ANGIOTENSIN-II-INDUCED PROTEIN-TYROSINE PHOSPHORYLATION IN NEONATAL RAT CARDIAC FIBROBLASTS

Angiotensin II has been demonstrated to act as a growth factor in rat cardiac fibroblasts. However, the signaling events that lead to fibrob last cell growth in response to angiotensin II remain to be elucidated . This study was designed to determine whether angiotensin II stimulat ed tyrosine phosphorylation of proteins in cardiac fibroblasts. Immuno blot analysis demonstrated rapid tyrosine phosphorylation of distinct substrates of 125, 95, 46-60, and 44 kDa in response to 10 nM angioten sin II. Tyrosine phosphorylation was maximal at 5 min and persisted fo r at least 180 min. Additional tyrosine-phosphorylated proteins of 185 , 145, and 85 kDa were detected in response to 10 ng/ml platelet-deriv ed growth factor BB. A cluster of 75-80-kDa proteins were phosphorylat ed in response to angiotensin II, phorbol ester, and platelet-derived growth factor. Angiotensin II-induced tyrosine phosphorylation was una ffected by phorbol ester-sensitive protein kinase C down-regulation an d could be partially blocked by pertussis toxin pretreatment. Angioten sin II stimulation resulted in increased cytosolic tyrosine kinase act ivity which was recovered by immunoprecipitation. Immunoblot analysis demonstrated tyrosine phosphorylation of p44(MAPK), and, in addition, we demonstrated for the first time tyrosine phosphorylation of p125(FA K), p46(SHC), and p56(SHC) in response to angiotensin II. The finding that angiotensin II and platelet-derived growth factor stimulated tyro sine phosphorylation of p46(SHC) and p56(SHC) suggested that this prot ein may serve as a common tyrosine kinase substrate in the mitogenic s ignaling cascade induced by G-protein-coupled receptors and growth fac tors and is consistent with the hypothesis that angiotensin II-induced tyrosine phosphorylation is involved in mitogenic signaling pathways in neonatal rat cardiac fibroblasts.