Identification of major nucleolar proteins as candidate mitotic substrates of cdc2 kinase.
Following the identification of the cdc2 kinase as a major element controlling entry of cells into mitosis, it is important to define the physiological target range of this enzyme. Here, we demonstrate that two major nucleolar proteins, nucleolin and NO38, are highly phosphorylated during mitosis. Importantly, the two nucleolar proteins are also phosphorylated by highly purified starfish cdc2 kinase in vitro, on sites that correspond to those observed specifically during mitosis in vivo. A repeated motif (TPXKK) is identified as the likely mitotic phosphoacceptor site in nucleolin, in that a synthetic peptide mimicking this site functions as both a substrate and a competitive inhibitor of cdc2 kinase. These results identify two novel candidate substrates for cdc2 kinase, and they implicate protein phosphorylation in controlling mitotic changes in nucleolar structure and activity.
Peter
M
M
Swiss Institute for Experimental Cancer Research (ISREC), Epalinges.
Nakagawa
J
J
Dorée
M
M
Labbé
J C
JC
Nigg
E A
EA
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D016203:CDC2 Protein Kinase; D002466:Cell Nucleolus / Q000378:metabolism* / Q000648:ultrastructure; D002474:Cell-Free System; D002645:Chickens; D007700:Kinetics; D051379:Mice; D008938:Mitosis; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D000090243:Nucleophosmin; D010449:Peptide Mapping; D010748:Phosphopeptides / Q000302:isolation & purification; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism; D016601:RNA-Binding Proteins; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity; D014357:Trypsin; D014407:Tumor Cells, Cultured / Q000378:metabolism; D000097796:Nucleolin
D000596:Amino Acids; D016203:CDC2 Protein Kinase; D009687:Nuclear Proteins; D000090243:Nucleophosmin; D010748:Phosphopeptides; D010750:Phosphoproteins; D011494:Protein Kinases; D016601:RNA-Binding Proteins; D014357:Trypsin; D000097796:Nucleolin