Figure 6.

CMs and CDs in Core K compartments with E and J Heads

(A) CMs and CDs in exponentially growing strains containing cysteines in the hinge, E heads, or Smc1/Scc1/Smc3 interfaces. Strains with cysteine pairs at interfaces (4C or 6C) and strains lacking just one of the cysteines (3C or 5C), carrying a 2.3-kb circular minichromosome, were treated with BMOE. DNAs associated with cohesion immunoprecipitates (Scc1-PK) were denatured with SDS and separated by agarose gel electrophoresis. Southern blotting reveals supercoiled monomers and nicked and supercoiled catenanes along with two forms of DNA unique to 6C cells, termed CMs and CDs (see Gligoris et al., 2014 for details). Full circularisation efficiencies (mean ± SD of 3 independent experiments): ES = 14 ± 2% (as measured by in-gel fluorescence) and 15% (as measured by western blot after engineered cleavage of Smc3, Figure S2A), EK = 7 ± 1%, and SK RING = 13 ± 6%. Representative of at least 3 independent experiments. See also Figure S4A.

(B) CMs and CDs in exponentially growing strains containing cysteines in the hinge, J heads, or Smc1/Scc1/Smc3 interfaces. Full circularisation efficiencies (mean ± SD of 3 independent experiments): JS = 11 ± 4%, JK = 7 ± 2%, and SK RING = 13 ± 6%. Representative of at least 3 independent experiments. See also Figure S4B. The lane corresponding to hinge/J heads 3C control is not shown.

(C) CMs and CDs in exponentially growing strains containing cysteines in the hinge, coiled coils, or Smc1/Scc1/Smc3 interfaces. Full circularisation efficiencies (mean ± SD of 3 independent experiments): CS = 22 ± 2%, CK = 12 ± 2%, and SK RING = 13 ± 6%. Representative of at least 3 independent experiments.

(D) Schematic representation of DNA topological association with cohesin compartments (see discussion for details).

See also Figure S4.