Polo kinase and Asp are needed to promote the mitotic organizing activity of centrosomes.
Interfering with the activity of polo-like kinases can lead to the formation of monopolar spindles. Polo-like kinases also regulate mitotic entry, activation of the anaphase-promoting complex and the necessary preconditions for cytokinesis. Similarities between the phenotypes of the Drosophila mutants asp and polo point towards a common role in spindle pole function. The abnormal spindles of asp mutants are bipolar but have disorganized broad poles at which gamma-tubulin has an abnormal distribution. Moreover, the synergism or of polo1 aspE3 double mutants indicates a possible involvement of these genes in a common process. Asp is a microtubule-associated protein of relative molecular mass 220,000 (Mr 220K) found at the face of the centrosome that contacts spindle microtubules. In partially purified centrosomes, it is required with gamma-tubulin to organize microtubule asters. Here, we show that Asp is the previously identified Mr 220K substrate of Polo kinase. Polo phosphorylates Asp in vitro, converting it into an MPM2 epitope. Polo and Asp proteins immunoprecipitate together and exist as part of a 25-38S complex. Extracts of polo-derived embryos are unable to restore the ability of salt-stripped centrosomes to nucleate microtubule asters. This can be rescued by addition of phosphorylated Asp or active Polo kinase.
Cancer Research Campaign Cell Cycle Genetics Group, University of Cambridge, Department of Genetics, Downing Street, Cambridge, UK CB2 3EH.
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do Carmo Avides
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Tavares
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Glover
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D016203:CDC2 Protein Kinase / Q000378:metabolism; D018385:Centrosome / Q000378:metabolism*; D004330:Drosophila / Q000196:embryology / Q000378:metabolism; D029721:Drosophila Proteins; D018985:Epitopes, B-Lymphocyte / Q000276:immunology; D051379:Mice; D008840:Microfilament Proteins / Q000276:immunology / Q000378:metabolism*; D008869:Microtubule-Associated Proteins / Q000276:immunology / Q000378:metabolism*; D008938:Mitosis / Q000502:physiology*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D051381:Rats
D000911:Antibodies, Monoclonal; D029721:Drosophila Proteins; D018985:Epitopes, B-Lymphocyte; D008840:Microfilament Proteins; D008869:Microtubule-Associated Proteins; C106174:asp protein, Drosophila; C072004:polo protein, Drosophila; D017346:Protein Serine-Threonine Kinases; D016203:CDC2 Protein Kinase