Figure 7. MARCH1 regulates insulin sensitivity by controlling surface INSR expression.

(a) Immunoblot analysis of insulin-stimulated INSR, IRS-1 and AKT activation in HeLa cells expressing the indicated shRNAs. (b) Immunoblot analysis of insulin-stimulated INSR, IRS1 and AKT activation in HeLa cells expressing the indicated constructs. (c) Insulin dose–response curves in HeLa cells expressing NS or MARCH1 shRNA #1, constructed using AKT pSer⁴⁷³ immunoblot densitometry (see also Supplementary Fig. 6d). (d) Biotinylation assay for cell surface expression of INSRα and INSRβ in serum-starved HeLa cells expressing NS or MARCH1 shRNA #1, normalized to Na-K ATPase intensity (see also Supplementary Fig. 7a). (e) Flow cytometric measurement of surface INSRα in HeLa cells expressing the indicated shRNA. (f) Immunoblot analysis for cell surface expression of INSRα and INSRβ in GPMVs isolated from serum-starved HeLa cells expressing indicated shRNAs. Data are mean±s.e.m. In (d), n=3 biological replicates. In all panels, *P<0.05, comparisons by unpaired t-test.