Dissecting the roles of Raf- and PI3K-signalling pathways in melanoma formation and progression in a zebrafish model.

Deregulated Ras signalling is implicated in most human neoplasia, exemplified by melanoma. Whereas Raf activation occurs almost ubiquitously in benign and malignant melanocytic neoplasms, implying an involvement in tumour initiation, phosphoinositide 3-kinase (PI3K) activation occurs predominantly in malignant neoplasms, implying an involvement in malignant progression. Here, we dissect the contributions of these two pathways to tumourigenesis in vivo, by modulating their activities in zebrafish melanocytes. Misexpression of oncogenic Ras (V12RAS) in founder fish induced frequent melanoma, beginning at larval stages, with concomitant activation of Raf-Mek-Erk and PI3K-Akt signalling. Misexpression of effector-domain mutants of V12RAS, or of various downstream effectors, confirmed a selective role for the Raf-Mek-Erk pathway in initiating neoplasia, but highlighted the requirement for additional Ras effector pathways for malignancy. The phenotype of animals with germ-line transmission of V12RAS resembled familial atypical mole and melanoma (FAMM) syndrome: melanocytes displayed hyperplasia, dysplasia, altered terminal differentiation and spontaneously progressed to invasive melanoma. Co-expressing a dominant-interfering form of PI3K abolished V12RAS-induced malignancy, demonstrating a direct role for PI3K signalling in the malignant progression of melanoma in vivo, and highlighting PI3K as a promising target for melanoma therapy.