title: The Ndc80 complex uses a tripartite attachment point to couple microtubule depolymerization to chromosome movement.
abstract: In kinetochores, the Ndc80 complex couples the energy in a depolymerizing microtubule to perform the work of moving chromosomes. The complex directly binds microtubules using an unstructured, positively charged N-terminal tail located on Hec1/Ndc80. Hec1/Ndc80 also contains a calponin homology domain (CHD) that increases its affinity for microtubules in vitro, yet whether it is required in cells and how the tail and CHD work together are critical unanswered questions. Human kinetochores containing Hec1/Ndc80 with point mutations in the CHD fail to align chromosomes or form productive microtubule attachments. Kinetochore architecture and spindle checkpoint protein recruitment are unaffected in these mutants, and the loss of CHD function cannot be rescued by removing Aurora B sites from the tail. The interaction between the Hec1/Ndc80 CHD and a microtubule is facilitated by positively charged amino acids on two separate regions of the CHD, and both are required for kinetochores to make stable attachments to microtubules. Chromosome congression in cells also requires positive charge on the Hec1 tail to facilitate microtubule contact. In vitro binding data suggest that charge on the tail regulates attachment by directly increasing microtubule affinity as well as driving cooperative binding of the CHD. These data argue that in vertebrates there is a tripartite attachment point facilitating the interaction between Hec1/Ndc80 and microtubules. We discuss how such a complex microtubule-binding interface may facilitate the coupling of depolymerization to chromosome movement.
authors.affiliation: Department of Biochemistry and Molecular Genetics, University of Virginia Medical Center, Charlottesville, VA 22908, USA.
authors.forename: John G
authors.initials: JG
authors.lastname: Tooley
authors.forename: Stephanie A
authors.initials: SA
authors.lastname: Miller
authors.forename: P Todd
authors.initials: PT
authors.lastname: Stukenberg
mesh_terms: D000818:Animals; D064107:Aurora Kinase B; D064127:Aurora Kinases; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000378:metabolism; D002453:Cell Cycle / Q000502:physiology; D020090:Chromosome Segregation / Q000502:physiology*; D002942:Circular Dichroism; D003598:Cytoskeletal Proteins; D005260:Female; D020868:Gene Silencing; D006367:HeLa Cells; D006801:Humans; D018386:Kinetochores / Q000378:metabolism*; D008840:Microfilament Proteins / Q000737:chemistry / Q000378:metabolism; D008870:Microtubules / Q000378:metabolism*; D008938:Mitosis; D008958:Models, Molecular; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D058105:Polymerization; D011485:Protein Binding / Q000235:genetics; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering; D008941:Spindle Apparatus / Q000378:metabolism; D014162:Transfection; D014714:Vertebrates; D000096985:Calponins
chemical_list: D064107:Aurora Kinase B; D064127:Aurora Kinases; D002135:Calcium-Binding Proteins; D003598:Cytoskeletal Proteins; D008840:Microfilament Proteins; D009687:Nuclear Proteins; D017346:Protein Serine-Threonine Kinases; D034741:RNA, Small Interfering; D000096985:Calponins; C120529:NDC80 protein, human; C576921:AURKB protein, human