FAN1-mediated translesion synthesis and POLQ/HELQ-mediated end joining generate interstrand crosslink-induced mutations

Abstract

Abstract DNA interstrand cross-links (ICLs) prevent strand separation during DNA replication and transcription and therefore are extremely cytotoxic. To counteract these effects cells evolved various specialized ICL repair pathways. The division of labor between these routes, and the extent to which repair impact on genetic integrity is, however, still poorly understood. Here, we have determined the mutagenic consequences of ICL repair in the animal model C. elegans. Plasmids containing a single, site-specific, psoralen crosslink were injected into the worm’s gonad, after which targeted sequencing of transgenic progeny animals was conducted to establish repair profiles. Through genetic dissection we identified two mechanisms by which psoralen ICLs induce genetic change: i) translesion synthesis through POLH and REV1/3-mediated bypass, giving rise to single nucleotide polymorphisms (SNVs), and ii) end joining through POLQ or HELQ action leading to deletions. Whilst we found no role for the Fanconi anemia genes FANCD2 and FANCI in causing nor suppressing these deletions, disruption of C. elegans TRAIP, which triggers the unloading of the CMG helicase at sites of blocked replication, resulted in a strikingly altered repair profile, arguing for a role for DNA replication in the aetiology of ICL-induced deletions. Translesion synthesis products were not affected by TRAIP deficiency. Instead, we found SNVs induced at ICLs to depend on the functionality of the Fanconi anemia-associated nuclease FAN1.