Cell cycle-dependent phosphorylation of centrosomes: Localization of phosphopeptide specific antibodies to the centrosome
✍ Scribed by Vandr�, Dale D.; Feng, Yang; Ding, Min
- Publisher
- John Wiley and Sons
- Year
- 2000
- Tongue
- English
- Weight
- 235 KB
- Volume
- 49
- Category
- Article
- ISSN
- 1059-910X
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✦ Synopsis
The microtubule nucleation capacity of the centrosome increases dramatically as cells progress from interphase into mitosis. The increase in nucleation capacity of the centrosome correlates with the cell cycle-dependent localization of the mitotic protein monoclonal-2 (MPM-2) phosphoepitopespecific antibody to the mitotic centrosome. Therefore, the phosphorylation state of centrosomal components may regulate the microtubule nucleation capacity of this organelle during mitosis. Neither the identity of the MPM-2 kinase(s) nor all of the MPM-2-reactive phosphoproteins associated with the centrosome have been fully elucidated. Only recently have the characteristics of the MPM-2 epitope site been defined, and we used this information to prepare polyclonal antibodies against synthetic phosphopeptides containing potential MPM-2 epitopes derived from the sequences of two MPM-2-reactive proteins, topoisomerase II, and microtubule associated protein 1B (MAP1B). We demonstrate that these phosphopeptide-specific antibodies also localize to the centrosome in a cell cycle-dependent fashion. Thus, polyclonal antibodies have been generated against defined phosphopeptides that reiterate many of the immunofluorescence staining properties exhibited by the MPM-2 antibody. These new phosphopeptide-specific antibodies will provide additional probes to examine the phosphorylation of centrosomal components and the functional consequences of their phosphorylation during mitosis. Microsc.