C57BL/6J mice overexpressing IFI202b, herein referred to as B6-Tg(Ifi202b) mice, were generated by Ozgene (Perth, WA, Australia). c-Myc-tagged Ifi202b cDNA downstream of the human ubiquitin C promoter was integrated into the ROSA26 locus. C57BL/6J wild-type (B6-wt) littermates were used as control animals. NZO mice were taken from our own colony (NZO/HIBomDife, Nuthetal, Germany). For details of housing and diet conditions, see electronic supplementary materials (ESM) Methods.
Characterisation of mice
Blood glucose, plasma variables (insulin, NEFA and triacylglycerol levels), glucose tolerance and body composition were assessed as described previously . Plasma adiponectin and leptin levels were measured using the Mouse Adiponectin/Acrp30 and Mouse/Rat Leptin ELISA (DY1119 and MOB00, respectively; R&D Systems, Minneapolis, MN, USA). Histological analysis of WAT, BAT and liver tissues was carried out using H&E staining (see ESM Methods for further details).
Intraperitoneal ITT and insulin sensitivity
For the i.p. ITT, 12- and 20-week-old non-fasted mice were injected with insulin (0.75 IU/kg body weight; Actrapid Penfill; Novo Nordisk, Mainz, Germany) and blood glucose levels were measured at various time points. To examine hepatic and muscle insulin sensitivity, after a 6 h fast, mice were injected with NaCl (control) or insulin (0.75 IU/kg body weight). Twenty minutes later, mice were killed by cervical dislocation under isoflurane anaesthesia and samples were taken for western blot analysis.
Ex vivo lipolysis assay
Gonadal WAT (WATgon) explants from 8-week-old mice were stimulated with 10 μmol/l isoprenaline (known as isoproterenol in the USA) (Sigma-Aldrich, St Louis, MO, USA) or isoprenaline plus insulin (178 μmol/l) in 160 μl of high-glucose DMEM (PAN-Biotech, Aidenbach, Germany). After 2 h, release of NEFA was detected (NEFA-HR(2); Wako Chemicals, Neuss, Germany).
Cell culture and differentiation
For details of murine 3T3-L1 and human SGBS cell culture and differentiation conditions, see ESM Methods. WT-1 brown pre-adipocytes  tested negative for mycoplasma and cultivated and differentiated as described , with modified dexamethasone concentration (5 μmol/l).
Overexpression of IFI202b and analysis of lipid droplet formation in 3T3-L1 pre-adipocytes
3T3-L1 pre-adipocytes were infected with a c-Myc-Ifi202b encoding adenovirus (ViraQuest, North Liberty, IA, USA). Empty adenovirus was used as a control. Lipid droplet formation in IFI202b-overexpressing 3T3-L1 adipocytes was assessed by fluorescent staining. A detailed protocol for IFI202b overexpression in pre-adipocytes can be found in ESM Methods.
siRNA-mediated knockdown of IFI202B/IFI16 in pre-adipocytes
Expression of IFI202b in murine 3T3-L1 and WT-1 pre-adipocytes, as well as IFI16 in human pre-adipocytes was suppressed via electroporation using specific siRNA (IFI16: J-020004-05-0050; Dharmacon, Lafayette, CO, USA) as described previously .
Isolation and differentiation of primary adipogenic precursor cells
Adipogenic precursor cells were isolated from subcutaneous WAT (WATsc) of 8-week-old mice fed a standard diet (V153x R/M-H; ssniff, Soest, Germany), as described . Stem cell antigen 1 (SCA1)-positive cells [22, 23] were purified by FACS (BD FACSAria III; BD Biosciences, San Jose, CA, USA) and differentiated, as previously described , with minor modifications.
RNA extraction and expression analysis
RNA from cells was isolated and reverse transcribed for quantitative real-time PCR analyses. Expression levels of Ifi202b, Zfp423, Pparg, Cebpa, Adipoq, Glut4 (also known as Slc2a4), Fabp4, Plin1, Hsd11b1, Pref1 (Dlk1), Ucp1, Cidea, Prdm16, Tcf21, Tle3, Ebf2, Ppargc1a, Ppara, Il4, Il6, Il10, Tnfa (Tnf) and Ccl2 were analysed in murine samples. In human samples, IFI16, ZNF423, CEBPA, PPARG, ADIPOQ, FASN, GLUT4 (SLC2A4), PLIN1 and IL6 were analysed. Genes were detected using specific TaqMan Gene Expression Assays and the 7500 Fast Real-Time PCR System (Thermo Fisher Scientific, Waltham, MA, USA) and gene expression profiling of WATgon of 8-week-old mice was performed by OakLabs (Henningsdorf, Germany). For details see ESM Methods.
Protein extraction and western blotting
Liver, muscle, BAT and WAT tissues from mice were homogenised and analysed by western blotting . A monoclonal anti-IFI202b antibody was raised against the C-terminal peptide 1–21: C-MSNRNLRSSTNSEFSEGQHQ. Primary antibodies against c-Myc (1:500 dilution; Clontech, Saint-Germain-en-Laye, France), uncoupling protein 1 (UCP1; 1:1000; ab10983; Abcam, Cambridge, UK), total Akt (t-Akt; 1:1000; no. 9272; Cell Signaling, Danvers, CO, USA), p-Akt (Ser473; 1:1000; no. 9271; Cell Signaling), HDAC1 (1:25,000; ab7028; Abcam) and GAPDH (1:20,000; AM4300; Thermo Fisher Scientific), and appropriate horseradish peroxidase-labelled secondary antibodies (Dianova, Hamburg, Germany) were applied. Experimental controls were used to validate antibodies.
IFI16 expression in human adipose tissue
Previously, IFI16 expression in adipose tissue was measured in 166 obese individuals . Here, we compared those exhibiting the 10% highest levels of IFI16 with those with the 10% lowest levels of expression. Human phenotyping , IFI16 expression analyses  and basal and insulin-stimulated glucose uptake into isolated adipocytes were measured as previously described . All study protocols were approved by the ethics committee of the University of Leipzig (Leipzig, Germany). All participants gave written informed consent before taking part in the study.
Tagging SNP analysis in humans
From the ongoing Tübingen Family (TÜF) study for type 2 diabetes, healthy adults at increased risk for type 2 diabetes were recruited . In the present study, a subset of 1896 participants with complete leukocyte and CRP measurements and a second subset of 420 subjects with complete IL6 measurements were selected for genotyping. Tagging SNPs covering the common genetic variation were identified and genotyping was conducted, as detailed in ESM Methods. Association analyses were performed by multiple linear regression analysis (least squares method) to adjust for the confounding variables sex, age and BMI. The study adhered to the Declaration of Helsinki, and its protocol was approved by the local ethics boards (Ethics Committees of the Eberhard Karls University Tübingen, Tübingen, Germany and the Karolinska Institute, Stockholm, Sweden). Informed written consent was obtained from all participants.
All data are reported as mean ± SEM. Statistical analysis was performed using Student’s t test for single comparisons and one-way ANOVA with Tukey’s post hoc test for differences between more than two groups. Differences between two groups over time were evaluated using a two-way ANOVA with Bonferroni’s or Sidak’s correction for multiple comparisons (GraphPad Prism version 6.00; GraphPad, La Jolla, CA, USA). For SNP analysis, JMP 11.2.0 (from SAS, Cary, NC, USA) was used for statistical analysis. Significance was accepted at p < 0.05, p < 0.01 and p < 0.001. Samples were randomised, and no data were omitted. The experimenters were not blind to group assignment.