Mitosis-specific Anchoring of γ Tubulin Complexes by Pericentrin Controls Spindle Organization and Mitotic Entry
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Published:2004-08
Issue:8
Volume:15
Page:3642-3657
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ISSN:1059-1524
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Container-title:Molecular Biology of the Cell
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language:en
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Short-container-title:MBoC
Author:
Zimmerman Wendy C.1, Sillibourne James1, Rosa Jack1, Doxsey Stephen J.1
Affiliation:
1. Department of Molecular Medicine, University of Massachusetts Medical School, Worcester, Massachusetts 01605
Abstract
Microtubule nucleation is the best known function of centrosomes. Centrosomal microtubule nucleation is mediated primarily by γ tubulin ring complexes (γ TuRCs). However, little is known about the molecules that anchor these complexes to centrosomes. In this study, we show that the centrosomal coiled-coil protein pericentrin anchors γ TuRCs at spindle poles through an interaction with γ tubulin complex proteins 2 and 3 (GCP2/3). Pericentrin silencing by small interfering RNAs in somatic cells disrupted γ tubulin localization and spindle organization in mitosis but had no effect on γ tubulin localization or microtubule organization in interphase cells. Similarly, overexpression of the GCP2/3 binding domain of pericentrin disrupted the endogenous pericentrin–γ TuRC interaction and perturbed astral microtubules and spindle bipolarity. When added to Xenopus mitotic extracts, this domain uncoupled γ TuRCs from centrosomes, inhibited microtubule aster assembly, and induced rapid disassembly of preassembled asters. All phenotypes were significantly reduced in a pericentrin mutant with diminished GCP2/3 binding and were specific for mitotic centrosomal asters as we observed little effect on interphase asters or on asters assembled by the Ran-mediated centrosome-independent pathway. Additionally, pericentrin silencing or overexpression induced G2/antephase arrest followed by apoptosis in many but not all cell types. We conclude that pericentrin anchoring of γ tubulin complexes at centrosomes in mitotic cells is required for proper spindle organization and that loss of this anchoring mechanism elicits a checkpoint response that prevents mitotic entry and triggers apoptotic cell death.
Publisher
American Society for Cell Biology (ASCB)
Subject
Cell Biology,Molecular Biology
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