Glutamate-induced protein phosphorylation in cerebellar granule cells: role of protein kinase C.
Protein phosphorylation in response to toxic doses of glutamate has been investigated in cerebellar granule cells. 32P-labelled cells have been stimulated with 100 microM glutamate for up to 20 min and analysed by one and two dimensional gel electrophoresis. A progressive incorporation of label is observed in two molecular species of about 80 and 43 kDa (PP80 and PP43) and acidic isoelectric point. Glutamate-stimulated phosphorylation is greatly reduced by antagonists of NMDA and non-NMDA glutamate receptors. The effect of glutamate is mimicked by phorbol esters and is markedly reduced by inhibitors of protein kinase C (PKC) such as staurosporine and calphostin C. PP80 has been identified by Western blot analysis as the PKC substrate MARCKS (myristoylated alanine-rich C kinase substrate), while antibody to GAP-43 (growth associated protein-43), the nervous tissue-specific substrate of PKC, failed to recognize PP43. Our results suggest that PKC is responsible for the early phosphorylative events induced by toxic doses of glutamate in cerebellar granule cells.
Eboli
M L
ML
Institute of General Pathology, Catholic University, Rome, Italy.
Mercanti
D
D
Ciotti
M T
MT
Aquino
A
A
Castellani
L
L
D000818:Animals; D002531:Cerebellum / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D019922:GAP-43 Protein; D018698:Glutamic Acid / Q000494:pharmacology*; D008562:Membrane Glycoproteins / Q000378:metabolism; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009474:Neurons / Q000187:drug effects* / Q000378:metabolism; D010703:Phorbol Esters / Q000494:pharmacology; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000502:physiology*; D051381:Rats; D017208:Rats, Wistar; D013268:Stimulation, Chemical; D013379:Substrate Specificity
D019922:GAP-43 Protein; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D010703:Phorbol Esters; D010750:Phosphoproteins; D018698:Glutamic Acid; D011493:Protein Kinase C