Phosphorylation of centrin during the cell cycle and its role in centriole separation preceding centrosome duplication.
Once during each cell cycle, mitotic spindle poles arise by separation of newly duplicated centrosomes. We report here the involvement of phosphorylation of the centrosomal protein centrin in this process. We show that centrin is phosphorylated at serine residue 170 during the G(2)/M phase of the cell cycle. Indirect immunofluorescence staining of HeLa cells using a phosphocentrin-specific antibody reveals intense labeling of mitotic spindle poles during prophase and metaphase of the cell division cycle, with diminished staining of anaphase and no staining of telophase and interphase centrosomes. Cultured cells undergo a dramatic increase in centrin phosphorylation following the experimental elevation of PKA activity, suggesting that this kinase can phosphorylate centrin in vivo. Surprisingly, elevated PKA activity also resulted intense phosphocentrin antibody labeling of interphase centrosomes and in the concurrent movement of individual centrioles apart from one another. Taken together, these results suggest that centrin phosphorylation signals the separation of centrosomes at prophase and implicates centrin phosphorylation in centriole separation that normally precedes centrosome duplication.
Lutz
W
W
Department of Biochemistry and Molecular Biology, Tumor Biology Program, Mayo Clinic Foundation, Rochester, Minnesota 55905, USA.
Lingle
W L
WL
McCormick
D
D
Greenwood
T M
TM
Salisbury
J L
JL
D000595:Amino Acid Sequence; D000906:Antibodies / Q000276:immunology; D002135:Calcium-Binding Proteins / Q000276:immunology / Q000378:metabolism*; D002453:Cell Cycle; D002502:Centrioles / Q000648:ultrastructure*; D002868:Chromosomal Proteins, Non-Histone / Q000276:immunology / Q000378:metabolism*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D006367:HeLa Cells; D006801:Humans; D010766:Phosphorylation; D008941:Spindle Apparatus
D000906:Antibodies; D002135:Calcium-Binding Proteins; D002868:Chromosomal Proteins, Non-Histone; C056608:caltractin; D017868:Cyclic AMP-Dependent Protein Kinases