Title: CENP-C Is a Structural Platform for Kinetochore Assembly
Abstract: Centromeres provide a region of chromatin upon which kinetochores are assembled in mitosis [1Przewloka M.R. Glover D.M. The kinetochore and the centromere: A working long distance relationship.Annu. Rev. Genet. 2009; 43: 439-465Crossref PubMed Scopus (88) Google Scholar, 2Santaguida S. Musacchio A. The life and miracles of kinetochores.EMBO J. 2009; 28: 2511-2531Crossref PubMed Scopus (341) Google Scholar]. Centromeric protein C (CENP-C) is a core component of this centromeric chromatin [3Saitoh H. Tomkiel J. Cooke C.A. Ratrie 3rd, H. Maurer M. Rothfield N.F. Earnshaw W.C. CENP-C, an autoantigen in scleroderma, is a component of the human inner kinetochore plate.Cell. 1992; 70: 115-125Abstract Full Text PDF PubMed Scopus (308) Google Scholar, 4Kwon M.S. Hori T. Okada M. Fukagawa T. CENP-C is involved in chromosome segregation, mitotic checkpoint function, and kinetochore assembly.Mol. Biol. Cell. 2007; 18: 2155-2168Crossref PubMed Scopus (87) Google Scholar] that, when depleted, prevents the proper formation of both centromeres and kinetochores [5Fukagawa T. Brown W.R. Efficient conditional mutation of the vertebrate CENP-C gene.Hum. Mol. Genet. 1997; 6: 2301-2308Crossref PubMed Scopus (83) Google Scholar, 6Heeger S. Leismann O. Schittenhelm R. Schraidt O. Heidmann S. Lehner C.F. Genetic interactions of separase regulatory subunits reveal the diverged Drosophila Cenp-C homolog.Genes Dev. 2005; 19: 2041-2053Crossref PubMed Scopus (70) Google Scholar, 7Tanaka K. Chang H.L. Kagami A. Watanabe Y. CENP-C functions as a scaffold for effectors with essential kinetochore functions in mitosis and meiosis.Dev. Cell. 2009; 17: 334-343Abstract Full Text Full Text PDF PubMed Scopus (78) Google Scholar, 8Milks K.J. Moree B. Straight A.F. Dissection of CENP-C-directed centromere and kinetochore assembly.Mol. Biol. Cell. 2009; 20: 4246-4255Crossref PubMed Scopus (80) Google Scholar, 9Przewloka M.R. Zhang W. Costa P. Archambault V. D'Avino P.P. Lilley K.S. Laue E.D. McAinsh A.D. Glover D.M. Molecular analysis of core kinetochore composition and assembly in Drosophila melanogaster.PLoS ONE. 2007; 2: e478Crossref PubMed Scopus (93) Google Scholar, 10Orr B. Sunkel C.E. Drosophila CENP-C is essential for centromere identity.Chromosoma. 2011; 120: 83-96Crossref PubMed Scopus (21) Google Scholar]. CENP-C localizes to centromeres throughout the cell cycle via its C-terminal part [6Heeger S. Leismann O. Schittenhelm R. Schraidt O. Heidmann S. Lehner C.F. Genetic interactions of separase regulatory subunits reveal the diverged Drosophila Cenp-C homolog.Genes Dev. 2005; 19: 2041-2053Crossref PubMed Scopus (70) Google Scholar, 8Milks K.J. Moree B. Straight A.F. Dissection of CENP-C-directed centromere and kinetochore assembly.Mol. Biol. Cell. 2009; 20: 4246-4255Crossref PubMed Scopus (80) Google Scholar], whereas its N-terminal part appears necessary for recruitment of some but not all components of the Mis12 complex of the kinetochore [8Milks K.J. Moree B. Straight A.F. Dissection of CENP-C-directed centromere and kinetochore assembly.Mol. Biol. Cell. 2009; 20: 4246-4255Crossref PubMed Scopus (80) Google Scholar]. We now find that all kinetochore proteins belonging to the KMN (KNL1/Spc105, the Mis12 complex, and the Ndc80 complex) network [1Przewloka M.R. Glover D.M. The kinetochore and the centromere: A working long distance relationship.Annu. Rev. Genet. 2009; 43: 439-465Crossref PubMed Scopus (88) Google Scholar] bind to the N-terminal part of Drosophila CENP-C. Moreover, we show that the Mis12 complex component Nnf1 interacts directly with CENP-C in vitro. To test whether CENP-C's N-terminal part was sufficient to recruit KMN proteins, we targeted it to the centrosome by fusing it to a domain of Plk4 kinase [11Dzhindzhev N.S. Yu Q.D. Weiskopf K. Tzolovsky G. Cunha-Ferreira I. Riparbelli M. Rodrigues-Martins A. Bettencourt-Dias M. Callaini G. Glover D.M. Asterless is a scaffold for the onset of centriole assembly.Nature. 2010; 467: 714-718Crossref PubMed Scopus (209) Google Scholar]. The Mis12 and Ndc80 complexes and Spc105 protein were then all recruited to centrosomes at the expense of centromeres, leading to mitotic abnormalities typical of cells with defective kinetochores. Thus, the N-terminal part of Drosophila CENP-C is sufficient to recruit core kinetochore components and acts as the principal linkage between centromere and kinetochore during mitosis.