title: Activation of protein kinase C increases phosphorylation of the L-myc trans-activator domain at a GSK-3 target site.
abstract: The L-myc protein migrates as three distinct differentially phosphorylated bands in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). This phosphorylation can be rapidly increased either by treatment with the protein kinase C (PKC) activator phorbol ester 12-O-tetradecanoyl-phorbol-13-acetate (TPA) or by inhibition of serine/threonine protein phosphatases with okadaic acid. In vitro mutagenesis and phosphoamino acid analyses define the N-terminal serine residues 38 and 42 of L-myc as critical targets for the PKC-dependent phosphorylation. These are the exclusive sites of phosphorylation in the N-terminal third of the L-myc protein, and can be phosphorylated in vitro by glycogen synthase kinase 3 beta (GSK-3 beta). A mutant L-myc protein in which these serines have been replaced by alanine residues does not show heterogeneous electrophoretic migration or hyperphosphorylation in response to PKC activation, and is not a substrate for GSK-3 beta in vitro. Similar potential phosphorylation sites are present in c-myc and N-myc in a highly conserved region thought to represent a transcriptional activation domain. We suggest that N-terminal phosphorylation of the L-myc protein is a means of rapid regulation of this oncoprotein, possibly mediated in vivo by the action of GSK-3.
authors.affiliation: Department of Virology, University of Helsinki, Finland.
authors.forename: K
authors.initials: K
authors.lastname: Saksela
authors.forename: T P
authors.initials: TP
authors.lastname: Mäkelä
authors.forename: K
authors.initials: K
authors.lastname: Hughes
authors.forename: J R
authors.initials: JR
authors.lastname: Woodgett
authors.forename: K
authors.initials: K
authors.lastname: Alitalo
mesh_terms: D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002453:Cell Cycle; D002478:Cells, Cultured; D004252:DNA Mutational Analysis; D004988:Ethers, Cyclic / Q000494:pharmacology; D038341:Glycogen Synthase Kinases; D006801:Humans; D066298:In Vitro Techniques; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011493:Protein Kinase C / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D016271:Proto-Oncogene Proteins c-myc / Q000378:metabolism*; D013329:Structure-Activity Relationship; D015534:Trans-Activators / Q000737:chemistry / Q000378:metabolism*
chemical_list: D004988:Ethers, Cyclic; D010750:Phosphoproteins; D016271:Proto-Oncogene Proteins c-myc; D015534:Trans-Activators; D010768:Phosphoserine; D019319:Okadaic Acid; D011494:Protein Kinases; D038341:Glycogen Synthase Kinases; D011493:Protein Kinase C; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases