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|Public on Oct 22, 2019
|53BP1 Enforces Distinct Pre- and Post-resection Blocks on Homologous Recombination (ChIP-seq)
|Genome binding/occupancy profiling by high throughput sequencing
|53BP1 activity drives genome instability and embryonic lethality in BRCA1-deficient cells by inhibiting homologous recombination (HR).53BP1’s anti-recombinogenic functions require phosphorylation-dependent interactions with two effector complexes, PTIP and RIF1/Shieldin. While RIF1/Shieldin is thought to block 5’-3’ nucleolytic processing of DNA ends, it remains unclear how PTIP antagonizes HR. Here we show that mutation of the PTIP interaction site in 53BP1 (S25A) increases Shieldin association with DNA damage. Despite excessive Shieldin “end-blocking” activity, the mutant protein allows end resection sufficient to rescue the lethality of BRCA1D11 mice. End resection in BRCA1D1153BP1S25A mice is rewired in a manner driven by DNA2 since Shieldin blocks EXO1-mediated nucleolytic processing. Despite ample resection, mutant cells fail to complete HR, as 53BP1/Shieldin also inhibits RNF168-mediated loading of PALB2/RAD51 onto ssDNA post-resection. As a result, BRCA1D1153BP1S25A mice exhibit hallmark features of HR insufficiency, including increased developmental neuronal apoptosis, premature aging and hypersensitivity to PARP inhibitors. Disruption of Shieldin or forced targeting of PALB2 to ssDNA in BRCA1D1153BP1S25A cells restores RNF168 recruitment, RAD51 nucleofilament formation, and PARPi resistance. Our study supports a model in which RIF1/Shieldin and PTIP associate independently with 53BP1 to regulate distinct end-resection pathways, and reveals a critical function of 53BP1/Shieldin post-resection that limits RNF168-mediated loading of RAD51.
|RPA ChIP-seq in WT, 53BP1-/-, BRCA1-/- MEFs at DSB sites upon induction of AsiSI. Rad51 ChIP-seq profile in WT, BRCA1-/- and BRCA1-/- 53BP1_S25A MEFs with or without FHA-PALB2 at AsiSI-induced DSB sites
|Callen E, Zong D, Wu W, Wong N, Stanlie A, Dumitrache LC, Byrum AK, Mendez-Dorantes C, Martinez P, Canela A, Maman Y, Day A, Kruhlak MJ, Blasco MA, Stark JM, Mosammaparast N, McKinnon PJ, Nussenzweig A
|Jul 03, 2019
|Last update date
|Jan 21, 2020
|National Cancer Institute
|Center for Cancer Research
|Laboratory of Genome Integrity
|9000 Rockville Pike
|This SubSeries is part of SuperSeries:
|53BP1 Enforces Distinct Pre- and Post-resection Blocks on Homologous Recombination