132infertility. We use this mouse model to define stages of gametogenesis impaired by deficient BRCA2 expression and find evidence for a role in meiosis and gametogenic success in both males and females. Materials and methods Isolation and characterization of BACsHuman genomic BAC library filters obtained from Roswell Park Cancer Institute (now available at BACPAC Resource Center at the Children's Hospital Oakland Research Institute in Oakland, California) were screened using probes derived from PCR amplified DNA fragments from the 5′ (nucleotides 331-850) and 3′ (nucleotides 9852-10420) ends of the human BRCA2 cDNA (NM 000059). Thirteen clones were obtained from the human BAC library and seven were hybridized to both end probes. To identify the clone with the largest upstream and downstream regions, the T7 and SP6 ends of each BAC clone were sequenced and compared with the published sequence of human BRCA2 region. Human BRCA2 BAC, RP11-777I19 was selected to generate transgenic mice. The insert size (~165 kb) was confirmed by pulsed field gel electrophoresis of NotI-digested BAC DNA.

BRCA1, a well-known tumor suppressor with multiple interacting partners, is predicted to have diverse biological functions. However, so far its only well-established role is in the repair of damaged DNA and cell cycle regulation. In this regard, the etiopathological study of low-penetrant variants of BRCA1 provides an opportunity to uncover its other physiologically important functions. Using this rationale, we studied the R1699Q variant of BRCA1, a potentially moderate-risk variant, and found that it does not impair DNA damage repair but abrogates the repression of microRNA-155 (miR-155), a bona fide oncomir. Mechanistically, we found that BRCA1 epigenetically represses miR-155 expression via its association with HDAC2, which deacetylates histones H2A and H3 on the miR-155 promoter. We show that overexpression of miR-155 accelerates whereas the knockdown of miR-155 attenuates the growth of tumor cell lines in vivo. Our findings demonstrate a new mode of tumor suppression by BRCA1 and suggest that miR-155 is a potential therapeutic target for BRCA1-deficient tumors.

Poly (ADP-ribose) polymerase (PARP) inhibitor (PARPi) olaparib has been approved for treatment of advanced ovarian cancer associated with BRCA1 and BRCA2 mutations. BRCA1- and BRCA2-mutated cells, which are homologous recombination (HR) deficient, are hypersensitive to PARPi through the mechanism of synthetic lethality. Here we examine the effect of PARPi on HR-proficient cells. Olaparib pretreatment, PARP1 knockdown or Parp1 heterozygosity of Brca2cko/ko mouse embryonic stem cells (mESCs), carrying a null (ko) and a conditional (cko) allele of Brca2, results in viable Brca2ko/ko cells. PARP1 deficiency does not restore HR in Brca2ko/ko cells, but protects stalled replication forks from MRE11-mediated degradation through its impaired recruitment. The functional consequence of Parp1 heterozygosity on BRCA2 loss is demonstrated by a significant increase in tumorigenesis in Brca2cko/cko mice. Thus, while olaparib efficiently kills BRCA2-deficient cells, we demonstrate that it can also contribute to the synthetic viability if PARP is inhibited before BRCA2 loss.