A quantitative description of Ndc80 complex linkage to human kinetochores

Nature Communications
Aussie SuzukiE D Salmon

Abstract

The Ndc80 complex, which mediates end-on attachment of spindle microtubules, is linked to centromeric chromatin in human cells by two inner kinetochore proteins, CENP-T and CENP-C. Here to quantify their relative contributions to Ndc80 recruitment, we combine measurements of kinetochore protein copy number with selective protein depletion assays. This approach reveals about 244 Ndc80 complexes per human kinetochore (∼14 per kinetochore microtubule), 215 CENP-C, 72 CENP-T and only 151 Ndc80s as part of the KMN protein network (1:1:1 Knl1, Mis12 and Ndc80 complexes). Each CENP-T molecule recruits ∼2 Ndc80 complexes; one as part of a KMN network. In contrast, ∼40% of CENP-C recruits only a KMN network. Replacing the CENP-C domain that binds KMN with the CENP-T domain that recruits both an Ndc80 complex and KMN network yielded functional kinetochores. These results provide a quantitative picture of the linkages between centromeric chromatin and the microtubule-binding Ndc80 complex at the human kinetochore.

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Citations

Mar 15, 2016·Nature Cell Biology·Aussie SuzukiKerry Bloom
Apr 4, 2016·Current Opinion in Structural Biology·Marion E PesentiAndrea Musacchio
Jul 5, 2016·Chromosoma·Harsh Nagpal, Tatsuo Fukagawa
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Feb 1, 2017·Nature Reviews. Genetics·Sebastian Müller, Geneviève Almouzni
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May 11, 2019·Cell Cycle·Masatoshi Hara, Tatsuo Fukagawa
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Sep 8, 2019·The Journal of Cell Biology·Jonathan Kuhn, Sophie Dumont
Nov 5, 2019·The Journal of Cell Biology·Reito WatanabeTatsuo Fukagawa
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Feb 24, 2021·The Journal of Cell Biology·Roshan L ShresthaMunira A Basrai
May 28, 2019·Current Opinion in Cell Biology·Alexandra F LongSophie Dumont
Apr 11, 2021·Seminars in Cell & Developmental Biology·Fioranna Renda, Alexey Khodjakov
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Jul 2, 2021·Science Advances·Kai WalsteinAndrea Musacchio
Nov 11, 2021·Cell Reports·Diana PapiniJonathan M G Higgins

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Methods Mentioned

BETA
fluorescence microscopy
gel filtration
transfections
transfection

Software Mentioned

MetaMorph

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