Abstract
Subcellular localization regulates BRCA1 function, and BRCA1 is exported to the cytoplasm following DNA damage in a p53-dependent manner. Because more than 50% of solid tumors harbor p53 mutations, it is possible that genetically wild-type (wt) BRCA1 is functionally abnormal through compromised nuclear-cytoplasmic shuttling in sporadic breast cancer patients with dysfunctional p53. In this study, we have investigated the mechanisms of p53-dependent BRCA1 subcellular distribution and DNA damage-induced nuclear export, as well as the impact on the resulting cytotoxic response to therapy in human breast cancer. We first show that p53 mediates BRCA1 nuclear export via protein-protein binding, rather than by modulation of its transcription. Furthermore, it is the C-terminal (BRCT) region of BRCA1 that is critical for its interaction with p53, and p53 may promote BRCA1 nuclear export by interrupting the association of BRCA1 with BARD1. In sporadic breast cancer specimens, dysfunctional p53 strongly correlates with nuclear retention of sequence-verified wt BRCA1. This p53-dependent BRCA1 shuttling determines cellular susceptibility to DNA damage as augmentation of cytosolic BRCA1 significantly enhances cancer cell susceptibility to ionizing radiation. Taken together, our data suggest that p53 dysfunction compromises nuclear export of wt BRCA1 as a mechanism to increase cellular resistance to DNA damage in sporadic breast cancer. We propose that targeting nuclear BRCA1 to the cytoplasm may offer a unique strategy to sensitize p53-deficient sporadic breast cancers to DNA damage-based therapy.
Highlights
BRCA1 serves critical roles in multiple cellular processes, such as chromatin remodeling, DNA replication, DNA repair, transcriptional regulation, cell-cycle checkpoint, and apoptosis [1, 2]
The effect of p53 transcriptional activity on the subcellular distribution of BRCA1 was determined in HCC1937 human breast cancer cells that are functionally null in both p53 and BRCA1 [16, 17] following co-expression of HA-tagged wt-BRCA1 with wt-p53 or 2 tumor-associated p53 mutants (p53-179Q and p53-273H)
To confirm the results obtained by cell fractionation, immunohistochemistry was conducted on these cells to assess BRCA1 localization as a function of p53 by co-immunostaining of exogenously expressed BRCA1 and p53 using anti-HA and anti-p53 antibodies (Fig. 1B)
Summary
BRCA1 serves critical roles in multiple cellular processes, such as chromatin remodeling, DNA replication, DNA repair, transcriptional regulation, cell-cycle checkpoint, and apoptosis [1, 2]. These BRCA1 functions are controlled by multiple mechanisms, including phosphorylation, protein–protein interactions, and subcellular localization. BRCA1 is a nucleocytoplasmic shuttling protein [3]. Nuclear BRCA1 functions in DNA repair [4, 5] and cell-cycle checkpoints [1], whereas cytoplasmic BRCA1 regulates centrosome func-.
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