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. 2016 Feb 22;35(6):635–653. doi: 10.15252/embj.201592532

Figure 2. SMC3 acetylation site mutants generate stable cohesin–chromatin interactions without ESCO1 and ESCO2.

Figure 2

  1. Quantification of iFRAP experiments using HeLa cells expressing SMC3‐LAP in the wild‐type form (wt) or with lysines 105‐106 mutated to glutamines (QQ) or arginines (RR) and synchronized in G1‐phase. Error bars denote s.e.m., n > 25 cells per condition.
  2. Quantification of chromatin residence time of SMC3‐LAP alleles in G1‐phase. Error bars denote s.e.m.
  3. Quantification of iFRAP experiments using cells synchronized in G2‐phase. Error bars denote s.e.m., n > 26 cells per condition.
  4. Quantification of stably bound SMC3‐LAP alleles in G2‐phase. Error bars denote s.e.m.
  5. Western blot of mouse fibroblasts expressing SMC3‐LAP in the presence or absence of endogenous Smc3 after gene deletion. Extracts were prepared after subculturing two times. Cells without SMC3‐LAP do not proliferate after deletion of Smc3.
  6. Western blot showing HeLa cell extracts after treatment with siRNA as indicated and synchronized in G2‐phase. Note that ESCO1 immunoblot signals were also reduced in sororin‐depleted cells. We currently do not know if this is an effect of sororin depletion, an off‐target effect or an artifact of unequal sample loading.
  7. iFRAP quantification of stably chromatin‐bound SMC3‐LAP after siRNA treatment using cells synchronized in G2‐phase. Error bars denote s.e.m., n > 11 cells per condition. Unpaired t‐test was used to compare conditions.