscope, processed through FIJI, and all data were analyzed in R studio. Biochemical assays were run on a gel, imaged, and images were edited in Adobe illustrator. All statistical analysis was done using PRISM. 

Usage notes

FIJI, R Studio, Adobe Illustrator, Prism.

Funding

National Institute of General Medical Sciences, Award: T32-GM7445-43

National Cancer Institute, Award: R35-CA263778