Role of recombination-dependent DNA damage response mechanisms in the resolution of benzo[a]pyrene-induced DNA replication stress
Final Report Abstract
Benzo[a]pyrene (B[a]P) is a well-characterised human carcinogen commonly found in the environment. Widespread exposure arises from cigarette smoke, air pollution, or barbequed foods and is therefore inevitable even for non-smokers. Upon intake, B[a]P forms bulky DNA adducts via its reactive metabolite (+)-anti-B[a]P-7,8-diol-9,10-epoxide (BPDE). These adducts mainly drive carcinogenicity of B[a]P as they are weak DNA repair substrates. Unrepaired BPDE adducts frequently persist into S-phase where they can impede replication fork progression, a process also known as DNA replication stress. Whilst resolution of replication stress by translesion synthesis is extensively studied at BPDE adducts, alternative recombination-dependent mechanisms are still poorly understood and were subject of this project. It is well established that replication stress frequently activates recombination via RAD51-dependent restart of stalled or collapsed forks. Surprisingly, DNA fibre analyses in mammalian cells revealed that BPDE-induced recombination is not associated with replication fork stalling. In contrast, RAD51 foci form under conditions when replication bypasses at bulky BPDE adducts, leading to the formation of single-stranded DNA (ssDNA) gaps. The recently discovered DNA primase PrimPol was identified as key mediator of ongoing replication and ssDNA gap formation on the BPDE-damaged template. Specifically, repriming by PrimPol is needed for initiation of recombination in response to BPDE, and RAD51 foci are lost when PrimPol is depleted or ectopically expressed as primase-dead variant. Recombination also depends on DNA end resection factors such as Mre11 and EXO1 in response to BPDE, suggesting that PrimPol-mediated ssDNA gaps require further extension for RAD51 loading. Taken together, this project uncovers a novel recombination pathway involving PrimPol to generate ssDNA gaps that govern RAD51-dependent resolution of B[a]P-induced replication stress. Similar involvement of PrimPol in UV-C-induced recombination highlights a broader importance of this new DNA damage response pathway for recombinogenic replication stress resolution.
Publications
- (2018) 'PARP1 and PARP2 stabilise replication forks at base excision repair intermediates through Fbh1-dependent Rad51 regulation', Nature Communications, 9: 746
Ronson GE, Piberger AL, Higgs MR, Olsen AL, Stewart GS, McHugh PJ, Petermann E, and Lakin ND
(See online at https://doi.org/10.1038/s41467-018-03159-2) - (2018) ‘BET inhibition induces HEXIM1- and RAD51-dependent conflicts between transcription and replication’, Cell Reports, 25: 2061-2069
Bowry A, Piberger AL, Rojas P, Saponaro M, and Petermann E
(See online at https://doi.org/10.1016/j.celrep.2018.10.079)
