Chromosome segregation, transcriptional regulation, and repair of DNA double-strand breaks require the cohesin protein complex. Cohesin holds the replicated chromosomes (sister chromatids) together to mediate sister chromatid cohesion. The mechanism of how cohesion is established is unknown. We found that in budding yeast, the head domain of the Smc3p subunit of cohesin is acetylated by the Eco1p acetyltransferase at two evolutionarily conserved residues, promoting the chromatin-bound cohesin to tether sister chromatids. Smc3p acetylation is induced in S phase after the chromatin loading of cohesin and is suppressed in G(1) and G(2)/M. Smc3 head acetylation and its cell cycle regulation provide important insights into the biology and mechanism of cohesion establishment.

In eukaryotes, the process of sister chromatid cohesion holds the two sister chromatids (the replicated chromosomes) together from DNA replication to the onset of chromosome segregation. Cohesion is mediated by cohesin, a four-subunit SMC (structural maintenance of chromosome) complex. Cohesin and cohesion are required for proper chromosome segregation, DNA repair, and gene expression. To carry out these functions, cohesion is regulated by elaborate mechanisms involving a growing list of cohesin auxiliary factors. These factors control the timing and position of cohesin binding to chromatin, activate chromatin-bound cohesin to become cohesive, and orchestrate the orderly dissolution of cohesion. The 45-nm ringlike architecture of soluble cohesin is compatible with dramatically different mechanisms for both chromatin binding and cohesion generation. Solving the mechanism of cohesion and its complex regulation presents significant challenges but offers the potential to provide important insights into higher-order chromosome organization and chromosome biology.

Vertebrate cells possess two different condensin complexes, known as condensin I and condensin II, that play a fundamental role in chromosome assembly and segregation during mitosis. Each complex contains a pair of structural maintenance of chromosomes (SMC) ATPases, a kleisin subunit and two HEAT-repeat subunits. Here we use recombinant human condensin subunits to determine their geometry within each complex. We show that both condensin I and condensin II have a pseudo-symmetrical structure, in which the N-terminal half of kleisin links the first HEAT subunit to SMC2, whereas its C-terminal half links the second HEAT subunit to SMC4. No direct interactions are detectable between the SMC dimer and the HEAT subunits, indicating that the kleisin subunit acts as the linchpin in holocomplex assembly. ATP has little, if any, effects on the assembly and integrity of condensin. Cleavage pattern of SMC2 by limited proteolysis is changed upon its binding to ATP or DNA. Our results shed new light on the architecture and dynamics of this highly elaborate machinery designed for chromosome assembly.

The Structural Maintenance of Chromosome (SMC) complex, termed cohesin, is essential for sister chromatid cohesion. Cohesin is also important for chromosome condensation, DNA repair, and gene expression. Cohesin is comprised of Scc3, Mcd1, Smc1, and Smc3. Scc3 also binds Pds5 and Wpl1, cohesin-associated proteins that regulate cohesin function, and to the Scc2/4 cohesin loader. We mutagenized SCC3 to elucidate its role in cohesin function. A 5 amino acid insertion after Scc3 residue I358, or a missense mutation of residue D373 in the adjacent stromalin conservative domain (SCD) induce inviability and defects in both cohesion and cohesin binding to chromosomes. The I358 and D373 mutants abrogate Scc3 binding to Mcd1. These results define an Scc3 region extending from I358 through the SCD required for binding Mcd1, cohesin localization to chromosomes and cohesion. Scc3 binding to the cohesin loader, Pds5 and Wpl1 are unaffected in I358 mutant and the loader still binds the cohesin core trimer (Mcd1, Smc1 and Smc3). Thus, Scc3 plays a critical role in cohesin binding to chromosomes and cohesion at a step distinct from loader binding to the cohesin trimer. We show that residues Y371 and K372 within the SCD are critical for viability and chromosome condensation but dispensable for cohesion. However, scc3 Y371A and scc3 K372A bind normally to Mcd1. These alleles also provide evidence that Scc3 has distinct mechanisms of cohesin loading to different loci. The cohesion-competence, condensation-incompetence of Y371 and K372 mutants suggests that cohesin has at least one activity required specifically for condensation.