Fig. 6. INPP4B regulates lipogenesis and AKT and PKC signaling pathways in mouse liver.

a RNA from Fig. 4(c–i) were analyzed for Srebf1 expression by qRT-PCR using 18S as control. b Proteins were extracted from livers of LFD WT, LFD Inpp4b^−/−, HFD WT, and HFD Inpp4b^−/− mice and used to compare levels of unprocessed SREBP1, processed SREBP1, and tubulin by western blotting. c Quantification of processed SREBP1 levels were presented as an average of three independent samples. The protein levels were normalized to tubulin and were shown in fold change from LFD WT. d Protein lysates from (b) were analyzed for pAKT, total AKT, pPKCβII, pPKCζ, pPKCε, and tubulin using western blotting. e, f Quantification of pAKT and pPKCβII protein from (d) (N = 3). g–i The Inpp4b^−/− animals were treated with vehicle DMSO (N = 4), 6 mg/kg of AZD5363 (N = 2) or BIM-I (N = 4) for 16 h. Livers were dissected and used for protein and RNA purification. One-way ANOVA (Dunnett’s multiple comparison test) was used. (g) Fifty µg of protein extracted from livers of Inpp4b^−/− animals treated with vehicle, BIM-I, or AZD5363 were used to compare levels of unprocessed and processed forms of SREBP1 and tubulin. (h) Quantification of processed or unprocessed SREBP1 proteins in (g) (N = 4). i RNAs purified from livers of Inpp4b^−/− animals treated with vehicle, BIM-I, or AZD5363 were used to compare expression of SREBP1 target genes Mogat1, Cyp2b13, and Cyp2b9 by qRT-PCR using 18S as control. j WT or Inpp4b^−/− mice were fasted overnight. Protein lysates from fasted and fed mice were isolated and analyzed for pAKT, precursor and processed forms of SREBP1, and tubulin using western blotting. k, l Quantification of pAKT and processed SREBP1 protein levels from two independent experiments (N = 4 for each individual bar) (^*p < 0.05, ^**p < 0.01, ^***p < 0.001, ^****p < 0.0001. Exact p values are provided in the Source data).