The Secret Life of Chromosome Loops upon DNA Double-Strand Break

Abstract

DNA double-strand breaks (DSBs) are harmful lesions that severely challenge genomic integrity, and recent evidence suggests that DSBs occur more frequently on the genome than previously thought. These lesions activate a complex and multilayered response called the DNA damage response, which allows to coordinate their repair with the cell cycle progression. While the mechanistic details of repair processes have been narrowed, thanks to several decades of intense studies, our knowledge of the impact of DSB on chromatin composition and chromosome architecture is still very sparse. However, the recent development of various tools to induce DSB at annotated loci, compatible with next-generation sequencing-based approaches, is opening a new framework to tackle these questions. Here we discuss the influence of initial and DSB-induced chromatin conformation and the strong potential of 3C-based technologies to decipher the contribution of chromosome architecture during DSB repair.

Keywords: DNA double-strand breaks repair; DSB clustering; chromatin; topologically associating domains; γH2AX.

Graphical abstract

Fig. 1

Contribution of the initial chromatin conformation into γH2AX establishment and programmed DSB induction and repair. (A) The initial chromosome conformation may dictate γH2AX spreading following DSB induction. In this model, ATM, the main H2AX kinase is locally recruited at the DSB. Once bound, it is able to phosphorylate H2AX containing nucleosomes brought to its physical proximity, thanks to chromatin dynamics that takes place within the TAD. Sustained signaling and ATM activation eventually trigger the phosphorylation of H2AX on the entire TAD. In this model, γH2AX distribution, as observed by ChIP-seq, should mimic the 3D chromatin conformation. (B) Chromosome conformation is critical during meiotic breaks formation by Spo11. During prophase, meiotic chromosomes are strongly reorganized with the formation of DNA loops anchored to a proteinaceous axis. Spo11 generates DSBs within DNA loops, which can further pair with the homologous chromosome in order to produce crossover and to complete meiosis. The 3D chromatin structure and the chromosomal axis are required for both DSB production by Spo11 and to ensure the “homologous bias” (i.e., the choice of the homologous chromosome rather than the sister chromatid, as a template for HR). (C) Chromosome conformation is also critical for the rearrangements that occur on immunoglobulin loci, in order to generate immunoglobulin isotypes (class switch recombination (CSR)) and the antibody repertoire (VDJ recombination). For example, during CSR (shown here), the long-range physical interactions between switch (S) sequences on the heavy chain locus (Igh) allow two DSBs to be rejoined.

Fig. 2

DSB-induced modification(s) of the chromosome conformation in cis to the break. Following DSB production and γH2AX spreading, the 3D conformation of damaged TAD could also be modified, due to the binding of cohesin, CTCF or repair proteins with potential function in chromatin architecture such as 53BP1 and RIF1. The DSB-induced histones modifications (including γH2AX spreading), nucleosome loss or/and generation of single strand DNA (resection) may also collectively change the dynamics of chromatin within TADs. Altogether, these changes could translate in enhanced mobility and efficient DSB repair.

Fig. 3

Changes in chromosome conformation upon damage in trans such as during DSB clustering. Both live cell imaging and 3C-based methods allowed to demonstrate that multiple DSBs can coalesce together within a single γH2AX focus. However, the mechanisms that ensure clustering are unclear and may entail various pathways. (A) The nucleoskeleton (both polymerized actin and/or microtubules) could allow for DSB mobilization and clustering in a directional manner. (B) The cytoskeleton could also contribute to clustering thanks to the transmission of forces from cytoskeleton to chromatin via the LINC complex, embedded in the nuclear envelope. In this context, the forces transmitted to chromatin may trigger a general increase in chromatin dynamics, increasing the probability of γH2AX collision/clustering. (C) Finally, the chromatin landscape established following damage could allow for compartmentalization, thanks to phase separation.