Mutations and structural variants arising during double-strand break repair.

Area of Science:

• Molecular Biology
• Genetics
• DNA Repair Mechanisms

Background:

• Double-strand break (DSB) repair pathways are critical for maintaining genomic integrity.
• DSB repair can be mutagenic, leading to various genetic alterations.
• Understanding the mechanisms of DSB repair is essential for comprehending genome evolution and disease.

Purpose of the Study:

• To investigate the mutagenic outcomes of DSB repair in budding yeast.
• To characterize the types and frequencies of mutations arising during repair.
• To elucidate the role of microhomology and homeology in DSB repair-associated mutations.

Main Methods:

• Induction of DSBs at the MAT locus in budding yeast using HO endonuclease.
• Repair of DSBs using a transcriptionally silent HMR::Kl-URA3 donor sequence.
• Analysis of mutation types, including deletions, insertions, and template switches, using DNA sequencing.

Main Results:

• DSB repair favors -1 deletions over +1 insertions in homonucleotide runs, contrasting with replication.
• Microhomology-bounded intragenic deletions (IDs) are 12 times more frequent than tandem duplications (TDs).
• Interchromosomal template switches (ICTS) occur at regions of short microhomology and require extensive adjacent homeology alignment.

Conclusions:

• DSB repair involves distinct mechanisms leading to specific mutation signatures.
• Microhomology plays a crucial role in generating structural variants during repair.
• Extensive homeology alignment is critical for interchromosomal template switching during DSB repair.