TYRPHOSTIN INHIBITS BOMBESIN STIMULATION OF TYROSINE PHOSPHORYLATION, C-FOS EXPRESSION, AND DNA-SYNTHESIS IN SWISS 3T3 CELLS

Treatment of quiescent Swiss 3T3 cells with 20 muM {[(3,4,5-trihydroxyphenyl)methylene]propanedinitrile} (tyrphostin) caused a 76% r eduction in the tyrosine phosphorylation of the M(r) 110,000-130,000 band induced by bombesin. This was accompanied by a 48% reduction in the tyrosine phosphorylation of the cytosolic tyrosine kinase p125 focal adhesion kinase. Preincubation with 20 muM tyrphostin did not inhibit either protein kinase A activation by forskolin or protein kinase C (PKC) activation by phorbol 12,13-dibutyrate in intact Swiss 3T3 cells. Similarly, 20 muM tyrphostin neither interfered with binding of bombesin to its receptor nor prevented bombesin-stimulated Ca2+ mobilization or PKC activation. Thus tyrphostin selectively inhibits tyrosine phosphorylation induced by bombesin in intact Swiss 3T3 cells. Consequently, we examined the contribution of this tyrosine phosphorylation pathway to the subsequent induction of c-fos and stimulation of mitogenesis by bombesin. Tyrphostin prevented both c-fos mRNA expression and DNA synthesis induced by bombesin. The incorporation of [H-3] thymidine was inhibited by tyrphostin in a dose-dependent manner (IC50 = 20 muM), and this effect was not reversed even at high concentrations of bombesin. These results provide evidence that tyrosine phosphorylation is a mitogenic signal for bombesin.