Skip to main content
. 2022 Oct 14;13:6080. doi: 10.1038/s41467-022-33749-0

Fig. 5. Suppression of autophagy by dephosphorylation of AMPKα through transcriptional induction of Ppp2r2d by AhR.

Fig. 5

a qRT-PCR assays for isoforms of PP2A subunit B in the liver of mice fed with Lieber-DeCarli diets for 4 weeks (n = 5 each). b Immunoblot (upper) or representative immunohistochemical images (lower) for PPP2R2D in the same liver samples as in (a) (n = 5 each). Scale bar: 100 μm. c qRT-PCR (left) or immunoblot (right) assays for PPP2R2D in AML-12 cells (100 mM ethanol, 48 h; repeated three times with similar results). d–f qRT-PCR, immunoblot, and immunohistochemical analyses for PPP2R2D (Lieber-DeCarli diet for 5 weeks; n = 5 each). Representative images were shown for (f). Scale bar: 100 μm. g Immunoblot of mPHs isolated from the mice fed as in (d) (n = 3 each). h Immunoblots of PPP2R2D in mPHs (100 mM ethanol, 48 h) or AML-12 cells (siAhR transfection, 48 h; 100 mM ethanol, 24 h) (repeated three times with similar results). i qRT-PCR assays (left) or immunoblot (right) for PPP2R2D in AML-12 cells treated with 100 μM kynurenine for 12 h or the indicated times (repeated three times with similar results). j Immunoblots for PPP2R2D in mPHs (100 μM kynurenine, 12 h) or AML-12 cells (transfection with siAhR, 24 h; 100 μM kynurenine, 12 h) (repeated three times with similar results). k ChIP assays using AhR immunoprecipitates from AML-12 cells (100 mM ethanol for 24 h; IgG immunoprecipitation, a negative control). One-tenth of cross-linked lysates served as the input control (n = 3 each). l Ppp2r2d promoter-reporter assays using WT or Mut reporters in AML-12 cells transfected with siAhR for 24 h (n = 3 each). m Immunoblots for p-AMPKα in mPHs treated with 100 mM ethanol for 48 h (left) or with 100 μM kynurenine for 12 h (right) after transfection with siPPP2R2D for 24 h (repeated three times with similar results). n Immunoblots for autophagy markers using the same samples as in (m) (repeated three times with similar results). Values are expressed as means ± SEM. Significantly different compared to Con, WT, Veh, or siCon. Data were analyzed via two-tailed Student’s t-test (a, c, i, k, l) or one-way ANOVA with Tukey HSD (d). Source data are provided as a Source Data file.