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J Cell Biol. 2015 Dec 21;211(6):1141-56. doi: 10.1083/jcb.201508072. Epub 2015 Dec 14.

Whole-proteome genetic analysis of dependencies in assembly of a vertebrate kinetochore.

Author information

1
Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, University of Edinburgh, Edinburgh EH9 3BF, Scotland, UK.
2
Department of Molecular Genetics, National Institute of Genetics and The Graduate University for Advanced Studies, Mishima, Shizuoka 411-8540, Japan Graduate School of Frontier Biosciences, Osaka University, Suita, Osaka 565-0871, Japan.
3
Department of Molecular Genetics, National Institute of Genetics and The Graduate University for Advanced Studies, Mishima, Shizuoka 411-8540, Japan.
4
Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, University of Edinburgh, Edinburgh EH9 3BF, Scotland, UK Department of Bioanalytics, Institute of Biotechnology, Technische Universität Berlin, 13355 Berlin, Germany.
5
Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, University of Edinburgh, Edinburgh EH9 3BF, Scotland, UK bill.earnshaw@ed.ac.uk.

Abstract

Kinetochores orchestrate mitotic chromosome segregation. Here, we use quantitative mass spectrometry of mitotic chromosomes isolated from a comprehensive set of chicken DT40 mutants to examine the dependencies of 93 confirmed and putative kinetochore proteins for stable association with chromosomes. Clustering and network analysis reveal both known and unexpected aspects of coordinated behavior for members of kinetochore protein complexes. Surprisingly, CENP-T depends on CENP-N for chromosome localization. The Ndc80 complex exhibits robust correlations with all other complexes in a "core" kinetochore network. Ndc80 associated with CENP-T interacts with a cohort of Rod, zw10, and zwilch (RZZ)-interacting proteins that includes Spindly, Mad1, and CENP-E. This complex may coordinate microtubule binding with checkpoint signaling. Ndc80 associated with CENP-C forms the KMN (Knl1, Mis12, Ndc80) network and may be the microtubule-binding "workhorse" of the kinetochore. Our data also suggest that CENP-O and CENP-R may regulate the size of the inner kinetochore without influencing the assembly of the outer kinetochore.

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PMID:
26668330
PMCID:
PMC4687880
DOI:
10.1083/jcb.201508072
[Indexed for MEDLINE]
Free PMC Article

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