Abstract FR:

The synthesis of the melanin pigments, the principal parameter of melanocytic differentiation, plays a key role in protection against deleterious effects of UV. Multiple clinical observations and experimental data show the key functionof the melanotropic hormones act on the melanocytes by inducing an increase in the intracellular cAMP concentration. In B16 mouse melanoma cells it was shown that cAMP leads to the inhibition of the PI3-K pathway and that this inhibition strongly stimulates the melanogenesis. But the molecular mechanisms involved in this process, however, remain to be elucidated. The focus of my thesis has been to try to understand how the inhibition of PI3-K controls melanocyte differentiation. Initially, we showed in the B16 melanoma cells that cAMP inhibits the activity of the protein kinase B PKB/AKT, the principal intracellular target of PI3-K, and leads to the downstream activation of GSK3b. Moreover, we observed that lithium, an inhibitor of GSK3b, inhibits the response to cAMP on tyrosinase promoter and that GSK3b synergises with MITF to stimulate the transcription of tyrosinase. Lastly, we demonstrated that cAMP increases the binding of MITF on its target sequence. In accordance with literature studies, these observations enabled us to propose that the activation of GSK3b by cAMP leads to MITF phosphorylation on serine 298. This phosphorylation stimulates the binding of MITF to its target sequence in the tyrosinase promoter, increases the transcription of the enzyme, and thus, stimulates the melanogenesis. In the second part of my studies, using a pharmacological inhibitor of the PI3-K, LY294002 we showed that the inhibition of the PI3-K induces an increase in the expression of two melanocytic enzymes tyrosinase and tyrp1 by a transcriptional mechanism which requires the transcription factor MITF. While LY294002 treatment induced an increase in MITF transcription, a dominant negative mutant of MITF did not allow any further stimulation in the expression of the melanocytic enzymes typically induced by the inhibition of the PI3-K pathway. In conclusion, we showed that the inhibition PI3-K pathway acts on two levels in the melanogenesis regulation. On one hand, it increases the MITF expression, and on the other hand, it increases binding on its target sequence to the tyrosinase promoter. Our work allows for better understanding of how the action of MITF in the PI3K pathway is implicated in the regulation of melanocyte differentiation.