Homologous recombination, i.e., homology-directed repair (HDR), is a major repair pathway for double-strand breaks (DSBs) including lesions arising during DNA replication. HDR mutants are characterized by genomic instability and sensitivity to DNA damaging agents such as inter-strand crosslinkers and poly (ADP-ribose) polymerase inhibitors. Several proteins central to the HDR pathway are tumor suppressors, notably the breast cancer suppressor BRCA2, which promotes the function of RAD51, the critical protein for homologous strand exchange. RAD51 paralogs are also tumor suppressors. The premise of this project is based on the established link of HDR with tumor suppression and therapy response. Thus, this proposal seeks to define roles of HDR factors and pathways, and the cellular responses to HDR deficiencies.
The specific aims are: 1. To determine the cellular responses to RAD51 paralog deficiency. We constructed isogenic human mammary epithelial cell lines with conditional mutations in genes for the five canonical RAD51 paralogs. We plan to compare these mutants for repair of spontaneous and induced DNA damage and replication fork protection. We hypothesize that their deficiency will lead to replication stress due to HDR deficiency. 2. To determine the cellular responses to BRCA2 deficiency, including the innate immune response and factors that affect the response. BRCA2 loss leads to severe replication stress and subsequent mitotic abnormalities. We propose to determine the effect of BRCA2 deficiency on the innate immune response that senses cytosolic DNA involving cGAS-STING. Factors that may intensify or abrogate the response to replication stress will be investigated. 3. To investigate the requirement for BRCA2 DNA binding. The most conserved region of BRCA2 is the DSS1and DNA-Binding Domain. We will investigate the role of this domain in cells and mice using a number of genetic and biochemical approaches. 4. Requirement for DSS1/SEM1 for survival and DNA repair in the context of BRCA2. The interaction with DSS1 is critical for BRCA2 function in the context of the full-length protein. We will address whether DSS1 is essential solely due to its interaction with BRCA2 or whether the requirement for DSS1 is through a BRCA2- independent role.

Public Health Relevance

Lesions that arise in the genome compromise its integrity and so must be repaired. Loss of repair or misrepair leads to chromosome loss and genomic rearrangements, which are associated with many tumor types, in particular breast and ovarian cancer. This project addresses fundamental questions about the tumor suppressors BRCA2 and RAD51 paralogs involved in repair between similar (homologous) sequences.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA185660-07
Application #
9983604
Study Section
Cancer Etiology Study Section (CE)
Program Officer
Oberdoerffer, Philipp
Project Start
2014-05-01
Project End
2024-04-30
Budget Start
2020-05-01
Budget End
2021-04-30
Support Year
7
Fiscal Year
2020
Total Cost
Indirect Cost
Name
Sloan-Kettering Institute for Cancer Research
Department
Type
DUNS #
064931884
City
New York
State
NY
Country
United States
Zip Code
10065
Brunet, Erika; Jasin, Maria (2018) Induction of Chromosomal Translocations with CRISPR-Cas9 and Other Nucleases: Understanding the Repair Mechanisms That Give Rise to Translocations. Adv Exp Med Biol 1044:15-25
Chen, Chun-Chin; Feng, Weiran; Lim, Pei Xin et al. (2018) Homology-Directed Repair and the Role of BRCA1, BRCA2, and Related Proteins in Genome Integrity and Cancer. Annu Rev Cancer Biol 2:313-336
Abreu, Carla M; Prakash, Rohit; Romanienko, Peter J et al. (2018) Shu complex SWS1-SWSAP1 promotes early steps in mouse meiotic recombination. Nat Commun 9:3961
Jahid, Sohail; Sun, Jian; Gelincik, Ozkan et al. (2017) Inhibition of colorectal cancer genomic copy number alterations and chromosomal fragile site tumor suppressor FHIT and WWOX deletions by DNA mismatch repair. Oncotarget 8:71574-71586
Chen, Chun-Chin; Kass, Elizabeth M; Yen, Wei-Feng et al. (2017) ATM loss leads to synthetic lethality in BRCA1 BRCT mutant mice associated with exacerbated defects in homology-directed repair. Proc Natl Acad Sci U S A 114:7665-7670
Vanoli, Fabio; Jasin, Maria (2017) Generation of chromosomal translocations that lead to conditional fusion protein expression using CRISPR-Cas9 and homology-directed repair. Methods 121-122:138-145
Vanoli, Fabio; Tomishima, Mark; Feng, Weiran et al. (2017) CRISPR-Cas9-guided oncogenic chromosomal translocations with conditional fusion protein expression in human mesenchymal cells. Proc Natl Acad Sci U S A 114:3696-3701
Feng, Weiran; Jasin, Maria (2017) BRCA2 suppresses replication stress-induced mitotic and G1 abnormalities through homologous recombination. Nat Commun 8:525
Feng, Weiran; Jasin, Maria (2017) Homologous Recombination and Replication Fork Protection: BRCA2 and More! Cold Spring Harb Symp Quant Biol 82:329-338
Kondrashova, Olga; Nguyen, Minh; Shield-Artin, Kristy et al. (2017) Secondary Somatic Mutations Restoring RAD51C and RAD51D Associated with Acquired Resistance to the PARP Inhibitor Rucaparib in High-Grade Ovarian Carcinoma. Cancer Discov 7:984-998

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