53BP1 and p53 synergize to suppress genomic instability and lymphomagenesis.
Ontology highlight
ABSTRACT:
SUBMITTER: Morales JC
PROVIDER: S-EPMC1413919 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Poly(ADP-ribose) polymerase 1 (PARP1) interacts genetically with the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) to suppress early-onset T-lineage lymphomas in the mouse, but the underlying mechanisms have remained unknown. To address this question, we analyzed a series of lymphomas arising in PARP1(-/-)/DNA-PKcs(-/-) (P1(-/-)/D(-/-)) mice. We found that, despite defective V(D)J recombination, P1(-/-)/D(-/-) lymphomas lacked clonal reciprocal translocations involving antigen-receptor loci. Instead, tumor cells were characterized by aneuploidy driven by two main mechanisms: p53 inactivation and abnormal chromosome disjunction due to telomere fusions (TFs). Aberrant accumulation of p53 was observed in 13/19 (68.4%) lymphomas. Sequence analysis revealed five p53 mutations: three missense point mutations (one transition in exon 8 and two transversions in exons 5 and 8, respectively), one in-frame 5-11 microindel in exon 7 and a 410-bp deletion encompassing exons 5-8, resulting in a truncated protein. Analysis of tumor metaphases using sequential telomere fluorescent in-situ hybridization and spectral karyotyping revealed that nine out of nine lymphomas contained TFs. Mutant but not wild-type p53 status was associated with frequent clonal and nonclonal TFs, suggesting that p53 normally limits the extent of telomere dysfunction during transformation. Chromosomes involved in TFs were more likely to be aneuploid than chromosomes not involved in TFs in the same metaphases, regardless of the p53 status, indicating that TFs promote aneuploidy via a mechanism that is distinct from p53 loss. Finally, analysis of radiation responses in P1(-/-)/D(-/-), and control primary cells and tissues indicates that loss of PARP1 increases in vivo radiosensitivity and genomic instability in DNA-PKcs-deficient mice without impairing p53 stabilization and effector functions, suggesting a more severe defect in double-strand break (DSB) repair in double mutants. Together, our findings uncover defective DSB repair leading to tumor suppressor inactivation and abnormal segregation of fused chromosomes as two novel mechanisms promoting tumorigenesis in thymocytes lacking PARP1 and DNA-PKcs.
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Project description:Misrepair of DNA double-strand breaks produced by the V(D)J recombinase (the RAG1/RAG2 proteins) at immunoglobulin (Ig) and T cell receptor (Tcr) loci has been implicated in pathogenesis of lymphoid malignancies in humans and in mice. Defects in DNA damage response factors such as ataxia telangiectasia mutated (ATM) protein and combined deficiencies in classical non-homologous end joining and p53 predispose to RAG-initiated genomic rearrangements and lymphomagenesis. Although we showed previously that RAG1/RAG2 shepherd the broken DNA ends to classical non-homologous end joining for proper repair, roles for the RAG proteins in preserving genomic stability remain poorly defined. Here we show that the RAG2 carboxy (C) terminus, although dispensable for recombination, is critical for maintaining genomic stability. Thymocytes from 'core' Rag2 homozygotes (Rag2(c/c) mice) show dramatic disruption of Tcr?/? locus integrity. Furthermore, all Rag2(c/c) p53(-/-) mice, unlike Rag1(c/c) p53(-/-) and p53(-/-) animals, rapidly develop thymic lymphomas bearing complex chromosomal translocations, amplifications and deletions involving the Tcr?/? and Igh loci. We also find these features in lymphomas from Atm(-/-) mice. We show that, like ATM-deficiency, core RAG2 severely destabilizes the RAG post-cleavage complex. These results reveal a novel genome guardian role for RAG2 and suggest that similar 'end release/end persistence' mechanisms underlie genomic instability and lymphomagenesis in Rag2(c/c) p53(-/-) and Atm(-/-) mice.
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Project description:BRCA1 is critical for DNA double-strand break (DSB) repair by homologous recombination (HR). BRCA1 deficient mice are embryonic lethal. Previous studies have shown that 53BP1 knockout (KO) rescues embryonic lethality of BRCA1 hypomorphic mutant mice by restoring HR. Here, we show that 53BP1 KO can partially rescue embryonic lethality of BRCA1 total KO mice, but HR is not restored in BRCA1-53BP1 double knockout (DKO) mice. As a result, BRCA1-53BP1 DKO cells are extremely sensitive to PARP inhibitors (PARPi). In addition to HR deficiency, BRCA1-53BP1 DKO cells have elevated microhomology-mediated end joining (MMEJ) activity and G2/M cell cycle checkpoint defects, causing severe genomic instability in these cells. Interestingly, BRCA1-53BP1 DKO mice rapidly develop thymic lymphoma that is 100% penetrant, which is not observed in any BRCA1 mutant mice rescued by 53BP1 KO. Taken together, our study reveals that 53BP1 KO can partially rescue embryonic lethality caused by complete BRCA1 loss without rescuing HR-related defects. This finding suggests that loss of 53BP1 can support the development of cancers with silenced BRCA1 expression without causing PARPi resistance.
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Project description:DNA constantly undergoes chemical modification due to endogenous and exogenous mutagens. The DNA base excision repair (BER) pathway is the frontline mechanism handling the majority of these lesions, and primarily involves a DNA incision and subsequent resealing step. It is imperative that these processes are extremely well-coordinated as unrepaired DNA single strand breaks (SSBs) can be converted to DNA double strand breaks during replication thus triggering genomic instability. However, the mechanism(s) governing the BER process are poorly understood. Here we show that accumulation of unrepaired SSBs triggers a p53/Sp1-dependent downregulation of APE1, the endonuclease responsible for the DNA incision during BER. Importantly, we demonstrate that impaired p53 function, a characteristic of many cancers, leads to a failure of the BER coordination mechanism, overexpression of APE1, accumulation of DNA strand breaks and results in genomic instability. Our data provide evidence for a previously unrecognized mechanism for coordination of BER by p53, and its dysfunction in p53-inactivated cells.
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Project description:The molecular pathways leading from genomic instability to cellular senescence and/or cell death remain incompletely characterized. Using mouse embryonic fibroblasts with constitutively increased DNA damage due to the absence of the full-length form of the tumor suppressor Brca1 (Brca1(Delta 11/Delta 11)), we show that deletion of p53 binding protein 1 (53BP1) selectivity abrogates senescence and cell death stimulated by reduced Brca1 activity. Furthermore, the embryonic lethality induced by Brca1 mutation can be alleviated by 53BP1 deletion. Adult Brca1(Delta 11/Delta 11)53BP1(-/-) manifest constitutively high levels of genomic instability, yet age relatively normally, with a surprisingly low incidence of overall tumor formation. Together, these in vitro and in vivo data suggest that 53BP1 is specifically required for the development of premature senescence and apoptosis induced by Brca1 deficiency. These observations may have important implications for Brca1-mediated tumor formation as well as for the molecular pathway leading from genomic instability to organismal aging.
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