Kinetochores

Radial arrays are best exemplified by mitotic half-spindles, which have a central MTOC, the centrosome. The centrosome consists of two centrioles (which are homologous with the basal body) surrounded by dense pericentriolar material (Kalt and Schliwa, 1993). In plant cells, the MTOC of the mitotic spindle consists of dense material only without centrioles. The plus ends of microtubules of the mitotic spindle are directed toward the equator of the cell. Some are free, and others attach to kinetochores on chromatids (see Figure 4). 

During anaphase, the two sets of daughter chromosomes separate and move toward the poles (anaphase A), and the poles move away from each other with an accompanying three- to fourfold elongation of the pole-to-pole distance by polar fibers but a shortening of the kinetochore fibers in anaphase B. 

During telophase, a midbody forms between the separating daughter cells (see Figure 4), and disassembly of the remaining spindle microtubules and kinetochores takes place. 

In terms of evolutionary biology, the complex mitotic process of higher animals and plants has evolved through a progression of steps from simple prokaryotic fission sequences. In prokaryotic cells, the two copies of replicated chromosomes become attached to specialized regions of the cell membrane and are separated by the slow intrusion of the membrane between them. In many primitive eukaryotes, the nuclear membrane participates in a similar process and remains intact the spindle microtubules are extranuclear but may indent the nuclear membrane to form parallel channels. In yeasts and diatoms, the nuclear membrane also remains intact, an intranuclear polar spindle forms and attaches at each pole to the nuclear envelope, and a single kinetochore microtubule moves each chromosome to a pole. In the cells of higher animals and plants, the mitotic spindle starts to form outside of the nucleus, the nuclear envelope breaks down, and the spindle microtubules are captured by chromosomes (Kubai, 1975 Heath, 1980 Alberts et al., 1989). 

Ault, A. Rieder, C.L. (1994). Centrosome and kinetochore movement during mitosis. Curr. Opin. Cell Biol. 6,41-49. 

Gorbsky, G.J. Borisy, G.G. (1989). Microtubules of the kinetochore fiber turn over in metaphase but not in anaphase. J. Cell Biol. 109, 653-662. 

Mitchison, T.J. (1988). Microtubule dynamics and kinetochore function in mitosis. Ann. Rev. Cell Biol. 4, 527-549. 

Burton PBJ, Raff MC, Kerr P, Yacoub MH, Barton PJR 1999 An intrinsic timer that controls cell-cycle withdrawal in cultured cardiac myocytes. Dev Biol 216 659—670 Chen X, Ko LJ, Jayaraman L, Prives C 1996 p53 levels, functional domains, and DNA damage determine the extent of the apoptotic response of tumor cells. Genes Dev 10 2438—2451 Duesbery NS, Choi T, Brown KD et al 1997 CENP-E is an essential kinetochore motor in maturing oocytes and is masked during mos-dependent cell cycle arrest at metaphase II. Proc Natl Acad Sci USA 94 9165-9170... 

Zecevic M, Catling AD, Eblen ST et al 1998 Active MAP kinase in mitosis localization at kinetochores and association with the motor protein CENP-E. J Cell Biol 142 1547—1558... 

Mailer Our hypothesis is that CSF is at the kinetochore. George Vande Woude published some data showing that the kinetochore protein CENP-E has certain masked epitopes only in a CSF-arrested spindle and not in a mitotic spindle. 

Mailer What might change that story are the phospho-specific antibodies for Rsk and MAPK, because they can pick up changes of just a few percent. The CSF on kinetochores may require only a tiny pool of those enzymes for regulation. 

Loss of sister chromatid cohesion would therefore be sufficient for the sudden movement of chromatids to opposite poles at the metaphase to anaphase transition. According to this hypothesis, a specific apparatus binds chromatids together during replication, holds them in an orientation that facilitates the attachment of sister kinetochores to spindles extending to opposite poles, and resists the splitting force that results from this bipolar attachment to the spindle. Destruction of this specialized cohesive structure triggers movement of chromatids to opposite poles at the onset of anaphase. 

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... 

Khodjakov A, ColeRW, Bajer AS,Rieder CL 1996 The force for poleward chromosome motion in Haemanthus cells acts along the length of the chromosome during metaphase but only at the kinetochore during anaphase. J Cell Biol 132 1093—1104... 

Lehner There is a paper by Goldstein (1981) in which he describes electron microscope (EM) studies of meiotic chromosomes. At meiosis I, chromosomes do not have two sister kinetochores. There is a maturation into two kinetochores between the two meiotic divisions. Have comparable EM analyses been done in yeast ... 

Goldstein LS 1981 Kinetochore structure and its role in chromosome orientation during the first meiotic division in male D. melanogaster. Cell 25 591-602... 

Blower MD, Karpen GH (2001) The role of Drosophila CID in kinetochore formation, cell-cycle progression and heterochromatin interactions. Nat Cell Biol 3 730-739 Blower MD, Sullivan BA, Karpen GH (2002) Conserved organization of centromeric chromatin in flies and humans. Dev Cell 2 319-330... 

Heun P, Erhardt S, Blower MD, Weiss S, Skora AD, Karpen GH (2006) Mislocalization of the Drosophila centromere-specific histone CiD promotes formation of functional ectopic kinetochores. Dev Cell 10 303-315... 

Oegema K, Desai A, Rybina S, Kirkham M, Hyman AA (2001) Functional analysis of kinetochore assembly in Caenorhabditis elegans. J Cell Biol 153 1209-1226 Okada M, Cheeseman IM, Hori T, Okawa K, McLeod IX, Yates JR, Desai A, Fukagawa T (2006) The CENP-H-I complex is required for the efficient incorporation of newly synthesized CENP-A into centromeres. Nat Cell Biol... 

Van Hooser AA, Ouspenski 11, Gregson HC, Starr DA, Yen TJ, Goldberg ML, Yokomori K, Earnshaw WC, Sullivan KE, Brinkley BR (2001) Specification of kinetochore-forming chromatin by the histone H3 variant CENP-A. J Cell Sci 114 3529-3542... 

Cse4 CID CENP-A Centromer identifier Kinetochore assembly RD/RI... 

Centromer-specific H3 variants (CenH3 s) are known as CENP-A in mammals, Cid in flies, Cse4 in yeast (Fig. 1 Table 1 reviewed in Smith 2002). CenH3 s are incorporated into nucleosomes independently of centromeric DNA replication. CenH3-containing nucleosomes associate with a number of factors with important roles in centromere structure and kinetochore assembly. Recently, the deposition complex... 

Centromere protein A (CENP-A), one of several variants of histone H3, is phosphorylated on Ser 7 by Aurora B kinase which is equivalent to Ser 10 of histone H3 (Zeitlin et al, 2001). Recent studies demonstrate that Aurora A kinase also phosphorylates CENP-A (S7) (Kunitoku et al, 2003) (Table 1). The presence of CENP-A in centromeric nucleosomes is required for kinetochore organization and function (Choo 2001). Loss of CENP-A phosphorylation function at Ser 7 caused a mislocalisation of Aurora B, a putative partner phosphatase (PPl-yl) and inner centromere protein (INCENP). H3.3, another variant of histone H3 is phosphorylated on Ser 31 in vivo (Table 1). H3.3 (S31) is a mitosis-specific modification that is present only in late prometaphase and metaphase. Furthermore, H3.3 (S31) is excluded from centromeres. However it is enriched in distinct chromosomal areas immediately adjacent to centromeres (Hake et al, 2005). 

Adams RR, Maiato H, Earnshaw WC, Carmena M (2001) Essential roles of Drosophila inner centromere protein (INCENP) and aurora B in histone H3 phosphorylation, metaphase chromosome ahgnment, kinetochore disjunction, chromosome segregation. J Cell Biol 153(4) 865-880 Ahn SH, Cheung WL, Hsu JY, Diaz RL, Smith MM, Alhs CD (2005a) Sterile 20 kinase phospho-rylates histone H2B at serine 10 during hydrogen peroxide-induced apoptosis in S. cerevisiae. Cell 120(l) 25-36... 

Kunitoku N, Sasayama T, Marumoto T, Zhang D, Honda S, Kobayashi O, Hatakeyama K, Ushio Y, Saya H, Hirota T (2003) CENP-A phosphorylation by Aurora-A in prophase is required for enrichment of Aurora-B at inner centromeres and for kinetochore function. Dev Cell 5(6) 853-864... 

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