Single Molecule Analyses of homologous Recombination Tracts – SMART

DNA double-strand breaks (DSBs) occur accidentally or are self-inflicted, such as during meiosis, where they produce chiasmata, ensuring accurate homolog segregation. DSB are a threat to genome integrity and therefore must be faithfully repaired, in order to avoid aneuploidy, genetic diseases or cancer. Our project will shed new light on the fine mechanisms of one type of DSB repair, homologous recombination, by using innovative approaches combining experimental and computational analyses.
Many aspects of DSB repair by homologous recombination are poorly understood, due to the inability of population-based studies to decipher their fine regulation. However, improving our understanding of homologous recombination mechanisms is important, in particular the step of DNA synthesis, susceptible to be error-prone and introduce mutations. Here, we combine two experimental systems (in budding yeast and human cells) where DSB form at defined genomic positions to study by single-molecule long-read sequencing/imaging the DNA synthesis tracts associated to DSB repair.
Using these approaches, we will address how meiotic DSBs are patterned in individual cells, how a DSB repairs using the sister versus a homolog chromatid, and determine the complexity of recombination events, in meiotic and somatic cells. We will compare the length of DSB repair DNA synthesis tracts with resection tracts and with the position of chromosomal features, in meiotic and somatic cells, during the G1 and G2 phases of the cell cycle. Finally, we will determine whether DSB repair-associated DNA synthesis is mutagenic, which has important consequences for the generation of genetic diversity but also the deregulation of cellular functions.
The originality of our project resides in the combination of two complementary experimental systems with cutting edge high-throughput genomic technologies and the tight collaboration between two wet labs and one computational lab.