3D genome organization contributes to genome instability at fragile sites

Dan Sarni,Takayo Sasaki,David M Gilbert,Juan Carlos Rivera-Mulia,Michal Irony Tur-Sinai,Brian Magnuson,Batsheva Kerem,Mats Ljungman,Karin Miron

doi:10.1038/s41467-020-17448-2

Abstract

Common fragile sites (CFSs) are regions susceptible to replication stress and are hotspots for chromosomal instability in cancer. Several features were suggested to underlie CFS instability, however, these features are prevalent across the genome. Therefore, the molecular mechanisms underlying CFS instability remain unclear. Here, we explore the transcriptional profile and DNA replication timing (RT) under mild replication stress in the context of the 3D genome organization. The results reveal a fragility signature, comprised of a TAD boundary overlapping a highly transcribed large gene with APH-induced RT-delay. This signature enables precise mapping of core fragility regions in known CFSs and identification of novel fragile sites. CFS stability may be compromised by incomplete DNA replication and repair in TAD boundaries core fragility regions leading to genomic instability. The identified fragility signature will allow for a more comprehensive mapping of CFSs and pave the way for investigating mechanisms promoting genomic instability in cancer.

Highlights

1234567890():,; DNA replication occurs in a temporal order known as the replication timing (RT) program, which is tightly regulated to ensure the faithful duplication of the genome[1]
Unchallenged cells were compared with cells treated with a low concentration (0.2 μM) of APH for 24 h, commonly used to induce common fragile sites (CFSs) expression (Fig. 1a), cells were exposed to APH for at least one cell cycle
It is worth noting that 2/3 of RT-variable regions (387) were delayed by APH (Supplementary Data 1). These loci are of interest for further investigation of CFS instability as mild replication stress induces under-replication associated with CFS expression[11,19]

Summary

Introduction

1234567890():,; DNA replication occurs in a temporal order known as the replication timing (RT) program, which is tightly regulated to ensure the faithful duplication of the genome[1]. Dormant origins activation is not always sufficient and the perturbed replication induces chromosomal instability at specific regions termed common fragile sites (CFSs)[9]. CFS instability was found to be both cell-type- and stress inducerspecific, raising the possibility that fragility is driven by perturbed regulation of organized yet dynamic cellular programs, such as DNA replication, transcription and genome organization. We explore the effect of mild replication stress induced by the DNA polymerase inhibitor aphidicolin (APH) on genome-wide replication and transcription programs, using Repli-seq and Bru-seq, and integrate these data with Hi-C data from the 4DN consortium. The results reveal a multilayer chromosomal fragility signature upon replication stress, comprised of a combination of a TAD boundary overlapping an actively transcribed large gene with APH-induced RT delay. The results suggest that replication, transcription, and genome organization combined underlie CFS expression

Methods

Results

Conclusion