Journal: Molecular Metabolism

Article Title: Baricitinib counteracts metaflammation, thus protecting against diet-induced metabolic abnormalities in mice

doi: 10.1016/j.molmet.2020.101009

Figure Lengend Snippet: Baricitinib attenuates the HD-induced JAK-STAT pathway. Western blotting analysis for phosphorylation of Tyr 1007/1008 JAK1/2 in the skeletal muscle (A) and kidneys (B) and normalized to total JAK1/2 and for phosphorylation of Tyr 690 on STAT2 in the skeletal muscle (C) and kidneys (D) and normalized to total STAT2. All of the data are expressed as mean ± SEM for n = 6 per group. ∗p< 0.05 vs ND and •p< 0.05 vs HD.

Article Snippet: The antibodies used were rabbit anti-Tyr 1007/1008 JAK2 (#3776), rabbit anti-total JAK2 (#3230), rabbit anti-Tyr 690 STAT2 (Bioss Antibodies, bs-3428R), rabbit anti-total STAT2 (#72604), rabbit p21 (#2947), rabbit anti-Ser 307 IRS-1 (#2381), mouse anti-total IRS-1 (#3194), rabbit anti-Ser 473 AKT (#4060), rabbit anti-total AKT (#9272), rabbit anti-Ser 9 GSK-3β (#9332) and rabbit anti-total GSK-3β (9315).

Techniques: Western Blot

Journal: The EMBO Journal

Article Title: IFNβ-dependent increases in STAT1, STAT2, and IRF9 mediate resistance to viruses and DNA damage

doi: 10.1038/emboj.2013.203

Figure Lengend Snippet: High levels of STAT1, STAT2, and IRF9 proteins protect cells from various RNA viruses in an IFN-independent manner. hTERT-HME1 cells transfected with empty vector (Vec), wild-type STAT1/STAT2/IRF9 (WT) or Y701F-STAT1/STAT2/IRF9 (YF) were used. ( A ) Cells were infected with 0.1 MOI of VSV or 1 MOI of EMCV, and the infected cells and cell-culture media were collected after 10 h (VSV) or 6 h (EMCV). The infectious viral titres in the collected samples were analysed by plaque assays on Vero cells. The data are represented as means of triplicate infections±s.d. An asterisk (*) represents P <0.01, by two-tailed t -test, compared to cells transfected with empty vector (Vec). ( B ) Cells were infected with VSV or EMCV (10–10 −5 MOI). After 48 h, the surviving cells were fixed with methanol (VSV) or 4% paraformaldehyde (EMCV) and stained with crystal violet. ( C ) Cells were infected with recombinant viral constructs (VSV, PIV3, or YFV) expressing GFP. After 8 h (VSV) or 48 h (PIV3 or YFV), GFP fluorescence was monitored by FACS analyses. The data are represented as mean GFP intensities (MFI)±s.d. of triplicate infections. An asterisk (*) represents P <0.01 and a cross (+) represents P <0.05, by two-tailed t -test, compared to Vec cells.

Article Snippet: Mouse monoclonal antibodies against STAT1 (BD Transduction), STAT2, IRF9 (ISGF3γ), and IRF1 (Santa Cruz Biotechnology), and rabbit polyclonal antibodies against Tyr 701-phosphorylated STAT1, Tyr 690-phophorylated STAT2 (Cell Signaling), and STAT1 (Upstate) were used for western analyses.

Techniques: Transfection, Plasmid Preparation, Infection, Cell Culture, Two Tailed Test, Staining, Recombinant, Construct, Expressing, Fluorescence

Journal: The EMBO Journal

Article Title: IFNβ-dependent increases in STAT1, STAT2, and IRF9 mediate resistance to viruses and DNA damage

doi: 10.1038/emboj.2013.203

Figure Lengend Snippet: U-ISGF3-induced genes have distinct ISREs. ( A ) BJ cells were treated with 3 IU/ml of IFNβ for 6 h, followed by microarray analyses for the expression of IFNβ-induced genes. To identify the U-ISGF3-induced genes, the cells were stably transfected with a lentiviral vector encoding Y701F-STAT1, without IFN treatment. The analysis criteria are described in ‘Materials and methods'. The genes marked with asterisks (*) are those with known anti-viral functions, and those marked with crosses (+) are known to be upregulated in DNA damage-resistant cancer cells. DDX58 is also known as RIG-I , and IFIH1 is known as MDA5. DDX60 and TMEM140 are newer names for FLJ20035 and FLJ11000 , respectively. ( B ) The ISRE sites in promoter regions (2500 base pairs upstream and 500 base pair downstream from the annotated transcription start sites) of 48 genes induced by ISGF3 and not by U-ISGF3 (listed in ), and those of 29 genes induced by U-ISGF3 were analysed and the results were visualized using the WebLogo software. The canonical ISRE shown in the top panel is annotated in Transfac as a standard sequence. The conserved ISRE sequences of the U-ISGF3-induced genes (bottom panel) are statistically different from the sequences of the genes induced by ISGF3 but not by U-ISGF3 (middle panel), applying symmetrized, position-averaged Kullback-Leibler distance ( P <0.05).

Article Snippet: Mouse monoclonal antibodies against STAT1 (BD Transduction), STAT2, IRF9 (ISGF3γ), and IRF1 (Santa Cruz Biotechnology), and rabbit polyclonal antibodies against Tyr 701-phosphorylated STAT1, Tyr 690-phophorylated STAT2 (Cell Signaling), and STAT1 (Upstate) were used for western analyses.

Techniques: Microarray, Expressing, Stable Transfection, Transfection, Plasmid Preparation, Software, Sequencing

Journal: The EMBO Journal

Article Title: IFNβ-dependent increases in STAT1, STAT2, and IRF9 mediate resistance to viruses and DNA damage

doi: 10.1038/emboj.2013.203

Figure Lengend Snippet: Biological significance of U-ISGF3-induced gene expression. ( A ) U-ISGF3 prolongs anti-viral effects. IFNβ induces the expression of a large number of ISGs (>100 genes) within minutes to hours through the action of ISGF3 (PY-701-STAT1, PY-690-STAT2, and IRF9), which binds to standard ISREs in ISG promoters. ISGs induced by an initial treatment with IFNβ include STAT1, STAT2, and IRF9, and the encoded proteins accumulate in their unphosphorylated forms for days after IFN stimulation. The accumulated STAT1, STAT2, and IRF9 proteins form U-ISGF3, which selectively binds to the distinct ISREs in promoters of a subset of ISGs (about 30 genes), most of which are anti-viral genes. The expression of these genes is prolonged, whereas the expression of the genes induced by ISGF3, but not by U-ISGF3, is terminated rapidly. ( B ) U-ISGF3 induces resistance to DNA damage. Chronic exposure to low doses of IFNβ increases the levels of U-ISGF3 and the U-ISGF3-induced proteins, with no increase in STAT phosphorylation. The proteins induced by U-ISGF3 increase resistance to DNA damage.

Article Snippet: Mouse monoclonal antibodies against STAT1 (BD Transduction), STAT2, IRF9 (ISGF3γ), and IRF1 (Santa Cruz Biotechnology), and rabbit polyclonal antibodies against Tyr 701-phosphorylated STAT1, Tyr 690-phophorylated STAT2 (Cell Signaling), and STAT1 (Upstate) were used for western analyses.

Techniques: Expressing