Neuron-specific Cdk5 kinase is responsible for mitosis-independent phosphorylation of c-Src at Ser75 in human Y79 retinoblastoma cells.
c-Src is phosphorylated at specific serine and threonine residues during mitosis in fibroblastic and epithelial cells. These sites are phosphorylated in vitro by the mitotic kinase Cdk1 (p34(cdc2)). In contrast, c-Src in Y79 human retinoblastoma cells, which are of neuronal origin, is phosphorylated at one of the mitotic sites, Ser75, throughout the cell cycle. The identity of the serine kinase that nonmitotically phosphorylates c-Src on Ser75 remains unknown. We now are able to show for the first time that Cdk5 kinase, which has the same consensus sequence as the Cdk1 and Cdk2 kinases, is required for the phosphorylation in asynchronous Y79 cells. The Ser75 phosphorylation was inhibited in a dose-dependent manner by butyrolactone I, a specific inhibitor of Cdk5-type kinases. Three stable subclones that have almost no kinase activity were selected by transfection of an antisense Cdk5-specific activator p35 construct into Y79 cells. The loss of the kinase activity caused an approximately 85% inhibition of the Ser75 phosphorylation. These results present compelling evidence that Cdk5/p35 kinase is responsible for the novel phosphorylation of c-Src at Ser75 in neuronal cells, raising the intriguing possibility that c-Src acts as an effector of Cdk5/p35 kinase during neuronal development.
Kato
G
G
Department of Biochemistry, Yamanashi Medical University, Nakakoma, Yamanashi, 409-3898, Japan. gkato@res.yamanashi-med.ac.jp
Maeda
S
S
D015107:4-Butyrolactone / Q000031:analogs & derivatives / Q000494:pharmacology; D001665:Binding Sites; D000081247:CSK Tyrosine-Protein Kinase; D051360:Cyclin-Dependent Kinase 5; D018844:Cyclin-Dependent Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D016373:DNA, Antisense / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D008938:Mitosis; D009474:Neurons / Q000201:enzymology*; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000737:chemistry / Q000378:metabolism*; D012175:Retinoblastoma / Q000201:enzymology; D012694:Serine / Q000737:chemistry; D014162:Transfection; D014407:Tumor Cells, Cultured; D019061:src-Family Kinases
D016373:DNA, Antisense; D004791:Enzyme Inhibitors; D012694:Serine; C082868:butyrolactone I; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; C000631346:CSK protein, human; D051360:Cyclin-Dependent Kinase 5; C495905:CDK5 protein, human; D018844:Cyclin-Dependent Kinases; D015107:4-Butyrolactone