MULTIPLE INTRACELLULAR SIGNALLINGS ARE INVOLVED IN THYROTROPIN-RELEASING-HORMONE (TRH)-INDUCED C-FOS AND JUN-B MESSENGER-RNA LEVELS IN CLONAL PROLACTIN CELLS

In mammosomatotropes GH3B6 cells, one of the primary responses to thyr otropin-releasing hormone (TRH) is the parallel induction of two proto -oncogenes, c-fos and jun B, which code for constituents of AP1 transc ription factor. To better understand the mode of action of TRH and to look for possible functions of c-fos and jun B in these cells, we have investigated the role of different intracellular signals in the induc tion of each proto-oncogene on the one hand, and on prolactin (PRL) re lease and PRL gene expression on the other hand. Northern and dot-blot analyses revealed that the activation of protein kinase C (PKC)-, Ca2 +- or adenylyl cyclase-dependent pathways acutely increased both c-fos and jun B transcripts. However, a gene specific responsiveness was re vealed using phorbol 12-myristate 13-acetate (TPA) and several combine d treatments. The simultaneous activation of PKC and Ca2+-dependent pa thways resulted in synergistic stimulations of c-fos mRNA levels only. Consistently, ionomycin plus low doses of TPA solely reproduced the p otent effect of TRH on c-fos transcripts. Data collected from TRH and TPA down-regulated cells indicated that TRH probably recruits TPA-depe ndent PKC isoforms for stimulating c-fos but not jun B transcripts. On the contrary, the TRH-induced stimulation of either proto-oncogene li kely involves Ca2+-dependent mechanisms because calcium agonists and t he peptide exert non-additive effects. Finally, the synergistic stimul ations observed in response to TRH combined with forskolin, indicate t hat adenylyl cyclase-dependent mechanisms are interconnected with TRH- induced proto-oncogene expression. The overall study also reveals that among the agonists tested, the dihydropyridine Bay K 8644 and forskol in only were capable to induce a long-lasting stimulation of c-fos and jun B mRNA levels, concomitant to increased levels of PRL transcripts , as does TRH. Considering that AP1 is assumed to be involved in signa l transmission from the cell surface to the nucleus, it might be thus proposed that a common stimulation of c-fos and jun B gene expression is possibly involved in the activation of the PRL gene. On the other h and, the systematic coincidence between acute PRL release and proto-on cogenes expression suggest a role for c-fos and jun B in the control o f genes involved in the secretory process.