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. Author manuscript; available in PMC: 2021 Jul 14.
Published in final edited form as: Nature. 2020 Jun 24;584(7819):98–101. doi: 10.1038/s41586-020-2428-0

Figure 3: Regulation of β-catenin acetylation by glycolysis and pHi.

Figure 3:

(a) Western blot analysis of whole cell extracts prepared from hiPS cells differentiated to PSM in vitro in CL medium, using anti-acetylated K49 β-catenin, anti-active β-catenin, anti-actin and anti-β-catenin antibodies (n=3).

(b) Western blot analysis using anti-acetylated K49 β-catenin, anti-laminB, anti-GAPDH and anti-β-catenin antibodies of cytoplasmic (Cyt) and nuclear (Nucl) extracts prepared from hiPS cells differentiating to PSM in vitro in CL medium (n=3).

(c-d) Western blot analysis using anti-acetylated K49 β-catenin, anti-active β-catenin, anti-actin and anti-β-catenin antibodies. (c) Whole cell extracts of day 2 hiPS cells differentiated to PSM in vitro in CL medium and cultured at different pH for 3h (n=4). (d) Whole cell extracts of day 2 hiPS cells differentiated to PSM in CL medium in vitro and cultured for 6 h in 2DG and glucose-free (Glu(−)) CL medium (n=3).

(e) Immunoprecipitations with an anti β-catenin antibody of extracts of 2-day chicken embryos cultured at different pH. Western blot analysis using anti-acetylated lysine, anti-acetylated K49 β-catenin and anti-β-catenin antibodies. IgG: control immunoprecipitation. (n=3).

(f) Immunoprecipitations of extracts of 2-day chicken embryos cultured at different glucose concentrations with an anti β-catenin antibody. Western blot analysis using anti-acetylated lysine, anti-acetylated K49 β-catenin and anti-β-catenin antibodies. IgG: control immunoprecipitation. (n=4).

(g) Western blot analysis using anti-acetylated lysine antibody (top) and anti-β-catenin antibodies (bottom). Recombinant β-catenin protein was incubated with 500 μM or 10 mM acetyl-CoA sodium salt in PBS at different pH conditions for 3 h at 37C. (n=3).

(h) Western blot analysis using anti-acetylated lysine antibody (top) and anti-β-catenin antibodies (bottom). Recombinant β-catenin protein was incubated with 10 mM acetyl-CoA sodium salt in PBS at pH6.5~pH7.5 for 3 h at 37C. (n=3).

For gel source data, see Supplementary Figure 1