Microsatellite break-induced replication generates highly mutagenized extrachromosomal circular DNAs.

Area of Science:

• Genetics
• Molecular Biology
• Genomics

Background:

• Extrachromosomal circular DNAs (eccDNAs) are prevalent in eukaryotic genomes.
• The formation and mutagenic potential of eccDNAs, particularly those derived from non-B DNA structures, remain incompletely understood.

Purpose of the Study:

• To investigate the mechanisms by which non-B DNA structures generate mutagenized eccDNAs.
• To characterize the mutational patterns and genomic alterations associated with eccDNA formation.
• To explore the influence of DNA repair factors and replication stress on eccDNA mutagenesis.

Main Methods:

• Inverse PCR of non-B microsatellites integrated at an ectopic chromosomal site.
• Analysis of mutations including base substitutions, insertions, deletions, and template switching.
• Assessment of the impact of drug-induced replication stress and depletion of DNA repair factors (Rad51, COPS2, POLη).

Main Results:

• Non-B DNA microsatellites generate highly mutagenized eccDNAs via replication-dependent processes.
• Mutagenesis extends bidirectionally from the non-B DNA sequences into flanking genomic regions.
• Distinct non-B DNA structures yield unique mutagenesis patterns, and replication stress or DNA repair factor depletion alters these profiles.
• Proposed an asynchronous capture model involving break-induced replication to explain eccDNA generation and associated genomic homologous recombination deficiency (HRD) scars.

Conclusions:

• Non-B DNA structures are potent sources of mutagenized eccDNAs, contributing to genomic instability.
• The findings provide insights into the origins of tumor eccDNAs, their role in neoantigen production, oncogenesis, and chemotherapy resistance.