Abstract
DNA double-strand breaks are repaired by multiple mechanisms that are roughly grouped into the categories of homology-directed repair and non-homologous end joining. End-joining repair can be further classified as either classical non-homologous end joining, which requires DNA ligase 4, or “alternative” end joining, which does not. Alternative end joining has been associated with genomic deletions and translocations, but its molecular mechanism(s) are largely uncharacterized. Here, we report that Drosophila melanogaster DNA polymerase theta (pol theta), encoded by the mus308 gene and previously implicated in DNA interstrand crosslink repair, plays a crucial role in DNA ligase 4-independent alternative end joining. In the absence of pol theta, end joining is impaired and residual repair often creates large deletions flanking the break site. Analysis of break repair junctions from flies with mus308 separation-of-function alleles suggests that pol theta promotes the use of long microhomologies during alternative end joining and increases the likelihood of complex insertion events. Our results establish pol theta as a key protein in alternative end joining in Drosophila and suggest a potential mechanistic link between alternative end joining and interstrand crosslink repair.
Highlights
DNA double-strand breaks (DSBs) and interstrand crosslinks pose serious threats to cell survival and genome stability
Consistent with this, we have previously shown that a DSB caused by excision of a P element transposon in flies is readily repaired by a DNA ligase 4-independent end-joining process [17]
Based on the results presented above, it seemed likely that pol h is involved in an end-joining process different from C-non-homologous end joining (NHEJ)
Summary
DNA double-strand breaks (DSBs) and interstrand crosslinks pose serious threats to cell survival and genome stability. Because these lesions compromise both strands of the double helix, they impede DNA replication and transcription and must be removed in a timely and coordinated manner. Interstrand crosslink repair has been shown to involve a DSB intermediate in some cases (reviewed in [1]). Error-free repair of DSBs can be accomplished through homologous recombination (HR) with an undamaged homologous template (reviewed in [2]). Non-homologous end joining (NHEJ) frequently creates small insertions and deletions during DSB repair, in cases where the broken ends cannot be readily ligated (reviewed in [3]). The use of translesion DNA polymerases during interstrand crosslink repair can result in mutations, due to the reduced fidelity of these polymerases [4,5]
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