Investigating DNA supercoiling in eukaryotic genomes.

Samuel Corless,Nick Gilbert

doi:10.1093/bfgp/elx007

Abstract

Supercoiling is a fundamental property of DNA, generated by polymerases and other DNA-binding proteins as a consequence of separating/bending the DNA double helix. DNA supercoiling plays a key role in gene expression and genome organization, but has proved difficult to study in eukaryotes because of the large, complex and chromatinized genomes. Key approaches to study DNA supercoiling in eukaryotes are (1) centrifugation-based or electrophoresis-based techniques in which supercoiled plasmids extracted from eukaryotic cells form a compacted writhed structure that migrates at a rate proportional to the level of DNA supercoiling; (2) in vivo approaches based on the preferential intercalation of psoralen molecules into under-wound DNA. Here, we outline the principles behind these techniques and discuss key discoveries, which have confirmed the presence and functional potential of unconstrained DNA supercoiling in eukaryotic genomes.

Highlights

A fundamental component of genome packaging and regulation is DNA supercoiling, a transition in DNA structure from a relaxed double helix to one that is over- or under-wound (Figure 1, Box 1)
DNA supercoiling plays a key role in gene expression and genome organization, but has proved difficult to study in eukaryotes because of the large, complex and chromatinized genomes
Key approaches to study DNA supercoiling in eukaryotes are (1) centrifugation-based or electrophoresis-based techniques in which supercoiled plasmids extracted from eukaryotic cells form a compacted writhed structure that migrates at a rate proportional to the level of DNA supercoiling; (2) in vivo approaches based on the preferential intercalation of psoralen molecules into under-wound DNA

Summary

Introduction

A fundamental component of genome packaging and regulation is DNA supercoiling, a transition in DNA structure from a relaxed double helix to one that is over- or under-wound (Figure 1, Box 1). Using centrifugation/electrophoresis and psoralen-based approaches over the past 35 years, the presence and distribution of DNA supercoiling in vivo in eukaryotes have been established, and recent advances have shown that unconstrained under-wound DNA is a general property of actively transcribed promoters and large-scale domains. These DNA structures are transient and can be disrupted by nicking the genome, inhibiting transcription or suppressing topoisomerase activity. These molecular probes will supplement current and future experimental observations determined using psoralen, to elucidate the presence and function of DNA supercoiling in the genomes of eukaryotes

Summary

Findings

Key Points