Activation of a microtubule-associated protein-2 kinase by insulin-like growth factor-I in bovine chromaffin cells.
Treatment of bovine chromaffin cells with insulin-like growth factor-I (IGF-I) caused the activation of a protein kinase that phosphorylates microtubule-associated protein-2 (MAP-2) in vitro. Activation of MAP-2 kinase by IGF-I varied with the time of treatment (maximal at 10-15 min) and the concentration of IGF-I (maximal at 10 nM). The IGF-I-activated MAP-2 kinase was localized to the soluble fraction of chromaffin cell extracts and required Mg2+ for activity. The IGF-I-activated kinase also phosphorylated myelin basic protein, but had little or no activity toward histones or ribosomal S6 protein. To examine the role of protein tyrosine phosphorylation in the activation of the MAP-2 kinase, we isolated phosphotyrosine (PTyr)-containing proteins from chromaffin cells by immunoaffinity adsorption on anti-PTyr-Sepharose beads. Anti-PTyr-Sepharose eluates from IGF-I-treated cells showed increased MAP-2 kinase activity; thus, the MAP-2 kinase (or a closely associated protein) appears to be a PTyr-containing protein. Treatment of anti-PTyr-Sepharose eluates or crude chromaffin cell extracts with alkaline phosphatase significantly decreased kinase activity toward myelin basic protein, indicating that phosphorylation of the IGF-I-activated kinase is required for its activity.
Cahill
A L
AL
Department of Pediatrics, University of Chicago, Illinois 60637.
Perlman
R L
RL
D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D001345:Autoradiography; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002412:Cations / Q000378:metabolism; D002417:Cattle; D002838:Chromaffin System / Q000166:cytology / Q000201:enzymology*; D004789:Enzyme Activation; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology*; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D013347:Subcellular Fractions / Q000201:enzymology; D014443:Tyrosine / Q000378:metabolism
D002412:Cations; D014443:Tyrosine; D008345:Manganese; D007334:Insulin-Like Growth Factor I; D011494:Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D000469:Alkaline Phosphatase; D008274:Magnesium