TYROSINE PHOSPHORYLATION OF PAXILLIN AND FOCAL ADHESION KINASE DURINGINSULIN-LIKE GROWTH FACTOR-I-STIMULATED LAMELLIPODIAL ADVANCE

In the current studies, we examined whether focal adhesion kinase (FAK ) and paxillin play a role in insulin-like growth factor-I (IGF-I)-sti mulated morphological changes in neuronal cells, In SH-SY5Y human neur oblastoma cells, 10 nM IGF-I enhanced the extension of lamellipodia wi thin 30 min, Scanning electron microscopy and staining with rhodamine- phalloidin showed that these lamellipodia displayed ruffles, filopodia , and a distinct meshwork of actin filaments, Immunofluorescent staini ng identified focal concentrations of FAK, paxillin, and phosphotyrosi ne within the lamellipodia, Immunoprecipitation experiments revealed t hat FAK and paxillin are tyrosine phosphorylated during IGF-I-stimulat ed lamellipodial extension, Maximal phosphorylation of FAK and paxilli n was observed 15-30 min after the addition of 10 nM IGF-I, whereas ma ximal IGF-I receptor phosphorylation occurred within 5 min, FAK, paxil lin, and IGF-I receptor tyrosine phosphorylation had similar concentra tion-response curves and were inhibited by the receptor blocking antib ody alpha IR-3. These results indicate that FAK and paxillin are tyros ine-phosphorylated during IGF-I-stimulated lamellipodial advance and s uggest that the tyrosine phosphorylation of these two proteins helps m ediate IGF-I stimulated cell and growth cone motility, These responses contrast directly with recent reports showing insulin-stimulated deph osphorylation of FAK and paxillin.