MRC Clinical Sciences Centre Cell Cycle Group, Luis Aragón

Background
Following DNA replication in S-phase, chromosomes are linked together (cohesion), compacted (condensation), untangled and then moved to opposite poles of the cell (segregation). All these events are coordinated and executed with precision to prevent aneuploidy, a condition of inappropriate chromosome number often associated with birth defects and cancer.

Late anaphase cell [+}
(Click image to enlarge)

Wild type yeast cell in late anaphase. The cell expresses the nucleolar marker Net1 fused to the cyan fluorescent protein (CFP) and contain a tetO chromosome tag inserted in the telomere of chromosome XII (visialised through tetR-Green Fluorescent Protein, GFP). The nuclei is shown in red, the ribosomal DNA in blue and the telomere tag in green.

The poleward movement of chromosomes to the two daughter cells relies on the spindle, a complex microtubule-based machine, which captures sister chromatids during metaphase and pulls them to opposite poles at anaphase. Chromosomes are not passive substrates during segregation, but actively contribute to the process by virtue of three specific structural features: (i) Each sister chromatid has a region that mediates the attachment and movement of chromosomes on the microtubules of the spindle, named centromere, (ii) Sister chromatids are linked by protein bridges (sister chromatid cohesion). The resolution of cohesion not only serves as a key regulatory step for the onset of chromosome segregation but it is also important during bipolar attachment of sister chromatids to the mitotic spindle, and (iii) Sister chromatids are compacted during segregation to minimise the entanglement of chromosomes while they move to the poles in anaphase. This is necessary to ensure that mitotic chromosomes are less than half as long as the spindle, thus preventing the lagging ends of segregating chromosomes from being severed by cytokinesis.

Smc6-9 mutant cell [+]
(Click image to enlarge)

Smc6-9 mutant cell in anaphase. The cell expresses the nucleolar marker Net1 fused to the green fluorescent protein (GFP). The nuclei are shown in red and the ribosomal DNA in green. Note that during anaphase rDNA separates into two in wild type cells but as it can be seen in the image rDNA separates into three or more signals in the smc6-9 mutant. The reason for this is that chromosome XII breaks at the rDNA region during mitosis in this mutant.

In eukaryotes cohesion and condensation are largely dependent upon two multiprotein complexes named cohesin and condensin respectively. The core subunits of cohesin and condensin belong to the SMC protein family of chromosomal ATPases.

SMC protein family
SMC proteins form complexes that have essential roles in chromosome behaviour. The members of the family are long proteins with two globular domains separated by two long coiled-coils and a central globular hinge region. The signature domain is a C-terminal 'DA' box. Both N- and C-terminal domains contain candidate Walker B motifs (ATP hydrolysis). In the genome of S. cerevisiae there are three structurally distinct SMC containing protein complexes: (i) cohesin, (ii) condensin, and (iii) the “SMC5/6 complex”.

(i) Cohesin
Eukaryotic sister chromatids are held together by proteins from the time of their replication during S phase, until they are segregated at anaphase. Cohesion is mediated by cohesin, which contains a core of two SMC proteins, Smc1p and Smc3p and two further subunits, Scc3p and Scc1p (Mcd1p/Rad21p). In budding yeast cohesin holds chromatids together. Cohesion is resolved during the metaphase to anaphase transition through the concerted cleavage of the Scc1p subunit by a cysteine protease, named separase. This resolution allows the spindle-mediated poleward movement of chromatids to the poles. The resolution of cohesion is also linked to cell cycle control through the spindle checkpoint.

(ii) Condensin
Eukaryotic chromosome condensation is a form of chromatin organisation required to densely compact chromosomes during mitosis. Condensation is mediated by the action of the condensin complex, which in S. cerevisiae consists of Smc2p, Smc4p, Brn1p, Ycs4p and Ycs5p.

Rap1 and Smc6 [+]
(Click image to enlarge)

Partial colocalisation of telomere protein Rap1 (red) and Smc6 (green). Nuclei are shown in blue.

Condensin was originally identified in Xenopus laevis egg extracts, where it is required for mitotic chromosome condensation. The present view is that condensin accomplishes condensation in vivo by reconfiguring chromatin, a feature consistent with the presence of secondary structures resembling motor proteins in members of the SMC family.

Studies in yeast support a role for the condensin complex in chromosome compaction in this organism. However, it has been also shown that S. cerevisiae condensin plays additional roles in the transmission of the rDNA locus. Recent experiments in a variety of organisms suggests that condensin is likely to perform several unexpected roles in widely diverse processes, including regulation of gene expression, cell-cycle checkpoints and centromere organisation.

(iii) The 'SMC5/6 complex'
Eukaryotic genomes have a third conserved SMC-containing protein complex: the 'SMC5/6'. The S. pombe, rad18 gene and its S. cerevisiae homologue, SMC6 (RHC18), are essential for proliferation and sensitive to ionising irradiation and UV. These studies suggest putative roles in higher-order chromosome organisation and DNA damage. The requirement of the SMC5/6 complex for viability is predicted to originate from a possible role linking replication, repair and mitotic control. The complex is likely to consist of multiple subunits including Smc5p, Smc6p, Nse1p and Nse2p, Nse3p, Nse4p Nse5p and Nse6p.