RESUMO
Human Cripto-1 (CR-1) plays an important role in regulating embryonic development while also regulating various stages of tumor progression. However, mechanisms that regulate CR-1 expression during embryogenesis and tumorigenesis are still not well defined. In the present study, we investigated the effects of two nuclear receptors, liver receptor homolog (LRH)-1 and germ cell nuclear factor receptor (GCNF) and epigenetic modifications on CR-1 gene expression in NTERA-2 human embryonal carcinoma cells and in breast cancer cells. CR-1 expression in NTERA-2 cells was positively regulated by LRH-1 through direct binding to a DR0 element within the CR-1 promoter, while GCNF strongly suppressed CR-1 expression in these cells. In addition, the CR-1 promoter was unmethylated in NTERA-2 cells, while T47D, ZR75-1, and MCF7 breast cancer cells showed high levels of CR-1 promoter methylation and low CR-1 mRNA and protein expression. Treatment of breast cancer cells with a demethylating agent and histone deacetylase inhibitors reduced methylation of the CR-1 promoter and reactivated CR-1 mRNA and protein expression in these cells, promoting migration and invasion of breast cancer cells. Analysis of a breast cancer tissue array revealed that CR-1 was highly expressed in the majority of human breast tumors, suggesting that CR-1 expression in breast cancer cell lines might not be representative of in vivo expression. Collectively, these findings offer some insight into the transcriptional regulation of CR-1 gene expression and its critical role in the pathogenesis of human cancer.
Assuntos
Neoplasias da Mama/metabolismo , Carcinoma Ductal de Mama/metabolismo , Carcinoma Embrionário/metabolismo , Metilação de DNA , Proteínas Ligadas por GPI/metabolismo , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Proteínas de Neoplasias/metabolismo , Membro 1 do Grupo A da Subfamília 6 de Receptores Nucleares/metabolismo , Regiões Promotoras Genéticas , Receptores Citoplasmáticos e Nucleares/metabolismo , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Sítios de Ligação , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Carcinoma Ductal de Mama/genética , Carcinoma Ductal de Mama/patologia , Carcinoma Embrionário/genética , Carcinoma Embrionário/patologia , Movimento Celular , Metilação de DNA/efeitos dos fármacos , Metilases de Modificação do DNA/antagonistas & inibidores , Metilases de Modificação do DNA/metabolismo , Decitabina , Relação Dose-Resposta a Droga , Células-Tronco de Carcinoma Embrionário/metabolismo , Células-Tronco de Carcinoma Embrionário/patologia , Feminino , Proteínas Ligadas por GPI/genética , Regulação da Expressão Gênica no Desenvolvimento , Regulação Neoplásica da Expressão Gênica , Genes Reporter , Inibidores de Histona Desacetilases/farmacologia , Humanos , Ácidos Hidroxâmicos/farmacologia , Peptídeos e Proteínas de Sinalização Intercelular/genética , Luciferases/biossíntese , Luciferases/genética , Células MCF-7 , Invasividade Neoplásica , Proteínas de Neoplasias/genética , Membro 1 do Grupo A da Subfamília 6 de Receptores Nucleares/genética , Interferência de RNA , RNA Mensageiro/metabolismo , Receptores Citoplasmáticos e Nucleares/genética , Fatores de Tempo , Análise Serial de Tecidos , Transcrição Gênica , Transfecção , Tretinoína/farmacologia , Ácido Valproico/farmacologiaRESUMO
Cripto-1 is a membrane-bound protein that is highly expressed in embryonic stem cells and in human tumors. In the present study, we investigated the effect of low levels of oxygen, which occurs naturally in rapidly growing tissues, on Cripto-1 expression in mouse embryonic stem (mES) cells and in human embryonal carcinoma cells. During hypoxia, Cripto-1 expression levels were significantly elevated in mES cells and in Ntera-2 or NCCIT human embryonal carcinoma cells, as compared with cells growing with normal oxygen levels. The transcription factor hypoxia-inducible factor-1alpha directly regulated Cripto-1 expression by binding to hypoxia-responsive elements within the promoter of mouse and human Cripto-1 genes in mES and NCCIT cells, respectively. Furthermore, hypoxia modulated differentiation of mES cells by enhancing formation of beating cardiomyocytes as compared with mES cells that were differentiated under normoxia. However, hypoxia failed to induce differentiation of mES cells into cardiomyocytes in the absence of Cripto-1 expression, demonstrating that Cripto-1 is required for hypoxia to fully differentiate mES cells into cardiomyocytes. Finally, cardiac tissue samples derived from patients who had suffered ischemic heart disease showed a dramatic increase in Cripto-1 expression as compared with nonischemic heart tissue samples, suggesting that hypoxia may also regulate Cripto-1 in vivo.