Replacement of Ser657 of protein kinase C-alpha by alanine leads to premature down regulation after phorbol-ester-induced translocation to the membrane.
Protein kinase C (PKC) is activated at the cell membrane by interacting with both the acidic lipid phosphatidylserine and the second messenger diacylglycerol. A direct activation of the kinase is also possible by substituting diacylglycerol with phorbol esters such as the tumor promoting agent 12-O-tetradecanoylphorbol-13-acetate (TPA). Transphosphorylation of the activation loop followed by autophosphorylation at sites located on various domains of the protein have been suggested to be required as permissive activation of the alpha and beta isoforms of PKC [Cazaubon, S., Bornancin, F. & Parker, P. (1994) Biochem. J. 301, 443-448; Keranen, L. M., Dutil, E. M. & Newton, A. C. (1995) Curr. Biol. 5, 1393-1403]. Ser657, located near the C-terminus of PKC-alpha, represents a site which is very conserved among the members of the PKC protein family. Circumstantial evidence suggested that this residue represents a possible site of phosphorylation. The conversion of Ser657 to alanine caused a 70% loss of the catalytic activity as well as a drastically increased down regulation upon translocation of this isozyme to the membrane when induced by phorbol ester. The faster electrophoretic mobility of the mutant protein compared to that of the wild-type enzyme suggested that Ser657 represents a phosphorylation site.
Gysin
S
S
Department of Research, University Hospital, Basel, Switzerland.
Imber
R
R
D000409:Alanine / Q000737:chemistry; D000818:Animals; D001665:Binding Sites; D001693:Biological Transport, Active / Q000187:drug effects; D002417:Cattle; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D015536:Down-Regulation; D004591:Electrophoresis, Polyacrylamide Gel; D007527:Isoenzymes / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018374:LLC-PK1 Cells; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D011493:Protein Kinase C / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D051571:Protein Kinase C-alpha; D012694:Serine / Q000737:chemistry; D013552:Swine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D007527:Isoenzymes; D012694:Serine; D011493:Protein Kinase C; D051571:Protein Kinase C-alpha; D013755:Tetradecanoylphorbol Acetate; D000409:Alanine