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PMID: 19450515 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't

Protein architecture of the human kinetochore microtubule attachment site.

Cell ·Vol. 137 ·No. 4 ·2009-05-15 ·Pages 672-84

Wan X, O'Quinn RP, Pierce HL, Joglekar AP, Gall WE, DeLuca JG, Carroll CW, Liu ST, Yen TJ, McEwen BF, Stukenberg PT, Desai A, Salmon ED

Abstract

Chromosome segregation requires assembly of kinetochores on centromeric chromatin to mediate interactions with spindle microtubules and control cell-cycle progression. To elucidate the protein architecture of human kinetochores, we developed a two-color fluorescence light microscopy method that measures average label separation, Delta, at <5 nm accuracy. Delta analysis of 16 proteins representing core structural complexes spanning the centromeric chromatin-microtubule interface, when correlated with mechanical states of spindle-attached kinetochores, provided a nanometer-scale map of protein position and mechanical properties of protein linkages. Treatment with taxol, which suppresses microtubule dynamics and activates the spindle checkpoint, revealed a specific switch in kinetochore architecture. Cumulatively, Delta analysis revealed that compliant linkages are restricted to the proximity of chromatin, suggested a model for how the KMN (KNL1/Mis12 complex/Ndc80 complex) network provides microtubule attachment and generates pulling forces from depolymerization, and identified an intrakinetochore molecular switch that may function in controlling checkpoint activity.

MeSH Terms
Cytoskeletal Proteins DNA-Binding Proteins/metabolism HeLa Cells Humans Kinetochores/chemistry,metabolism Metaphase Microscopy, Fluorescence Microtubule-Associated Proteins/metabolism Microtubules/chemistry,metabolism Nuclear Proteins
Chemicals
Cytoskeletal Proteins DNA-Binding Proteins Knl1 protein, human MIS12 protein, human Microtubule-Associated Proteins NDC80 protein, human Nuclear Proteins
Authors & Affiliations
13 authors, click to expand affiliations / ORCID
Wan Xiaohu
Department of Biology, University of North Carolina, Chapel Hill, Chapel Hill, NC 27599, USA.
O'Quinn Ryan P
Pierce Heather L
Joglekar Ajit P
Gall Walt E
DeLuca Jennifer G
Carroll Christopher W
Liu Song-Tao
Yen Tim J
McEwen Bruce F
Stukenberg P Todd
Desai Arshad
Salmon E D
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Article Info
Journal
Cell
Abbr.
Cell
ISSN
1097-4172
DOI
10.1016/j.cell.2009.03.035
Published
2009-05-15
Pages
672-84
Language
English
Region
United States
NLM ID
0413066
PMCID
PMC2699050
Subset
IM
Grants
NCI NIH HHS · CA06927 · United States
NIGMS NIH HHS · R01 GM060678 · United States
NIGMS NIH HHS · GM60678 · United States
NIGMS NIH HHS · R01 GM060678-08 · United States
NIGMS NIH HHS · R37 GM024364 · United States
NIGMS NIH HHS · GM074215 · United States
NIGMS NIH HHS · GM44762 · United States
NIGMS NIH HHS · GM24364 · United States
NIGMS NIH HHS · R37 GM024364-30 · United States
NIGMS NIH HHS · R37 GM024364-29 · United States
NCI NIH HHS · K01 CA125051 · United States
NIGMS NIH HHS · R01 GM074215 · United States
NIGMS NIH HHS · GM06627 · United States
NIGMS NIH HHS · GM86877 · United States
NIGMS NIH HHS · R37 GM024364-31 · United States
External Links
PubMed source DOI: 10.1016/j.cell.2009.03.035 PMC Full Text View journal details
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