The alternative Epac /cAMP pathway and the MAPK pathway mediate hCG induction

JULIETA MAYMÓ; ANTONIO PÉREZ PÉREZ; BERNARDO MASKIN; JOSÉ DUEÑAS; JUAN CARLOS CALVO; VÍCTOR SÁNCHEZ-MARGALET; CECILIA L. VARONE

PLOS ONE

PUBLIC LIBRARY SCIENCE

Lugar: San Francisco; Año: 2012 vol. 7 p. 46216 - 46220

1932-6203

Leptin is the product of the obese gene, originally considered as an adipocyte-derived signaling molecule for the central control of metabolism. However, pleiotropic effects of leptin have been identified in reproduction and pregnancy, particularly in the placenta, where it may work as an autocrine hormone, mediating angiogenesis, growth and immunomodulation. In the present work, we demonstrated that recombinant human chorionic gonadotropin (hCG) added to JEG-3 choriocarcinoma cell line or to placental explants induces endogenous leptin expression, when analyzed by quantitative RT-PCR. On the other hand, we found that hCG increased cAMP intracellular levels in BeWo cells in a dose-dependent manner and stimulated cAMP response element (CRE) activity, evaluated by transient transfection. Moreover, cotransfection with an expression plasmid of a dominant negative mutant of CREB caused a significant inhibition of hCG stimulation of leptin promoter activity. These results demonstrate that hCG indeed activates cAMP/PKA pathway, and that this pathway is involved in leptin expression. Nevertheless, we found leptin induction by hCG is dependent on cAMP levels. Treatment with (Bu) cAMP in combination with low and non stimulatory hCG concentrations led to an increase in leptin expression, whereas stimulatory concentrations showed the opposite effect. The involvement of PKA on leptin induction by hCG was investigated. We found that specific PKA inhibition by H89 caused a significant increase of hCG leptin induction, suggesting that probably high cAMP levels might inhibit hCG effect. It was found that hCG enhancement of leptin mRNA expression involved the MAPK pathway. In this work, we demonstrated that hCG leptin induction through the MAPK signaling pathway is inhibited by PKA. In this sense, we observed that ERK1/2 phosphorylation increased when hCG treatment was combined with H89. In view of these results, the involvement of the alternative cAMP/Epac signaling pathway in hCG dependent leptin expression was studied. We observed that a cAMP analogue that specifically activates Epac (CPT-OMe) stimulated leptin placental expression by hCG. In addition, the overexpression of Epac and Rap1 proteins increased leptin promoter activity and enhanced hCG effect in transient transfection assays. In conclusion, we provide evidence suggesting that hCG induction of leptin gene expression in placental cells is mediated not only by activation of the MAPK signaling pathway but also by the alternative cAMP/Epac signaling pathway. Moreover, we have demonstrated that PKA activation interferes with this mechanism. These data provide novel insights into the signaling molecules and mechanisms regulating leptin gene expression in placenta by hCG.