Mayor, Thibault and Hacker, Ulrike and Stierhof, York-Dieter and Nigg, Erich A. (2002) The mechanism regulating the dissociation of the centrosomal protein C-Nap1 from mitotic spindle poles. Journal of Cell Science, 115 (16). pp. 3275-3284. ISSN 0021-9533. http://resolver.caltech.edu/CaltechAUTHORS:20111031-142910138
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The centrosomal protein C-Nap1 is thought to play an important role in centrosome cohesion during interphase of the cell cycle. At the onset of mitosis, when centrosomes separate for bipolar spindle formation, C-Nap1 dissociates from centrosomes. Here we report the results of experiments aimed at determining whether the dissociation of C-Nap1 from mitotic centrosomes is triggered by proteolysis or phosphorylation. Specifically, we analyzed both the cell cycle regulation of endogenous C-Nap1 and the fate of exogenously expressed full-length C-Nap1. Western blot analyses suggested a reduction in the endogenous C-Nap1 level during M phase, but studies using proteasome inhibitors and destruction assays performed in Xenopus extracts argue against ubiquitin-dependent degradation of C-Nap1. Instead, our data indicate that the mitotic C-Nap1 signal is reduced as a consequence of M-phase-specific phosphorylation. Overexpression of full-length C-Nap1 in human U2OS cells caused the formation of large structures that embedded the centrosome and impaired its microtubule nucleation activity. Remarkably, however, these centrosome-associated structures did not interfere with cell division. Instead, centrosomes were found to separate from these structures at the onset of mitosis, indicating that a localized and cell-cycle-regulated activity can dissociate C-Nap1 from centrosomes. A prime candidate for this activity is the centrosomal protein kinase Nek2, as the formation of large C-Nap1 structures was substantially reduced upon co-expression of active Nek2. We conclude that the dissociation of C-Nap1 from mitotic centrosomes is regulated by localized phosphorylation rather than generalized proteolysis.
|Additional Information:||© 2002 The Company of Biologists Limited. Accepted May 26, 2002. We thank Olaf Kelm for providing Xenopus extracts and all colleagues in our laboratory for discussions. In its initial stage, this work was supported by the Swiss National Science Foundation (31-50576.97) and the Canton of Geneva.|
|Subject Keywords:||C-Nap1; Nek2; centrosome; mitotic spindle; phosphorylation|
|Usage Policy:||No commercial reproduction, distribution, display or performance rights in this work are provided.|
|Deposited By:||Tony Diaz|
|Deposited On:||01 Nov 2011 21:12|
|Last Modified:||26 Dec 2012 14:21|
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