Abstract
BackgroundBRCA1 is a key regulatory protein participating in cell cycle checkpoint and DNA damage repair networks. BRCA1 plays important roles in protecting numerous cellular processes in response to cell damaging signals. Transforming growth factor-beta (TGF-β) is a potent regulator of growth, apoptosis and invasiveness of tumor cells. TFG-β activates Smad signaling via its two cell surface receptors, the TbetaRII and ALK5/TbetaRI, leading to Smad-mediated transcriptional regulation.Methodology/Principal FindingsHere, we report an important role of BRCA1 in modulating TGF-β signaling during oxidative stress responses. Wild-type (WT) BRCA1, but not mutated BRCA1 failed to activate TGF-β mediated transactivation of the TGF-β responsive reporter, p3TP-Lux. Further, WT-BRCA1, but not mutated BRCA1 increased the expression of Smad3 protein in a dose-dependent manner, while silencing of WT-BRCA1 by siRNA decreased Smad3 and Smad4 interaction induced by TGF-β in MCF-7 breast cancer cells. BRCA1 interacted with Smad3 upon TGF-β1 stimulation in MCF-7 cells and this interaction was mediated via the domain of 298–436aa of BRCA1 and Smad3 domain of 207–426aa. In addition, H2O2 increased the colocalization and the interaction of Smad3 with WT-BRCA1. Interestingly, TGF-β1 induced Smad3 and Smad4 interaction was increased in the presence of H2O2 in cells expressing WT-BRCA1, while the TGF-β1 induced interaction between Smad3 and Smad4 was decreased upon H2O2 treatment in a dose-dependent manner in HCC1937 breast cancer cells, deficient for endogenous BRCA1. This interaction between Smad3 and Smad4 was increased in reconstituted HCC1937 cells expressing WT-BRCA1 (HCC1937/BRCA1). Further, loss of BRCA1 resulted in H2O2 induced nuclear export of phosphor-Smad3 protein to the cytoplasm, resulting decreased of Smad3 and Smad4 interaction induced by TGF-β and in significant decrease in Smad3 and Smad4 transcriptional activities.Conclusions/SignificanceThese results strongly suggest that loss or reduction of BRCA1 alters TGF-β growth inhibiting activity via Smad3 during oxidative stress responses.
Highlights
Patients who inherit genetic defects in BRCA1 and BRCA2 have an increased lifetime risk of developing breast cancer
BRCA1 activates a Transforming growth factor-beta (TGF-b)-responsive reporter To understand whether inactivation of BRCA1 by germline mutations affects the TGF-b signaling pathway, we investigated the effect of BRCA1 mutations on TGF-b1 transactivation activity by performing a cell-based reporter assay with a TGF-bresponsive reporter, p3TP-Lux [19]
We uncovered an important role of BRCA1 in the TGF-b signaling pathway during oxidative stress responses
Summary
Patients who inherit genetic defects in BRCA1 and BRCA2 have an increased lifetime risk of developing breast cancer. BRCA1 is a multifunctional protein that has been implicated in many cellular processes, including genomic stability, the cell-cycle checkpoint, DNA-damage repair, apoptosis, and gene transcription [1]. BRCA1 has two important structural motifs, including a highly conserved amino-terminal RING finger motif and tandem BRCT motifs at its C-terminus [2,3]. The BRCT region of BRCA1 is essential for its DNA repair, transcriptional regulation and tumor suppressor functions [5]. Germline mutations in BRCA1 were often seen in the two regions [6], suggesting that the RING finger and BRCT motifs play an important role in the development of breast and ovarian cancers. BRCA1 is a key regulatory protein participating in cell cycle checkpoint and DNA damage repair networks. BRCA1 plays important roles in protecting numerous cellular processes in response to cell damaging signals. TFG-b activates Smad signaling via its two cell surface receptors, the TbetaRII and ALK5/TbetaRI, leading to Smad-mediated transcriptional regulation
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