Extended Data Fig. 4: SNHG6 regulates FAF2-mTOR interaction in hepatoma.
(A) Co-IP detection of the FAF2-mTOR interaction at cholesterol starvation. HepG2 cells expressing SFB-FAF2 and GFP-mTOR plasmids (48 h) were subjected to MCD (0.5% w/v) for 3 h; (B) Cholesterol mediated FAF2-mTOR interaction in a dose-dependent manner. HepG2 cells expressing SFB-FAF2 and GFP-mTOR plasmids (48 h) were replenished indicated cholesterol concentrations for 3 h; (C) Co-IP detection of the FAF2-mTOR interaction at arginine treatment. HepG2 cells expressing indicated plasmids were pre-cultured in amino acid deficiency medium for 3 h and were changed to arginine sufficiency medium for 1 h; (D) Co-IP detection of the FAF2-mTOR interaction at insulin treatment. HepG2 cells expressing indicated plasmids were pre-cultured in serum deficiency medium for 3 h and then were subjected to insulin stimulation for 1 h. (E) Co-IP detection of the FAF2-mTOR interaction upon glucose treatment. HepG2 cells expressing indicated plasmids, followed by glucose starvation for 8 h and then glucose replenishment for 8 h. (F,G) The regulatory function of SNHG6 on FAF2-mTOR interaction was detected by Co-IP assay. GFP-mTOR, SFB-FAF2 and SNHG6 plasmids (F) or SNHG6 interfering RNA (G) were transfected to HepG2 cells for 48 h as described conditions. (H) Co-IP detection of the FAF2-mTOR interaction upon RNase treatment. Indicated plasmids were transfected into HepG2 cells for 48 h, whose cell lysates were incubated with RNase treatment for 15 mins in vitro, followed S-protein beads immunoprecipitation and IB detection using indicated antibodies. (I) Co-IP detection FAF2-mTOR interaction in Huh7 cells. shScr/shSNHG6 Huh7 cells were transfected with SFB-FAF2 plasmids for 48 h, followed by cholesterol replenishment (50 μM, 2 h). (J) Coomassie blue staining of GST-tagged FAF2 truncation.
