Article
- The EMBO Journal (2005) 24, 3927 - 3939
- doi:10.1038/sj.emboj.7600848
Published online: 27 October 2005
Subject Category:
Unstable microtubule capture at kinetochores depleted of the centromere-associated protein CENP-F
Pascale Bomont1, Paul Maddox1, Jagesh V Shah1,a, Arshad B Desai1 and Don W Cleveland1
- Department of Cellular and Molecular Medicine and Ludwig Institute for Cancer Research, University of California at San Diego, La Jolla, CA, USA
Correspondence to:
Don W Cleveland, Department of Cellular and Molecular Medicine, Ludwig Institute for Cancer Research, 3080 CMM-East, University of California at San Diego, 9500 Gilman Drive, La Jolla, CA 92093, USA. Tel.: +1 858 534 7811; Fax: +1 858 534 7659; E-mail: dcleveland@ucsd.edu
aPresent address: Department of Systems Biology, Harvard Medical School, 4 Blackfan Circle, Boston, MA 02115, USA
Received 22 July 2005; Accepted 30 September 2005
Abstract
Centromere protein F (CENP-F) (or mitosin) accumulates to become an abundant nuclear protein in G2, assembles at kinetochores in late G2, remains kinetochore-bound until anaphase, and is degraded at the end of mitosis. Here we show that the absence of nuclear CENP-F does not affect cell cycle progression in S and G2. In a subset of CENP-F depleted cells, kinetochore assembly fails completely, thereby provoking massive chromosome mis-segregation. In contrast, the majority of CENP-F depleted cells exhibit a strong mitotic delay with reduced tension between kinetochores of aligned, bi-oriented sister chromatids and decreased stability of kinetochore microtubules. These latter kinetochores generate mitotic checkpoint signaling when unattached, recruiting maximum levels of Mad2. Use of YFP-marked Mad1 reveals that throughout the mitotic delay some aligned, CENP-F depleted kinetochores continuously recruit Mad1. Others rebind YFP-Mad1 intermittently so as to produce 'twinkling', demonstrating cycles of mitotic checkpoint reactivation and silencing and a crucial role for CENP-F in efficient assembly of a stable microtubule–kinetochore interface.
Keywords:
- centromere associated protein CENP-F,
- kinetochore,
- Mad1,
- Mad2,
- microtubule
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