RESUMO
Breast cancer 1 (BRCA1)-associated breast cancers are mostly basal-like high-grade ductal carcinomas that frequently overexpress epidermal growth factor receptor (EGFR). Aberrant EGFR expression is correlated with disease progression, resistance to radiation and chemotherapy, and poor clinical prognosis. Although BRCA1 is involved in multiple cellular processes, its functional role in EGFR regulation remains enigmatic. Here, we report a previously unrecognized posttranscriptional mechanism by which BRCA1 regulates EGFR expression through the induction of miR-146a. We demonstrate that EGFR expression correlates negatively with BRCA1, whereas miR-146a levels increase with BRCA1. We show that BRCA1 binds to MIR146A promoter and activates transcription, which in turn attenuates EGFR expression. Knockdown of miR-146a in BRCA1-overexpressing cells negated this effect and suppressed its ability to inhibit proliferation and transformation. In archived triple-negative breast cancer samples, we show a strong positive correlation between BRCA1 and miR-146a expression. We also show that low expression of miR-146a strongly predicts positive lymph node status and is associated with distinctively poor overall survival of patients. Together, these observations provide an insight into a novel BRCA1î²miR-146aî²EGFR paradigm by which BRCA1 carries out an aspect of tumor suppressor function that is potentially amenable to therapeutic intervention.
Assuntos
Proteína BRCA1/genética , Receptores ErbB/genética , MicroRNAs/genética , Neoplasias de Mama Triplo Negativas/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , Feminino , Humanos , Linfonodos/patologia , Regiões Promotoras Genéticas/genética , Processamento Pós-Transcricional do RNA/genética , Ativação Transcricional/genética , Neoplasias de Mama Triplo Negativas/patologia , Proteínas Supressoras de Tumor/genéticaRESUMO
Mammalian selenocysteine-containing proteins characterized with respect to function are involved in redox processes and exhibit distinct expression patterns and cellular locations. A recently identified 15-kDa selenoprotein (Sep15) has no homology to previously characterized proteins, and its function is not known. Here we report the intracellular localization and identification of a binding partner for this selenoprotein which implicate Sep15 in the regulation of protein folding. The native Sep15 isolated from rat prostate and mouse liver occurred in a complex with a 150-kDa protein. The latter protein was identified as UDP-glucose:glycoprotein glucosyltransferase (UGTR), the endoplasmic reticulum (ER)-resident protein, which was previously shown to be involved in the quality control of protein folding. UGTR functions by glucosylating misfolded proteins, retaining them in the ER until they are correctly folded or transferring them to degradation pathways. To determine the intracellular localization of Sep15, we expressed a green fluorescent protein-Sep15 fusion protein in CV-1 cells, and this protein was localized to the ER and possibly other perinuclear compartments. We determined that Sep15 contained the N-terminal signal peptide that was essential for translocation and that it was cleaved in the mature protein. However, C-terminal sequences of Sep15 were not involved in trafficking and retention of Sep15. The data suggest that the association between Sep15 and UGTR is responsible for maintaining the selenoprotein in the ER. This report provides the first example of the ER-resident selenoprotein and suggests a possible role of the trace element selenium in the quality control of protein folding.
Assuntos
Glucosiltransferases/metabolismo , Proteínas/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Primers do DNA , Glucosiltransferases/química , Masculino , Camundongos , Dados de Sequência Molecular , Próstata/metabolismo , Ligação Proteica , Ratos , Selenoproteínas , Frações Subcelulares/metabolismoRESUMO
Selenium has been implicated in cancer prevention, but the mechanism and possible involvement of selenoproteins in this process are not understood. To elucidate whether the 15-kDa selenoprotein may play a role in cancer etiology, the complete sequence of the human 15-kDa protein gene was determined, and various characteristics associated with expression of the protein were examined in normal and malignant cells and tissues. The 51-kilobase pair gene for the 15-kDa selenoprotein consisted of five exons and four introns and was localized on chromosome 1p31, a genetic locus commonly mutated or deleted in human cancers. Two stem-loop structures resembling selenocysteine insertion sequence elements were identified in the 3'-untranslated region of the gene, and only one of these was functional. Two alleles in the human 15-kDa protein gene were identified that differed by two single nucleotide polymorphic sites that occurred within the selenocysteine insertion sequence-like structures. These 3'-untranslated region polymorphisms resulted in changes in selenocysteine incorporation into protein and responded differently to selenium supplementation. Human and mouse 15-kDa selenoprotein genes manifested the highest level of expression in prostate, liver, kidney, testis, and brain, and the level of the selenoprotein was reduced substantially in a malignant prostate cell line and in hepatocarcinoma. The expression pattern of the 15-kDa protein in normal and malignant tissues, the occurrence of polymorphisms associated with protein expression, the role of selenium in differential regulation of polymorphisms, and the chromosomal location of the gene may be relevant to a role of this protein in cancer.
