Fig. 4: ENDOG activates the mTORC2-AKT-ACLY axis by competitively binding with 14-3-3γ.

a Co-IP analyses. HepG2 cells were cotransfected with 14-3-3γ-Myc and ENDOG-Flag/ pLVX3-Flag plasmids for 48 h. b Co-IP analyses. HepG2 cells were transfected with ENDOG-Flag or pLVX3-Flag plasmids for 48 h. c Endogenous Co-IP analyses in wild-type and ENDOG knockout HepG2 cells. d, e ENDOG releases to the cytoplasm in HepG2 after being treated with 200 μM oleic acid for 24 h. Cyto, cytoplasm; Mito, mitochondria. n = 4 biologically independent samples. f Representative images of costaining of Tim23 (mitochondrial inner membrane protein) and ENDOG in HepG2 after being treated with 200 μM oleic acid for 24 h. g Endogenous Co-IP analyses in wild-type HepG2 cells after being treated with 200 μM oleic acid for 24 h. h, i Western blot and quantitative results of AKT-ACLY signaling after transfection with ENDOG and 14-3-3γ / ENDOG plasmids together for 48 h. n = 4 biologically independent samples. j–l Representative images of Nile red, the quantitative results of lipid area and measurement of triglycerides after transfection with ENDOG and 14-3-3γ / ENDOG plasmids together for 24 h and then treated with 200 μM oleic acid for another 24 h. n = 10 biologically independent samples for Nile red staining and 4 for triglyceride measurement. Three experiments were repeated independently with similar results in (a, b, c, f, g). Statistical significance was determined by unpaired Student’s t-test (two-tailed) in (i, k, l); error bars are mean ± SD. Source data and exact P values are provided in a Source data file. *P < 0.05; **P < 0.01; ***P < 0.001.