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Mol Biol Cell. 2009 Nov;20(22):4790-803. doi: 10.1091/mbc.E09-07-0560. Epub 2009 Sep 30.

Murine CENP-F regulates centrosomal microtubule nucleation and interacts with Hook2 at the centrosome.

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1
Stahlman Cardiovascular Research Laboratories, Program in Developmental Biology, Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, TN 37232-6300, USA.

Abstract

The microtubule (MT) network is essential in a broad spectrum of cellular functions. Many studies have linked CENP-F to MT-based activities as disruption of this protein leads to major changes in MT structure and function. Still, the basis of CENP-F regulation of the MT network remains elusive. Here, our studies reveal a novel and critical localization and role for CENP-F at the centrosome, the major MT organizing center (MTOC) of the cell. Using a yeast two-hybrid screen, we identify Hook2, a linker protein that is essential for regulation of the MT network at the centrosome, as a binding partner of CENP-F. With recently developed immunochemical reagents, we confirm this interaction and reveal the novel localization of CENP-F at the centrosome. Importantly, in this first report of CENP-F(-/-) cells, we demonstrate that ablation of CENP-F protein function eliminates MT repolymerization after standard nocodazole treatment. This inhibition of MT regrowth is centrosome specific because MT repolymerization is readily observed from the Golgi in CENP-F(-/-) cells. The centrosome-specific function of CENP-F in the regulation of MT growth is confirmed by expression of truncated CENP-F containing only the Hook2-binding domain. Furthermore, analysis of partially reconstituted MTOC asters in cells that escape complete repolymerization block shows that disruption of CENP-F function impacts MT nucleation and anchoring rather than promoting catastrophe. Our study reveals a major new localization and function of CENP-F at the centrosome that is likely to impact a broad array of MT-based actions in the cell.

PMID:
19793914
PMCID:
PMC2777108
DOI:
10.1091/mbc.E09-07-0560
[Indexed for MEDLINE]
Free PMC Article

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