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De novo mutations arising in the germline can drive genome evolution, but they can also affect embryonic development or cause genetic disorders when transmitted to offspring. Our goal is to understand mutagenic DNA repair mechanisms that operate in mouse and human germ cells, with a focus on meiosis and end-joining pathways. To this end, we use genetic and molecular assays combined with deep sequencing and computational analyses.
End joining at meiotic double-strand breaks (DSBs)
In meiosis, SPO11 forms numerous DSBs to ensure recombination between homologous chromosomes (Fig. A). However, while essential for correct chromosome segregation and healthy gamete production, DSBs are intrinsically mutagenic. In particular, as we showed in mice, closely spaced DSBs can undergo end joining, leading to de novo indels and structural variants (Fig. B). These events are enhanced in the absence of ATM, the kinase that regulates DSB formation.
End-joining pathways are well characterized in mammalian mitotic cells but their use in meiotic cells is largely unexplored. We are utilizing a combination of experimental approaches to elucidate the molecular mechanisms of end joining at SPO11 DSBs, uncover the spectrum of resulting genomic changes, and understand how meiotic cells normally suppress this mutagenic repair.
Topoisomerase II (TOP2)–driven genome instability in germ cells
TOP2 resolves topological stress associated with transcription, replication, and chromatin organization. This essential activity involves the transient formation of DSBs. While these DSBs are normally rapidly resealed, TOP2 can become trapped on DNA, resulting in persistent DSBs that need to be repaired. We are studying TOP2-induced DSBs in germ cells, their repair and mutagenic consequences using mouse mutants, biochemical and genomic approaches, and microscopy.
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