Abstract
Kinetochores are large proteinaceous structure on the surface of chromosomes’ primary constriction during mitosis. They link chromosomes to spindle microtubules and also regulate the spindle assembly checkpoint, which is crucial for correct chromosome segregation in all eukaryotes. The better known core networks of kinetochores include the KMN network (K, KNL1; M, Mis12 complex; N, Ndc80 complex)and CCAN (constitutive centromere-associated network). However, the detailed molecular mechanism of the kinetochore protein network remains unclear. This study demonstrates that CENP-H and CENP-K form quite stable subcomplex by TAP (tandem affinity purification) with HEK 293 cells which express TAP-CENP-K, with the ratio of purified CENP-H and CENP-K being close to 1: 1 even with high salt. Bioinformatic analysis suggests that CENP-H and CENP-K are enriched with coiled-coil regions. This implies that CENP-H and CENP-K form heterodimeric coiled-coils. Furthermore, the functional regions which form the complex are respectively located on their N- and C-terminals, but the association between the C-terminals is more complex. It is possible that this is the first identified heterodimeric coiled-coils within the inner kinetochore, which is directly involved in the attachment between kinetochores and the spindle microtubules.
Similar content being viewed by others
References
Cleveland D W, Mao Y, Sullivan K F. Centromeres and kinetochores: from epigenetics to mitotic checkpoint signaling. Cell, 2003, 4: 407–421 10.1016/S0092-8674(03)00115-6
Maiato H, DeLuca J, Salmon E D, et al. The dynamic kinetochore-microtubule interface. J Cell Sci, 2004, Pt 23: 5461–5477 10.1242/jcs.01536
Kline-Smith S L, Sandall S, Desai A. Kinetochore-spindle microtubule interactions during mitosis. Curr Opin Cell Biol, 2005, 1: 35–46 10.1016/j.ceb.2004.12.009
Cheeseman I M, Desai A. Molecular architecture of the kinetochore-microtubule interface. Nat Rev Mol Cell Biol 2008, 1: 33–46 10.1038/nrm2310
Cheeseman I M, Niessen S, Anderson S, et al. A conserved protein network controls assembly of the outer kinetochore and its ability to sustain tension. Genes Dev, 2004, 18: 2255–2268 15371340, 10.1101/gad.1234104, 1:CAS:528:DC%2BD2cXnslCnuro%3D
Tanaka T U, Desai A. Kinetochore-microtubule interactions: the means to the end. Curr Opin Cell Biol, 2008, 1: 53–63
Cheeseman I M, Chappie J S, Wilson-Kubalek E M, et al. The conserved KMN network constitutes the core microtubule-binding site of the kinetochore. Cell 2006, 5: 983–997 10.1016/j.cell.2006.09.039
Foltz D R, Jansen L E, Black B E, et al. The human CENP-A centromeric nucleosome-associated complex. Nat Cell Biol, 2006, 5: 458–469 10.1038/ncb1397
He D, Zeng C, Woods K, et al. CENP-G: a new centromeric protein that is associated with the alpha-1 satellite DNA subfamily. Chromosoma, 1998, 3: 189–197 10.1007/s004120050296
Okada M, Cheeseman I M, Hori T, et al. The CENP-H-I complex is required for the efficient incorporation of newly synthesized CENP-A into centromeres. Nat Cell Biol, 2006, 5: 446–457 10.1038/ncb1396
Black B E, Brock M A, Bedard S, et al. An epigenetic mark generated by the incorporation of CENP-A into centromeric nucleosomes. Proc Natl Acad Sci USA, 2007, 12: 5008–5013 10.1073/pnas.0700390104
McClelland S E, Borusu S, Amaro A C, et al. The CENP-A NAC/CAD kinetochore complex controls chromosome congression and spindle bipolarity. EMBO J, 2007, 24: 5033–5047 10.1038/sj.emboj.7601927
Hori T, Okada M, Maenaka K, et al. CENP-O-Class proteins form a stable complex and are required for proper kinetochore function. Mol Biol Cell, 2007, 3: 843–854 10.1091/mbc.E07-06-0556
Cheeseman I M, Hori T, Fukagawa T, et al. KNL1 and the CENP-H/I/K complex coordinately direct kinetochore assembly in vertebrates. Mol Biol Cell, 2007, 2: 587–594 10.1091/mbc.E07-10-1051
Liu S T, Hittle J C, Jablonski S A, et al. Human CENP-I specifies localization of CENP-F, MAD1 and MAD2 to kinetochores and is essential for mitosis. Nat Cell Biol, 2003, 4: 341–345 10.1038/ncb953
Ciferri C, Pasqualato S, Screpanti E, et al. Implications for kinetochore-microtubule attachment from the structure of an engineered Ndc80 complex. Cell, 2008, 3: 427–439 10.1016/j.cell.2008.03.020
Kline S L, Cheeseman I M, Hori T, et al. The human Mis12 complex is required for kinetochore assembly and proper chromosome segregation. J Cell Biol, 2006, 1: 9–17 10.1083/jcb.200509158
Wei R R, Al-Bassam J, Harrison S C. The Ndc80/HEC1 complex is a contact point for kinetochore-microtubule attachment. Nat Struct Mol Biol, 2007, 1: 54–59 10.1038/nsmb1186
McCleland M L, Gardner R D, Kallio M J, et al. The highly conserved Ndc80 complex is required for kinetochore assembly, chromosome congression, and spindle checkpoint activity. Genes Dev, 2003, 1: 101–114 10.1101/gad.1040903
Wei R R, Sorger P K, Harrison S C. Molecular organization of the Ndc80 complex, an essential kinetochore component. Proc Natl Acad Sci USA, 2005, 15: 5363–5367 10.1073/pnas.0501168102
Lupas A, Van Dyke M, Stock J. Predicting coiled coils from protein sequences. Science, 1991, 5009: 1162–1164 10.1126/science.252.5009.1162
McDonnell A V, Jiang T, Keating A E, et al. Paircoil2: improved prediction of coiled coils from sequence. Bioinformatics, 2006, 3: 356–358
Earnshaw W C, Rothfield N. Identification of a family of human centromere proteins using autoimmune sera from patients with scleroderma. Chromosoma, 1985, 3–4: 313–321 10.1007/BF00328227
Fukagawa T, Mikami Y, Nishihashi A, et al. CENP-H, a constitutive centromere component, is required for centromere targeting of CENP-C in vertebrate cells. EMBO J, 2001, 16: 4603–4617 10.1093/emboj/20.16.4603
Westermann S, Cheeseman I M, Anderson S, et al. Architecture of the budding yeast kinetochore reveals a conserved molecular core. J Cell Biol, 2003, 2: 215–222 10.1083/jcb.200305100
Mikami Y, Hori T, Kimura H, et al. The functional region of CENP-H interacts with the Nuf2 complex that localizes to centromere during mitosis. Mol Cell Biol, 2005, 5: 1958–1970 10.1128/MCB.25.5.1958-1970.2005
Dong H, Paramonov S E, Hartgerink J D. Self-assembly of alpha-helical coiled coil nanofibers. J Am Chem Soc, 2008, 41: 13691–13695 10.1021/ja8037323
Kajava A V, Potekhin S A, Corradin G, et al. Organization of designed nanofibrils assembled from alpha-helical peptides as determined by electron microscopy. J Pept Sci, 2004, 5: 291–297 10.1002/psc.520
Pandya M J, Spooner G M, Sunde M, et al. Sticky-end assembly of a designed peptide fiber provides insight into protein fibrillogenesis. Biochemistry, 2000, 30: 8728–8734 10.1021/bi000246g
Potekhin S A, Melnik T N, Popov V, et al. De novo design of fibrils made of short alpha-helical coiled coil peptides. Chem Biol, 2001, 11: 1025–1032 10.1016/S1074-5521(01)00073-4
Author information
Authors and Affiliations
Corresponding author
Additional information
Supported by the National Key Scientific Program (Grant No. 2006CB910100).
Rights and permissions
About this article
Cite this article
Qiu, S., Wang, J., Yu, C. et al. CENP-K and CENP-H may form coiled-coils in the kinetochores. SCI CHINA SER C 52, 352–359 (2009). https://doi.org/10.1007/s11427-009-0050-3
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s11427-009-0050-3