title: B23/nucleophosmin serine 4 phosphorylation mediates mitotic functions of polo-like kinase 1.
abstract: Phosphoprotein profiling by Kinetworks trade mark analysis of M-phase-arrested HeLa cells by nocodazole treatment revealed that a novel mitosis-specific phosphorylation event occurred in the nucleolar protein B23/nucleophosmin at a conserved Ser-4 residue. Consistent with the resemblance of the Ser-4 phosphorylation site to the Polo-like kinase 1 (Plk1) consensus recognition sequence, inhibition of Plk1 by a kinase-defective mutation (K82M) abrogated B23 Ser-4 phosphorylation, whereas activation of Plk1 by a constitutively active mutation (T210D) enhanced its phosphorylation following in vivo transfection and in vitro phosphorylation assays. Depletion of endogenous Plk1 by RNA interference abolished B23 Ser-4 phosphorylation. The physical interaction of Plk1 and B23 was further demonstrated by their co-immunoprecipitation and glutathione S-transferase fusion protein pull-down assays. Interference of Ser-4 phosphorylation of B23 induced multiple mitotic defects in HeLa cells, including aberrant numbers of centrosomes, elongation and fragmentation of nuclei, and incomplete cytokinesis. The phenotypes of B23 mutants are reminiscent of a subset of those described previously in Plk1 mutants. Our findings provide insights into the biochemical mechanism underlying the role of Plk1 in mitosis regulation through the identification of Ser-4 in B23 as a major physiological substrate of Plk1.
authors.affiliation: Kinexus Bioinformatics Corp., Vancouver, British Columbia V6T 1Z3.
authors.forename: Hong
authors.initials: H
authors.lastname: Zhang
authors.forename: Xiaoqing
authors.initials: X
authors.lastname: Shi
authors.forename: Harry
authors.initials: H
authors.lastname: Paddon
authors.forename: Maggie
authors.initials: M
authors.lastname: Hampong
authors.forename: Wei
authors.initials: W
authors.lastname: Dai
authors.forename: Steven
authors.initials: S
authors.lastname: Pelech
mesh_terms: D018797:Cell Cycle Proteins; D004789:Enzyme Activation / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008938:Mitosis / Q000502:physiology*; D009154:Mutation; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D000090243:Nucleophosmin; D010766:Phosphorylation; D011485:Protein Binding; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D012694:Serine; D013379:Substrate Specificity; D000097869:Polo-Like Kinase 1
chemical_list: D018797:Cell Cycle Proteins; D009687:Nuclear Proteins; D000090243:Nucleophosmin; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D012694:Serine; D000097869:Polo-Like Kinase 1; C000716967:NPM1 protein, human