The recruitment of cohesins to pericentric chromatin in some organisms appears to require heterochromatin associated with repetitive DNA. However, neocentromeres and budding yeast centromeres lack flanking repetitive DNA, indicating that cohesin recruitment occurs through an alternative pathway. Here, we demonstrate that all budding yeast chromosomes assemble cohesin domains that extend over 20–50 kb of unique pericentric sequences flanking the conserved 120-bp centromeric DNA. The assembly of these cohesin domains requires the presence of a functional kinetochore in every cell cycle. A similar enhancement of cohesin binding was also observed in regions flanking an ectopic centromere. At both endogenous and ectopic locations, the centromeric enhancer amplified the inherent levels of cohesin binding that are unique to each region. Thus, kinetochores are enhancers of cohesin association that act over tens of kilobases to assemble pericentric cohesin domains. These domains are larger than the pericentric regions stretched by microtubule attachments, and thus are likely to counter microtubule-dependent forces. Kinetochores mediate two essential segregation functions: chromosome movement through microtubule attachment and biorientation of sister chromatids through the recruitment of high levels of cohesin to pericentric regions. We suggest that the coordination of chromosome movement and biorientation makes the kinetochore an autonomous segregation unit.
References
[1]
Aagaard L, Schmid M, Warburton P, Jenuwein T (2000) Mitotic phosphorylation of SUV39H1, a novel component of active centromeres, coincides with transient accumulation at mammalian centromeres. J Cell Sci 113: 817–829.
[2]
Amor DJ, Choo KH (2002) Neocentromeres: Role in human disease, evolution, and centromere study. Am J Hum Genet 71: 695–714.
[3]
Bernard P, Maure JF, Partridge JF, Genier S, Javerzat JP, et al. (2001) Requirement of heterochromatin for cohesion at centromeres. Science 294: 2539–2542.
[4]
Blat Y, Kleckner N (1999) Cohesins bind to preferential sites along yeast chromosome III, with differential regulation along arms versus the centric region. Cell 98: 249–259.
[5]
Bloom KS, Carbon J (1982) Yeast centromere DNA is in a unique and highly ordered structure in chromosomes and small circular minichromosomes. Cell 29: 305–317.
[6]
Bohlander SK, Espinosa R, Le Beau MM, Rowley JD, Diaz MO (1992) A method for the rapid sequence-independent amplification of microdissected chromosomal material. Genomics 13: 1322–1324.
[7]
Campbell D, Doctor JS, Feuersanger JH, Doolittle MM (1981) Differential mitotic stability of yeast disomes derived from triploid meiosis. Genetics 98: 239–255.
[8]
Clarke L, Carbon J (1980) Isolation of a yeast centromere and construction of functional small circular chromosomes. Nature 287: 504–509.
[9]
Doheny KF, Sorger PK, Hyman AA, Tugendreich S, Spencer F, et al. (1993) Identification of essential components of the kinetochore. Cell 73: 761–774.
[10]
Espelin CW, Kaplan KB, Sorger PK (1997) Probing the architecture of a simple kinetochore using DNA-protein crosslinking. J Cell Biol 139: 1383–1396.
[11]
Furuya K, Takahashi K, Yanagida M (1998) Faithful anaphase is ensured by Mis4, a sister chromatid cohesion molecule required in S phase and not destroyed in G1 phase. Genes Dev 12: 3408–3418.
[12]
Gerton JL, DeRisi J, Shroff R, Lichten M, Brown PO, et al. (2000) Inaugural article: Global mapping of meiotic recombination hotspots and coldspots in the yeast . Proc Natl Acad Sci U S A 97: 11383–11390.
[13]
Glynn F, Megee PC, Yu HG, Mistrot C, Unal E, et al. (2004) Genome-wide mapping of the cohesion complex in the yeast . PLoS Biol 2(9): e259. doi: 10.1371/journal.pbio.0020259.
[14]
Goh PY, Kilmartin JV (1993) NDC10: A gene involved in chromosome segregation in . J Cell Biol 121: 503–512.
[15]
Gonzalez C, Casal Jimenez J, Ripoll P, Sunkel CE (1991) The spindle is required for the process of sister chromatid separation in Drosophila neuroblasts. Exp Cell Res 192: 10–15.
[16]
Goshima G, Yanagida M (2000) Establishing biorientation occurs with precocious separation of the sister kinetochores, but not the arms, in the early spindle of budding yeast. Cell 100: 619–633.
[17]
Guacci V, Koshland D, Strunnikov A (1997) A direct link between sister chromatid cohesion and chromosome condensation revealed through the analysis of MCD1 in . Cell 91: 47–57.
[18]
Hanna JS, Kroll ES, Lundblad V, Spencer FA (2001) CTF18 and CTF4 are required for sister chromatid cohesion. Mol Cell Biol 21: 3144–3158.
[19]
Hartman T, Stead K, Koshland D, Guacci V (2000) Pds5p is an essential chromosomal protein required for both sister chromatid cohesion and condensation in . J Cell Biol 151: 613–626.
[20]
He X, Asthana S, Sorger PK (2000) Transient sister chromatid separation and elastic deformation of chromosomes during mitosis in budding yeast. Cell 101: 763–775.
[21]
He X, Rines DR, Espelin CW, Sorger PK (2001) Molecular analysis of kinetochore-microtubule attachment in budding yeast. Cell 106: 195–206.
[22]
Hecht A, Strahl-Bolsinger S, Grunstein M (1996) Spreading of transcriptional repressor SIR3 from telomeric heterochromatin. Nature 383: 92–96.
[23]
Iyer VR, Horak CE, Scafe CS, Botstein D, Snyder M, et al. (2001) Genomic binding sites of the yeast cell-cycle transcription factors SBF and MBF. Nature 409: 533–538.
[24]
Jiang W, Lechner J, Carbon J (1993) Isolation and characterization of a gene (CBF2) specifying a protein component of the budding yeast kinetochore. J Cell Biol 121: 513–519.
[25]
Klein F, Mahr P, Galova M, Buonomo SB, Michaelis C, et al. (1999) A central role for cohesins in sister chromatid cohesion, formation of axial elements, and recombination during yeast meiosis. Cell 98: 91–103.
[26]
Laloraya S, Guacci V, Koshland D (2000) Chromosomal addresses of the cohesin component Mcd1p. J Cell Biol 151: 1047–1056.
[27]
Lee JT, Lu N, Han Y (1999) Genetic analysis of the mouse X inactivation center defines an 80-kb multifunction domain. Proc Natl Acad Sci U S A 96: 3836–3841.
[28]
Losada A, Hirano T (2001) Intermolecular DNA interactions stimulated by the cohesin complex in vitro. Implications for sister chromatid cohesion. Curr Biol 11: 268–272.
[29]
Losada A, Hirano M, Hirano T (1998) Identification of Xenopus SMC protein complexes required for sister chromatid cohesion. Genes Dev 12: 1986–1997.
[30]
Matsuzaki H, Nakajima R, Nishiyama J, Araki H, Oshima Y (1990) Chromosome engineering in by using a site-specific recombination system of a yeast plasmid. J Bacteriol 172: 610–618.
[31]
Megee PC, Koshland D (1999) A functional assay for centromere-associated sister chromatid cohesion. Science 285: 254–257.
[32]
Megee PC, Mistrot C, Guacci V, Koshland D (1999) The centromeric sister chromatid cohesion site directs Mcd1p binding to adjacent sequences. Mol Cell 4: 445–450.
[33]
Meluh PB, Koshland D (1997) Budding yeast centromere composition and assembly as revealed by in vivo cross-linking. Genes Dev 11: 3401–3412.
[34]
Michaelis C, Ciosk R, Nasmyth K (1997) Cohesins: Chromosomal proteins that prevent premature separation of sister chromatids. Cell 91: 35–45.
[35]
Mythreye K, Bloom KS (2003) Differential kinetochore protein requirements for establishment versus propagation of centromere activity in . J Cell Biol 160: 833–843.
[36]
Nicklas RB, Ward SC (1994) Elements of error correction in mitosis: Microtubule capture, release, and tension. J Cell Biol 126: 1241–1253.
[37]
Nonaka N, Kitajima T, Yokobayashi S, Xiao G, Yamamoto M, et al. (2002) Recruitment of cohesin to heterochromatic regions by Swi6/HP1 in fission yeast. Nat Cell Biol 4: 89–93.
[38]
Panizza S, Tanaka T, Hochwagen A, Eisenhaber F, Nasmyth K (2000) Pds5 cooperates with cohesin in maintaining sister chromatid cohesion. Curr Biol 10: 1557–1564.
[39]
Renauld H, Aparicio OM, Zierath PD, Billington BL, Chhablani SK, et al. (1993) Silent domains are assembled continuously from the telomere and are defined by promoter distance and strength, and by SIR3 dosage. Genes Dev 7: 1133–1145.
Saffery R, Irvine DV, Griffiths B, Kalitsis P, Wordeman L, et al. (2000) Human centromeres and neocentromeres show identical distribution patterns of >20 functionally important kinetochore-associated proteins. Hum Mol Genet 9: 175–185.
[42]
Saunders M, Fitzgerald-Hayes M, Bloom K (1988) Chromatin structure of altered yeast centromeres. Proc Natl Acad Sci U S A 85: 175–179.
[43]
Skibbens RV, Rieder CL, Salmon ED (1995) Kinetochore motility after severing between sister centromeres using laser microsurgery: Evidence that kinetochore directional instability and position is regulated by tension. J Cell Sci 108: 2537–2548.
[44]
Skibbens RV, Corson LB, Koshland D, Hieter P (1999) Ctf7p is essential for sister chromatid cohesion and links mitotic chromosome structure to the DNA replication machinery. Genes Dev 13: 307–319.
[45]
Sumner AT (1991) Scanning electron microscopy of mammalian chromosomes from prophase to telophase. Chromosoma 100: 410–418.
[46]
Tanaka T, Cosma MP, Wirth K, Nasmyth K (1999) Identification of cohesin association sites at centromeres and along chromosome arms. Cell 98: 847–858.
[47]
Tanaka T, Fuchs J, Loidl J, Nasmyth K (2000) Cohesin ensures bipolar attachment of microtubules to sister centromeres and resists their precocious separation. Nat Cell Biol 2: 492–499.
[48]
Tomonaga T, Nagao K, Kawasaki Y, Furuya K, Murakami A, et al. (2000) Characterization of fission yeast cohesin: Essential anaphase proteolysis of Rad21 phosphorylated in the S phase. Genes Dev 14: 2757–2570.
[49]
Toth A, Ciosk R, Uhlmann F, Galova M, Schleiffer A, et al. (1999) Yeast cohesin complex requires a conserved protein, Eco1p(Ctf7), to establish cohesion between sister chromatids during DNA replication. Genes Dev 13: 320–333.
[50]
Wang Z, Castano IB, De Las Penas A, Adams C, Christman MF (2000) Pol kappa: A DNA polymerase required for sister chromatid cohesion. Science 289: 774–779.
[51]
Watanabe Y, Nurse P (1999) Cohesin Rec8 is required for reductional chromosome segregation at meiosis. Nature 400: 461–464.
