Specific amino acid restriction inhibits attachment and spreading of human melanoma via modulation of the integrin/focal adhesion kinase pathway and actin cytoskeleton remodeling
We found previously that restriction of tyrosine (Tyr) and phenylalanine (Phe) inhibited growth and metastasis of B16BL6 murine melanoma and arrested these cells in the G0-G1 phase of the cell cycle. Here, we report that deprivation of these two amino acids in vitro induces apoptosis in B16BL6 and in human A375 melanoma cells but not in nontransformed, neonatal murine epidermal cells or human infant foreskin fibroblasts. Four days after deprivation of Tyr and Phe in vitro, 37% of B16BL6 and 51% of A375 melanoma cells were undergoing apoptosis. Apoptosis was not associated with elevation in intracellular calcium or alteration in p53 or c-myc protein expression. Expression and Tyr phosphorylation of focal adhesion kinase (FAK) were inhibited in both melanoma cell lines by deprivation of Tyr and Phe but not by deprivation of glutamine or serum. Tyr phosphorylation of FAK in Tyr- and Phe-deprived melanoma cells was enhanced within 30 min of refeeding with complete DMEM. FAK protein expression recovered within 60 min, and cell viability recovered within 24 h. Genistein, a tyrosine kinase inhibitor that specifically inhibits Tyr phosphorylation of FAK, did not induce apoptosis in A375 melanoma cells at a concentration of 50 microM. Genistein prevented the recovery of cell viability upon refeeding with Tyr and Phe to previously deprived A375 melanoma cells. These data collectively indicate that apoptosis induced by Tyr and Phe deprivation is FAK-dependent.