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. 2016 Aug 31;7:12639. doi: 10.1038/ncomms12639

Figure 6. Insulin signalling represses MARCH1 transcription through FOXO1.

Figure 6

(a) qRT-PCR measurement of March1 expression in wild-type mouse WAT after 6 h fasting (basal) or after 2 h of insulin stimulation (clamp). n=8 (basal) and 9 (clamp) mice per group. (b) qRT-PCR measurements of MARCH1 expression (n=3) in the indicated cell lines after acute insulin stimulation at the indicated doses. (c) qRT-PCR measurement of FOXO1 mRNA expression (n=3) in HeLa cells expressing either a NS or FOXO1 shRNA. (d) qRT-PCR measurement of MARCH1 mRNA expression (n=3) in HeLa cells expressing either a NS or FOXO1 shRNA. (e) qRT-PCR measurements of MARCH1 expression (n=3) in HeLa (left) or HepG2 (right) cells expressing empty vector, wild-type FOXO1, constitutively active FOXO1 and/or PGC-1α and treated with insulin as indicated. (f) Luciferase reporter assay (n=3) for wild-type MARCH1 promoter (left) or a mutant MARCH1 promoter with a mutated FOXO1-binding site (right) in HeLa cells expressing empty vector, wild-type FOXO1, constitutively active FOXO1 (FOXO1 CA), and/or PGC-1α and treated with insulin as indicated. (g) ChIP measurement of FOXO1 protein enrichment on the MARCH1 or ACTIN promoter with and without insulin treatment (n=3) as indicated. Data are mean±s.e.m. In all panels, *P<0.05, comparisons by t-test. qRT-PCR, quantitative real-time PCR.