title: Protein architecture of the human kinetochore microtubule attachment site.
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.
authors.affiliation: Department of Biology, University of North Carolina, Chapel Hill, Chapel Hill, NC 27599, USA.
authors.forename: Xiaohu
authors.initials: X
authors.lastname: Wan
authors.forename: Ryan P
authors.initials: RP
authors.lastname: O'Quinn
authors.forename: Heather L
authors.initials: HL
authors.lastname: Pierce
authors.forename: Ajit P
authors.initials: AP
authors.lastname: Joglekar
authors.forename: Walt E
authors.initials: WE
authors.lastname: Gall
authors.forename: Jennifer G
authors.initials: JG
authors.lastname: DeLuca
authors.forename: Christopher W
authors.initials: CW
authors.lastname: Carroll
authors.forename: Song-Tao
authors.initials: ST
authors.lastname: Liu
authors.forename: Tim J
authors.initials: TJ
authors.lastname: Yen
authors.forename: Bruce F
authors.initials: BF
authors.lastname: McEwen
authors.forename: P Todd
authors.initials: PT
authors.lastname: Stukenberg
authors.forename: Arshad
authors.initials: A
authors.lastname: Desai
authors.forename: E D
authors.initials: ED
authors.lastname: Salmon
mesh_terms: D003598:Cytoskeletal Proteins; D004268:DNA-Binding Proteins / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000737:chemistry* / Q000378:metabolism*; D008677:Metaphase; D008856:Microscopy, Fluorescence; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008870:Microtubules / Q000737:chemistry* / Q000378:metabolism*; D009687:Nuclear Proteins
chemical_list: D003598:Cytoskeletal Proteins; D004268:DNA-Binding Proteins; C472643:Knl1 protein, human; C492664:MIS12 protein, human; D008869:Microtubule-Associated Proteins; C120529:NDC80 protein, human; D009687:Nuclear Proteins