Research
BRCA1 interaction network that suppresses tumor development
We have previously identified a BRCA1 C-terminal BRCT domain associated complex, termed the BRCA1-A complex. BRCA1 is recruited to DNA damage sites through the BRCA1-A complex in response to ionizing radiation (IR).
Our studies aim to delineate a functional role of this complex in the DNA damage response, BRCA1 signaling, and tumor suppression. Abraxas is the central adaptor protein in the BRCA1-A complex and binds to BRCA1 BRCT domains in a phosphorylation-dependent manner. We have generated Abraxas knockout mice and demonstrated that Abraxas plays an important role in suppressing tumor development in mice and that the interaction between Abraxas and BRCA1 is critical for Abraxas’ function in repair of DNA and maintenance of genome stability (Castillo et al., Cell Rep 8:807-817, 2014).
Our study show that Abraxas promotes BRCA1 dimerization in response to IR. We found that IR-induced phosphorylation of a serine residue next to the BRCA1-binding motif in Abraxas is critical for stabilizing the BRCT domains in forming a dimer. The phosphorylation is required for efficient accumulation of BRCA1 at DNA double strand breaks and cellular sensitivity to IR. Importantly, we identify that two previously uncharacterized BRCA1 germline mutations, F1662S and M1663K, occur at residues at the dimer interface and either of the two mutants disrupts BRCA1 BRCT dimer formation in vitro and in vivo (Wu et al., Mol Cell 61:434-448, 2016).
Project keywords:
Mouse Model/Tumor Suppression/Cancer Metastasis
Chromatin modification that regulates transcription inhibition and DNA repair at DNA double strand breaks
Post-translational modifications by covalent attachment of ubiquitin to proteins, known as ubiquitination, play important regulatory roles in the DNA damage response. Ubiquitin can be conjugated to the substrate by monoubiquitination or polyubiquitination with additional ubiquitin molecules conjugated through one of the seven lysine residues or through the N-terminal methionine residue, forming polyubiquitin chains of distinct linkages. Lys63-linkage ubiquitination on damaged chromatin has been shown to play important roles in recruiting DNA damage repair proteins to the damage sites, including 53BP1 and BRCA1. We have uncovered an ATM-dependent, RNF8-and Ube2S-catalyzed Lys11-linkage-specific ubiquitin modification on damaged chromatin that regulates repression of transcription at double strand breaks (DSB), revealing Lys11-linkage ubiquitin modification as a new signaling and regulatory platform in the response to DSBs (Paul and Wang, Mol Cell 66:458-472, 2017).
Our studies demonstrate that there is crosstalk between linkage-specific ubiquitin signaling at DNA damage sites. Cezanne and Cezanne2 are members of the ovarian tumor (OTU) subfamily deubiquitinating enzyme (DUB). Using a ubiquitin binding domain protein array screen, we identify that the UBA domains of Cezanne and Cezanne2 selectively bind to Lys63-linked polyubiquitin. Our work presents a model that Cezanne serves as a “reader” of the Lys63-linkage localizing to DNA damage sites and an “eraser” of the Lys11-linkage ubiquitination to promote the recruitment of Rap80/BRCA1-A complex and 53BP1 to DNA damage sites, indicating a crosstalk between linkage-specific ubiquitination at DNA damage sites. This work not only provides insights into the regulation of DNA repair and transcription at DSBs through linkage-specific ubiquitin modification, it also identifies Cezanne and Cezanne2 as two new players in the ubiquitin landscape at DSBs to facilitate recruitment of DNA repair proteins (Wu et al., Genes Dev 33:1702-1717, 2019).
Project keywords:
Chromatin Modification/Ubiquitin/DNA Damage Signaling/Genome Editing/Proteomic Analysis
Replication stress response that maintains genome stability and prevents cancer
Research from our laboratory has identified and analyzed Abro1, a paralog protein of Abraxas. We found that Abro1 forms an Abro1/BRISC complex in a similar manner as Abraxas forms the Abraxas/BRCA1-A complex (Hu et al., J. Biol Chem 286:11734-11745, 2011).
Recently, by generating and analyzing Abro1-/- mice and MEFs, we found that Abro1 is a new component in the protection of stalled replication fork integrity mechanisms. Abro1-deficiency results in increased chromosome instability, and Abro1-null mice appear to be tumor prone. Moreover, we show that Abro1 protects fork stability by inhibiting DNA2 nuclease-mediated degradation of nascent strand DNA at stalled forks, a mechanism that is distinct from the BRCA2-dependent pathway that inhibits MRE11-mediated degradation of stalled forks. It provides a new aspect of regulated fork protection mechanisms for maintenance of genome stability (Xu et al., Genes Dev 66:458-472, 2017).
Project keywords:
DNA Replication Stress Response/Chromosome Stability/Mouse Model