Recurrent DNA break clusters drive replication-stress-induced copy number variants and genome diversification.

Area of Science:

• Genetics
• Genomics
• Molecular Biology

Background:

• Copy number variants (CNVs) are linked to neurological disorders, psychiatric conditions, and cancer.
• The mechanisms generating CNVs, particularly recurrent ones, under replication stress are not fully understood.

Purpose of the Study:

• To investigate the role of recurrent DNA-break clusters (RDCs) in initiating CNVs during replication stress.
• To elucidate the mechanisms driving both recurrent and non-recurrent CNV formation.

Main Methods:

• Chemically induced replication stress in murine neural progenitor cells.
• Bulk and single-cell whole-genome sequencing.
• CRISPR/Cas9-mediated transcriptional suppression.
• Analysis of DNA repair pathways (Pol θ and NHEJ).

Main Results:

• Recurrent CNVs were enriched at late-replicating RDCs within actively transcribed genes.
• Single-cell sequencing revealed non-recurrent CNVs arising from RDCs during replication timing transitions.
• CRISPR/Cas9 suppression of RDCs abolished CNV generation.
• CNV formation mechanism varied based on DNA repair context (Pol θ dependence).

Conclusions:

• Recurrent DNA-break clusters (RDCs) are common initiating lesions for replication-stress-induced CNVs.
• RDCs drive genome diversification through both recurrent and non-recurrent CNV formation.
• DNA repair pathways significantly influence CNV generation at RDCs.