DNA repair in motion: from molecular interactions to break-repair dynamics

Abstract

DNA double-strand breaks are among the most harmful forms of DNA damage and can lead to genomic instability and cancer when repaired incorrectly. Cells therefore employ multiple repair pathways whose activity is tightly regulated by factors including cell-cycle stage, chromatin context, and repair proteins such as BRCA1. This thesis investigates DNA repair from three complementary perspectives: repair dynamics, BRCA1 protein interactions, and the function of the BRCA1-A complex.

First, NANOBREAKR, a nanopore sequencing-based method, was developed to simultaneously distinguish broken, intact, and repaired DNA molecules, enabling direct analysis of repair kinetics, pathway usage, and structural repair outcomes. Second, systematic analysis of more than 4,000 variants in the BRCA1 BRCT domains revealed how individual mutations differently affect interactions with repair proteins and homologous recombination, providing insight into the functional consequences of disease-associated BRCA1 variants. Finally, studies of the BRCA1-A complex showed that it can regulate DNA end processing and resection independently of direct BRCA1 binding. Together, these findings provide new tools and mechanistic insights for understanding dynamic DNA repair processes, BRCA1 function, genome maintenance, and the interpretation of cancer-associated mutations.