Figure 2. GTF2IRD1 is a cold-inducible transcription factor that forms a complex with PRDM16 and EHMT1 in the BAT.
(A) Identification of nuclear-localized components in a PRDM16 transcriptional complex purified from differentiated beige adipocytes. Nuclear localized proteins and canonical DNA-binding transcription factors were listed based on the annotation database in UniProt.
(B) Gene expression profile of the identified transcriptional factors in interscapular BAT, inguinal WAT, and epididymal WAT. The color scale shows z-scored FPKM representing the mRNA level of each gene in blue (low expression)-white-red (high expression) scheme.
(C) Gene expression profile of the BAT-enriched transcriptional factors in cultured primary brown adipocytes and white adipocytes.
(D) Expression of Gtf2ird1 in BAT from mice housed at 22°C or 6°C for 3 days. n=5.
(E) Immunoblotting for GTF2IRD1 protein from mice in (D). β-actin was used as loading control.
(F) Expression of Gtf2ird1 mRNA in the indicated adipose tissues of mice treated with vehicle (saline) or CL316, 243 at a dose of 1 mg/kg for 7 days. n=5. * P<0.05, ** P<0.01, Data in D and F are represented as mean ± SEM.
(G) A EHMT1 complex was immunopurified from differentiated brown adipocytes. Endogenous GTF2IRD1 was detected by immunoblotting. Inputs are shown in lower panels.
(H) In vitro binding assay of 35S-labeled GTF2IRD1 and the indicated GST-fusion fragments of PRDM16. Coomassie brilliant blue for GST-proteins (bottom panel).