Ep709y

Western blots were performed as previously described in ref. ^{22 (link)}. The following antibodies were used: rabbit mAb Anti-C/EBPα (EP709Y, Abcam), rabbit mAb Anti-C/EBPα (EP708Y, Abcam), rabbit pAb Anti-GAPDH (sc-25778, Santa Cruz Biotech.), rabbit mAb Anti-GAPDH (14C10, CST). Donkey-anti-rabbit-HRP (NA934V, GE-Healthcare) or goat-anti-rabbit (sc-2004, Santa Cruz Biotech.) served as secondary antibodies. For chemiluminescence detection, we used Western Blot Ultra or Western Blot Premium Substrate (Licor) and analyzed the membranes using C-Digit Chemiluminescent Western Blot Scanner (Licor). Evaluation of data were performed using ImageJ software (NIH). Full western blot gel scans are included as Supplementary Figs. 5–8.

Protein was obtained by collecting 3T3-L1 cells or WAT in lysis buffer, containing 150 mM NaCl, 50 mM Tris, 1% sodium deoxycholate, 0.1% SDS, and 1% Triton X-100 as well as fresh protease inhibitor PMSF. Protein was run by gel electrophoresis on SDS-PAGE, followed by transferring to PVDF membrane (Roche). Membranes were sealed with 5% BSA/TBST for 2 h, followed by incubation with primary antibodies at 4 °C overnight. The membranes were washed by phosphate buffer saline (PBS) three times and 5 min each time before incubated with secondary antibodies. Protein bands were captured by the GelView 6000 Pro (BLT, Guangzhou, China). Primary antibodies used were shown as following: anti-HMGB2 (EPR6301, Abcam), anti-C/EBPβ (E299, Abcam), anti-C/EBPα antibody (EP709Y, Abcam), anti-PPARγ antibody (EPR18516, Abcam), anti-FABP4 antibody (EPR3579, Abcam), anti-GAPDH antibody (Loading Control, Abcam), and anti-β-Tubulin antibody (EPR16778, Abcam).