Cohesin and its friends

Genetics is the method of last resort when other approaches reach their limits. The identification of mutants such as desynaptic in maize (Maguire et al 1991) and MeiS332 in Drosophila (Davis 1971, Goldstein 1980), in which sister chromatids dissociate prematurely during meiosis, provided the first inkling that sister chromatid cohesion might be mediated by special proteins (Kerrebrock et al 1995). Despite its important role during meiosis, MeiS332 is dispensable for mitotic divisions and is therefore unlikely to be a universal component of the cohesion apparatus.

Genetic studies in yeast have meanwhile uncovered a multi-subunit complex called cohesin that is essential for sister chromatid cohesion not only in yeast (Toth et al 1999) but also in vertebrates (Losada et al 1998). An important breakthrough in cohesin's identification was the discovery that proteolysis (Holloway et al 1993), mediated by a ubiquitin protein ligase responsible for destroying mitotic cyclins (Irniger et al 1995, King et al 1995), is needed for sister chromatid separation. This ligase, known as the anaphase-promoting complex (APC) or cyclosome (Zachariae & Nasmyth 1999), was initially thought to mediate proteolysis of cohesion proteins. Its role in sister separation turns out to be less direct; it in fact mediates destruction of an inhibitor of the sister-separating apparatus (Yamamoto etal 1996a, Cohen-Fix etal 1996, Funabiki etal 1996a, Ciosk et al 1998). Nevertheless, the premise that the APC destroyed cohesion proteins provided a new impetus to the search for proteinaceous bridges connecting sister chromatids. Screens for mutations that permitted separation of sister chromatids in

FIG. 4. Cohesin and friends. (A), a model illustrating how yeast proteins required for cohesion connect sister chromatids during DNA replication and maintain this association until the onset of anaphase. (B), electron micrographs of homodimers of the Bacillus subtilis SMC protein. Three commonly observed conformations are shown. Size bar, 20 nm. Reprinted with permission from Melby et al (1998). (C) A speculative model for how cohesin might join sister chromatids together, which is based on the premise that cohesin forms large supercoiled loops analogous to those proposed for condensin (shown alongside).

FIG. 4. Cohesin and friends. (A), a model illustrating how yeast proteins required for cohesion connect sister chromatids during DNA replication and maintain this association until the onset of anaphase. (B), electron micrographs of homodimers of the Bacillus subtilis SMC protein. Three commonly observed conformations are shown. Size bar, 20 nm. Reprinted with permission from Melby et al (1998). (C) A speculative model for how cohesin might join sister chromatids together, which is based on the premise that cohesin forms large supercoiled loops analogous to those proposed for condensin (shown alongside).

cells lacking APC activity have now identified at least eight proteins essential for sister chromatid cohesion (Toth et al 1999, Michaelis et al 1997, Guacci et al 1997, Furuya et al 1998, Skibbens et al 1999). Remarkably, the function of all these proteins seem to be intimately connected (Fig. 4A).

Four of these proteins, Smc1, Smc3, Scc1 (also called Mcd1 and Rad21) and Scc3 form a multi-subunit complex called cohesin (Toth et al 1999, Losada et al 1998). Indeed, Mcd1/Scc1 was independently isolated as a dosage suppressor of an Smc1 mutation (Guacci et al 1997). All four cohesin subunits are required both for establishing cohesion during S phase and (at least in yeast) for maintaining it until the onset of anaphase. Two other proteins, Scc2 (Mis4) and Scc4, form a separate complex that is required for the association of cohesin with chromosomes (Ciosk et al 2000). Cohesin binds to specific chromosomal loci (including centromeres) for much of interphase (Tanaka et al 1999, Blat & Kleckner 1999, Megee et al 1999) but it can only establish cohesion between sister chromatids during DNA replication, possibly when sister DNA molecules emerge from replication forks (Uhlmann & Nasmyth 1998). Establishment of sister cohesion is therefore an integral part of S phase.

Another protein, Spo76 is required for orderly sister chromatid cohesion in Sordaria (van Heemst et al 1999). Spo76 has homologues in many organisms, and is called Pds5 in budding yeast (V. Guacci & D. Koshland, personal communication) and BimD in Aspergillus nidulans (Denison et al 1993). In budding yeast a protein called Eco1 or Ctf7 is essential for establishing cohesion during S phase but not for maintaining it during G2 or M phases (Toth et al 1999, Skibbens et al 1999). Its fission yeast homologue Eso1 is also required for establishing sister chromatid cohesion (Tanaka et al 2000). Of all known cohesion proteins the cohesin complex may lie at the heart of the cohesion process because cleavage of one ofits subunits is essential for the separation ofsister chromatids, at least in yeast (Uhlmann et al 1999). Xenopus cohesin is also needed for proper sister chromatid cohesion (Losada et al 1998). Nevertheless, it is still uncertain how, or indeed if, cohesin holds sisters together during metaphase in animal cells, as most of it dissociates from chromosomes by pro-metaphase (Losada et al 1998). It is therefore possible that other important players remain to be identified.

Two cohesin subunits, Smc1 and Smc3, are members of a large family of related proteins whose evolution predates the split between eukaryotes and bacteria (Hirano 1999). All Smc proteins have related globular domains at their N- and C-termini, joined by two long stretches of a-helical coiled-coil, which are linked by a central flexible hinge. Bacterial Smc proteins form anti-parallel homodimers whose terminal globular domains are proposed to form an active ATPase. The flexibility of the hinge region allows the Smc homodimer to adopt either a V or a linear shape (Fig. 4B) (Melby et al 1998). It remains to be seen whether cohesin contains an Smc1/3 heterodimer or Smc1 and Smc3 homodimers.

Little is known about the properties of cohesin in vitro, except that fragments from the C-terminal domain of Smc3p and its coiled-coil region can bind DNA (Akhmedov et al 1999). Smcl and Smc3 belong to a subfamily of eukaryotic Smc proteins, which includes Smc2 and Smc4. The latter two proteins are components of the condensin complex, which is necessary for mitotic chromosome condensation (Hirano & Mitchison 1994, Saka et al 1994, Hirano et al 1997). Condensin possesses ATPase activity and is capable of forming large supercoiled loops by introducing a global positive writhe (Kimura et al 1999). These positive supercoils might be the driving force for mitotic chromosome condensation. The presence of a pair of Smc proteins in both condensin and cohesin suggests that these two complexes might have similar though not identical activities. Cohesin might, for example, introduce large constrained supercoils, like those produced by condensin, at equivalent positions on each sister chromatid. Cohesin's Scc1 subunit might help link together equivalent coils from each sister (Fig. 4C). An ability to coil chromosomes would explain how cohesin contributes to chromosome compaction (Guacci et al 1997).

In animal cells, condensin binds to chromosomes at about the same time that most cohesin dissociates from them (Losada et al 1998), between prophase and pro-metaphase. It is possible that condensin's ability to condense chromosomes as cells enter mitosis depends on the prior dissociation of most cohesin. The connections between sister DNA molecules might otherwise interfere with the locally processive coiling of each chromatid on itself. Cohesin could also contribute to chromosome compaction during interphase and early stages of mitosis by providing longditudinal links along chromatids as well as horizontal ones between sisters (Guacci et al 1997).

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