Depolarization-induced tyrosine phosphorylation in PC12h cells.
Depolarization induced by KCl was found to induce tyrosine phosphorylation of cellular proteins in PC12h cells. By Western blotting with anti-phosphotyrosine antibody, we detected tyrosine phosphorylation of proteins with molecular weights of 120, 110, 105, 95, 75, 70, 66, 44, and 42 kDa in response to KCl. The immunoprecipitates from KCl-treated cells with the antibody contained large amounts of tyrosine-phosphorylated proteins and increased activity of tyrosine kinase. Incubation of the immunoprecipitates with [gamma-32P]ATP resulted in tyrosine phosphorylation of two proteins with the molecular weights of 120 and 140 kDa. These effects were completely abolished by the addition of EGTA before KCl treatment, suggesting that the depolarization-induced tyrosine phosphorylation may require calcium entry into the cells from the medium. Increased activity of tyrosine kinase phosphorylating the 120 and 140 kDa proteins was also recovered from cells stimulated with nerve growth factor, basic fibroblast growth factor, epidermal growth factor, and vasoactive intestinal peptide. Among them, depolarization by KCl elicited the strongest effect. These results indicate that a protein tyrosine kinase that phosphorylate the 120 and 140 kDa proteins is phosphorylated or activated in response to calcium ion, cAMP, and growth factors acting through tyrosine kinase receptors.
Okumura
N
N
Division of Metabolism, Osaka University.
Okada
M
M
Nakagawa
H
H
D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005346:Fibroblast Growth Factors / Q000494:pharmacology; D008564:Membrane Potentials / Q000502:physiology; D009414:Nerve Growth Factors / Q000494:pharmacology; D016716:PC12 Cells; D010766:Phosphorylation; D011189:Potassium Chloride / Q000494:pharmacology; D011233:Precipitin Tests; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D051381:Rats; D014443:Tyrosine / Q000378:metabolism*
D009414:Nerve Growth Factors; D014443:Tyrosine; D005346:Fibroblast Growth Factors; D004815:Epidermal Growth Factor; D011189:Potassium Chloride; D011505:Protein-Tyrosine Kinases; D002118:Calcium