RESUMO
Heptahelical opioid receptors are implicated in the transcriptional regulation of neuronal development. Here we demonstrated that activation of mu-opioid receptors in human neuroblastoma SH-SY5Y cells led to the activation of signal transducer and activator of transcription 3 (STAT3), a transcription factor central to the regulation of numerous biological processes. The mu-opioid-induced activation of STAT3 is sensitive to receptor was further shown to pertussis toxin treatment and required JAK and Src tyrosine kinases, but not phosphatidylinositol 3-kinase. This mu-opioid-induced response was mediated via the extracellular signal-regulated protein kinase in a Raf-1-independent manner. The present study provides a foundation to explore the importance of STAT3 signaling in the regulation of neuronal growth and differentiation by the mu-opioid receptor.
Assuntos
Proteínas de Ligação a DNA/metabolismo , MAP Quinase Quinase Quinase 1 , Sistema de Sinalização das MAP Quinases/fisiologia , Neuroblastoma , Receptores Opioides mu/metabolismo , Transativadores/metabolismo , Linhagem Celular Tumoral/efeitos dos fármacos , Linhagem Celular Tumoral/enzimologia , Subunidades alfa Gi-Go de Proteínas de Ligação ao GTP/metabolismo , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno , MAP Quinase Quinase Quinases/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Toxina Pertussis/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-raf/metabolismo , Fator de Transcrição STAT3 , Tirosina/metabolismo , Quinases da Família src/metabolismoRESUMO
The hematopoietic-specific G(q) subfamily members, Galpha(16) and Galpha(14) proteins have recently been shown to be capable of stimulating the signal transducer and activator of transcription 3 (STAT3) as well as STAT1. In the present study we examined whether this activation was STAT-member specific as well as determining the possible involvement of Gbetagamma dimers. Despite clear stimulation of STAT3, the constitutively active mutants of Galpha(16) (Galpha(16)QL) and Galpha(14) (Galpha(14)QL) failed to induce the phosphorylation of several STAT family members, including STAT2, STAT4 and STAT5 in human embryonic kidney 293 cells. On the other hand, transient expression of specific combinations of Gbetagamma complexes induced STAT3 phosphorylation. Among the 48 combinations tested, 13 permutations of Gbetagamma stimulated STAT3 phosphorylation and all of them contain the neuronal-specific Ggamma(2), Ggamma(4), Ggamma(7) and Ggamma(9). These results suggested that the activation of STAT family members by Galpha(16) or Galpha(14) was selective and that distinct combinations of Gbetagamma complexes can also regulate the STAT signaling pathway.
Assuntos
Subunidades alfa de Proteínas de Ligação ao GTP/metabolismo , Subunidades beta da Proteína de Ligação ao GTP/metabolismo , Subunidades gama da Proteína de Ligação ao GTP/metabolismo , Fator de Transcrição STAT3/metabolismo , Dimerização , Subunidades alfa de Proteínas de Ligação ao GTP/genética , Subunidades alfa de Proteínas de Ligação ao GTP/farmacologia , Subunidades beta da Proteína de Ligação ao GTP/genética , Subunidades beta da Proteína de Ligação ao GTP/farmacologia , Subunidades gama da Proteína de Ligação ao GTP/genética , Subunidades gama da Proteína de Ligação ao GTP/farmacologia , Células HeLa , Hematopoese , Humanos , Mutação/genética , Neurônios/metabolismo , Fosforilação , Ligação Proteica , Transdução de Sinais , Transgenes/genéticaRESUMO
The pseudomalignant nature of the placenta prompted us to search for tumor suppressor gene hypermethylation, a phenomenon widely reported in cancer, in the human placenta. Nine tumor suppressor genes were studied. Hypermethylation of the Ras association domain family 1 A (RASSF1A) gene was found in human placentas from all three trimesters of pregnancy but was absent in other fetal tissues. Hypermethylation of Rassf1 was similarly observed in placentas from the rhesus monkey but not the mouse. An inverse relationship between RASSF1A promoter methylation and gene expression was demonstrated by bisulfite sequencing of microdissected placental cells and immunohistochemical staining of placental tissue sections using an anti-RASSF1A antibody. Treatment of choriocarcinoma cell lines, JAR and JEG3, by 5-aza-2'-deoxycytidine and trichostatin A led to reduction in RASSF1A methylation but increased expression. These observations extend the analogy between the primate placenta and malignant tumors to the epigenetic level.