Purified maturation promoting factor phosphorylates pp60c-src at the sites phosphorylated during fibroblast mitosis.
We have previously shown that overexpressed chicken pp60c-src has retarded mobility, novel serine/threonine phosphorylation, and enhanced kinase activity during NIH 3T3 cell mitosis. Here we show that novel mitotic phosphorylations occur at Thr 34, Thr 46, and Ser 72. The possibility, previously raised, that Ser 17 is dephosphorylated during mitosis is excluded. The phosphorylated sites lie in consensus sequences for phosphorylation by p34cdc2, the catalytic component of maturation promoting factor (MPF). Furthermore, highly purified MPF from metaphase-arrested Xenopus eggs phosphorylated both wild-type and kinase-defective pp60c-src at these sites. Altered phosphorylation alone is sufficient to account for the large retardation in mitotic pp60c-src electrophoretic mobility: phosphorylation of normal pp60c-src by MPF retarded mobility and dephosphorylation of mitotic pp60c-src restored normal mobility. These results suggest that pp60c-src is one of the targets for MPF action, which may account in part for the pleiotropic changes in protein phosphorylation and cellular architecture that occur during mitosis.
Shenoy
S
S
Department of Molecular and Cell Biology, Pennsylvania State University, University Park 16802.
Choi
J K
JK
Bagrodia
S
S
Copeland
T D
TD
Maller
J L
JL
Shalloway
D
D
D000595:Amino Acid Sequence; D000818:Animals; D002478:Cells, Cultured; D002645:Chickens; D005347:Fibroblasts / Q000166:cytology; D006133:Growth Substances / Q000302:isolation & purification / Q000378:metabolism*; D016200:Maturation-Promoting Factor; D000090204:Mesothelin; D051379:Mice; D008815:Mice, Inbred Strains; D008938:Mitosis; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000302:isolation & purification; D010748:Phosphopeptides / Q000302:isolation & purification; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D016392:Proto-Oncogene Proteins pp60(c-src); D012694:Serine; D013912:Threonine; D014357:Trypsin
D006133:Growth Substances; C000715528:Msln protein, mouse; D010446:Peptide Fragments; D010748:Phosphopeptides; D011518:Proto-Oncogene Proteins; D013912:Threonine; D012694:Serine; D011494:Protein Kinases; D016392:Proto-Oncogene Proteins pp60(c-src); D016200:Maturation-Promoting Factor; D014357:Trypsin; D000090204:Mesothelin