The BRCA2-MEILB2-BRME1 complex governs meiotic recombination and impairs the mitotic BRCA2-RAD51 function in cancer cells.
Ontology highlight
ABSTRACT:
SUBMITTER: Zhang J
PROVIDER: S-EPMC7188823 | biostudies-literature |
REPOSITORIES: biostudies-literature
Similar Datasets
Project description:The meiosis-specific synaptonemal complex protein SYCP3 has been reported to be aberrantly expressed in tumours. However, in contrast to its well-defined function in meiosis, its possible role in mitotic cells is entirely unknown. Here, we show that SYCP3 is expressed in a range of primary tumours and that it impairs chromosomal integrity in mitotic cells. Expression of SYCP3 inhibits the homologous recombination (HR) pathway mediated by RAD51, inducing hypersensitivity to DNA-damaging agents such as a poly(ADP-ribose) polymerase (PARP) inhibitor and chromosomal instability. SYCP3 forms a complex with BRCA2 and inhibits its role in HR. These findings highlight a new mechanism for chromosomal instability in cancer and extend the range of PARP-inhibitor sensitive tumours to those expressing SYCP3.
| S-EPMC3246250 | biostudies-literature
Project description:Heterozygous mutations in the tumor suppressor BRCA2 confer a high risk of breast and other cancers in humans. BRCA2 maintains genome stability in part through the regulation of Rad51-dependent homologous recombination. Much about its precise function in the DNA damage responses is, however, not yet known. We have made null mutations in the Drosophila homolog of BRCA2 and measured the levels of homologous recombination, non-homologous end-joining, and single-strand annealing in the pre-meiotic germline of Drosophila males. We show that repair by homologous recombination is dramatically decreased in Drosophila brca2 mutants. Instead, large flanking deletions are formed, and repair by the non-conservative single-strand annealing pathway predominates. We further show that during meiosis, Drosophila Brca2 has a dual role in the repair of meiotic double-stranded breaks and the efficient activation of the meiotic recombination checkpoint. The eggshell patterning defects that result from activation of the meiotic recombination checkpoint in other meiotic DNA repair mutants can be strongly suppressed by mutations in brca2. In addition, Brca2 co-immunoprecipitates with the checkpoint protein Rad9, suggesting a direct role for Brca2 in the transduction of the meiotic recombination checkpoint signal.
| S-EPMC2233675 | biostudies-literature
Project description:Mutation of the breast cancer susceptibility gene, BRCA2, leads to breast and ovarian cancers. Mechanistic insight into the functions of human BRCA2 has been limited by the difficulty of isolating this large protein (3,418 amino acids). Here we report the purification of full-length BRCA2 and show that it both binds RAD51 and potentiates recombinational DNA repair by promoting assembly of RAD51 onto single-stranded DNA (ssDNA). BRCA2 acts by targeting RAD51 to ssDNA over double-stranded DNA, enabling RAD51 to displace replication protein-A (RPA) from ssDNA and stabilizing RAD51-ssDNA filaments by blocking ATP hydrolysis. BRCA2 does not anneal ssDNA complexed with RPA, implying it does not directly function in repair processes that involve ssDNA annealing. Our findings show that BRCA2 is a key mediator of homologous recombination, and they provide a molecular basis for understanding how this DNA repair process is disrupted by BRCA2 mutations, which lead to chromosomal instability and cancer.
| S-EPMC2952063 | biostudies-literature
Project description:Meiotic recombination is required for proper homologous chromosome segregation in plants and other eukaryotes. The eukaryotic RAD51 gene family has seven ancient paralogs with important roles in mitotic and meiotic recombination. Mutations in mammalian RAD51 homologs RAD51C and XRCC3 lead to embryonic lethality. In the model plant Arabidopsis thaliana, RAD51C and XRCC3 homologs are not essential for vegetative development but are each required for somatic and meiotic recombination, but the mechanism of RAD51C and XRCC3 in meiotic recombination is unclear. The non-lethal Arabidopsis rad51c and xrcc3 null mutants provide an opportunity to study their meiotic functions. Here, we show that AtRAD51C and AtXRCC3 are components of the RAD51-dependent meiotic recombination pathway and required for normal AtRAD51 localization on meiotic chromosomes. In addition, AtRAD51C interacts with both AtRAD51 and AtXRCC3 in vitro and in vivo, suggesting that these proteins form a complex (es). Comparison of AtRAD51 foci in meiocytes from atrad51, atrad51c, and atxrcc3 single, double and triple heterozygous mutants further supports an interaction between AtRAD51C and AtXRCC3 that enhances AtRAD51 localization. Moreover, atrad51c-/+ atxrcc3-/+ double and atrad51-/+ atrad51c-/+ atxrcc3-/+ triple heterozygous mutants have defects in meiotic recombination, suggesting the role of the AtRAD51C-AtXRCC3 complex in meiotic recombination is in part AtRAD51-dependent. Together, our results support a model in which direct interactions between the RAD51C-XRCC3 complex and RAD51 facilitate RAD51 localization on meiotic chromosomes and RAD51-dependent meiotic recombination. Finally, we hypothesize that maintenance of RAD51 function facilitated by the RAD51C-XRCC3 complex could be highly conserved in eukaryotes.
| S-EPMC5470734 | biostudies-literature
Project description:Homologous recombination (HR) is one of the major DNA double-strand break (DSB) repair pathways in mammalian cells. Defects in HR trigger genomic instability and result in cancer predisposition. The defining step of HR is homologous strand exchange directed by the protein RAD51, which is recruited to DSBs by BRCA2. However, the regulation of the BRCA2-RAD51 axis remains unclear. Here we report that ubiquitination of RAD51 hinders RAD51-BRCA2 interaction, while deubiquitination of RAD51 facilitates RAD51-BRCA2 binding and RAD51 recruitment and thus is critical for proper HR. Mechanistically, in response to DNA damage, the deubiquitinase UCHL3 is phosphorylated and activated by ATM. UCHL3, in turn, deubiquitinates RAD51 and promotes the binding between RAD51 and BRCA2. Overexpression of UCHL3 renders breast cancer cells resistant to radiation and chemotherapy, while depletion of UCHL3 sensitizes cells to these treatments, suggesting a determinant role of UCHL3 in cancer therapy. Overall, we identify UCHL3 as a novel regulator of DNA repair and reveal a model in which a phosphorylation-deubiquitination cascade dynamically regulates the BRCA2-RAD51 pathway.
| S-EPMC5204351 | biostudies-literature
Project description:In most organisms, the primary function of homologous recombination (HR) is to allow genome protection by the faithful repair of DNA double-strand breaks. The vital step of HR is the search for sequence homology, mediated by the RAD51 recombinase, which is stimulated further by proteins mediators such as the tumor suppressor BRCA2. The biochemical interplay between RAD51 and BRCA2 is unknown in Leishmania or Trypanosoma. Here we show that the Leishmania infantum BRCA2 protein possesses several critical features important for the regulation of DNA recombination at the genetic and biochemical level. A BRCA2 null mutant, generated by gene disruption, displayed genomic instability and gene-targeting defects. Furthermore, cytological studies show that LiRAD51 can no longer localize to the nucleus in this mutant. The Leishmania RAD51 and BRCA2 interact together and the purified proteins bind single-strand DNA. Remarkably, LiBRCA2 is a recombination mediator that stimulates the invasion of a resected DNA double-strand break in an undamaged template by LiRAD51 to form a D-loop structure. Collectively, our data show that LiBRCA2 and LiRAD51 promote HR at the genetic and biochemical level in L. infantum, the causative agent of visceral leishmaniasis.
| S-EPMC3413117 | biostudies-literature
Project description:Homologous recombination promotes proper segregation of chromosomes during meiosis. Programmed double-strand breaks (DSBs) initiate recombination and are repaired preferentially using the homolog rather than the sister chromatid template. In yeast, activation of Mek1 kinase upholds this bias. Mek1 is also a proposed effector kinase in the recombination checkpoint that responds to aberrant DNA and/or axis structures. Elucidating a role for Mek1 in this checkpoint has been difficult, because a mek1 null mutation causes rapid repair of DSBs using a sister chromatid, thus bypassing formation of checkpoint-activating lesions. Here we analyzed a MEK1 gain-of-function allele to test if it would enhance interhomolog bias and/or the checkpoint response.When Mek1 activation was artificially maintained through glutathione S-transferase-mediated dimerization, there was an enhanced skew toward interhomolog recombination and reduction of intersister events, including multichromatid joint molecules. Increased interhomolog events were specifically repaired as noncrossovers rather than as crossovers. Ectopic Mek1 dimerization was also sufficient to impose interhomolog bias in the absence of recombination checkpoint functions, thereby uncoupling these two processes. Finally, the stringency of the checkpoint response was enhanced in mutants with weak recombination defects by blocking prophase exit in a subset of cells in which arrest is not absolute.We propose that Mek1 plays dual roles during meiotic prophase I by phosphorylating targets directly involved in the recombination checkpoint, as well as targets involved in sister chromatid recombination. We discuss how regulation of pachytene exit by Mek1 or similar kinases could influence checkpoint stringency, which may differ among species and between sexes.
| S-EPMC2989795 | biostudies-literature
Project description:BackgroundThe Rad51 recombinase assembles on DNA to execute homologous DNA recombination (HR). This process is essential to repair replication-associated genomic lesions before cells enter mitosis, but how it is started and stopped during the cell cycle remains poorly understood. Rad51 assembly is regulated by the breast cancer suppressor Brca2, via its evolutionarily conserved BRC repeats, and a distinct carboxy (C)-terminal motif whose biological function is uncertain. Using "hit-and-run" gene targeting to insert single-codon substitutions into the avian Brca2 locus, we report here a previously unrecognized role for the C-terminal motif.ResultsWe show that the avian C-terminal motif is functionally cognate with its human counterpart and identify point mutations that either abolish or enhance Rad51 binding. When these mutations are introduced into Brca2, we find that they affect neither the assembly of Rad51 into nuclear foci on damaged DNA nor DNA repair by HR. Instead, foci disassemble more rapidly in a point mutant that fails to bind Rad51, associated with faster mitotic entry. Conversely, the slower disassembly of foci in a point mutant that constitutively binds Rad51 correlates with delayed mitosis. Indeed, Rad51 foci do not persist in mitotic cells even after G2 checkpoint suppression, suggesting that their disassembly is a prerequisite for chromosome segregation.ConclusionsWe conclude that Rad51 binding by the C-terminal Brca2 motif is dispensable for the execution of HR but instead links the disassembly of Rad51 complexes to mitotic entry. This mechanism may ensure that HR terminates before chromosome segregation. Our findings assign a biological function for the C-terminal Brca2 motif in a mechanism that coordinates DNA repair with the cell cycle.
| S-EPMC2719694 | biostudies-literature
Project description:RAD51 recombinase polymerizes at the site of double-strand breaks (DSBs) where it performs DSB repair. The loss of RAD51 causes extensive chromosomal breaks, leading to apoptosis. The polymerization of RAD51 is regulated by a number of RAD51 mediators, such as BRCA1, BRCA2, RAD52, SFR1, SWS1, and the five RAD51 paralogs, including XRCC3. We here show that brca2-null mutant cells were able to proliferate, indicating that RAD51 can perform DSB repair in the absence of BRCA2. We disrupted the BRCA1, RAD52, SFR1, SWS1, and XRCC3 genes in the brca2-null cells. All the resulting double-mutant cells displayed a phenotype that was very similar to that of the brca2-null cells. We suggest that BRCA2 might thus serve as a platform to recruit various RAD51 mediators at the appropriate position at the DNA-damage site.
| S-EPMC3136442 | biostudies-literature
Project description:Mutations in the tumor suppressor BRCA2 predominantly predispose to breast cancer. Paradoxically, while loss of BRCA2 promotes tumor formation, it also causes cell lethality, although how lethality is triggered is unclear. Here, we generate BRCA2 conditional non-transformed human mammary epithelial cell lines using CRISPR-Cas9. Cells are inviable upon BRCA2 loss, which leads to replication stress associated with under replication, causing mitotic abnormalities, 53BP1 nuclear body formation in the ensuing G1 phase, and G1 arrest. Unexpected from other systems, the role of BRCA2 in homologous recombination, but not in stalled replication fork protection, is primarily associated with supporting human mammary epithelial cell viability, and, moreover, preventing replication stress, a hallmark of pre-cancerous lesions. Thus, we uncover a DNA under replication-53BP1 nuclear body formation-G1 arrest axis as an unanticipated outcome of homologous recombination deficiency, which triggers cell lethality and, we propose, serves as a barrier that must be overcome for tumor formation.BRCA2 mutations promote tumour formation while also paradoxically causing cell lethality. Here the authors generate conditional BRCA2 loss in a non-transformed human mammary cell line and see increased replication stress due to under-replication of DNA.
| S-EPMC5597640 | biostudies-literature