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Analysis of IRF-7 (pS477/pS479) in transfected human epithelial cells. The 293 fetal kidney cell line was either co-transfected with TBK1 and IRF-7 expression vectors (dashed histogram) or un-transfected (solid line). After 24 hours, the cells were fixed (BD Cytofix™ Fixation Buffer, Cat. No. 554655) for 10 minutes at 37°C, then permeabilized (BD Phosflow™ Perm Buffer III, Cat. No. 558050) on ice for at least 30 minutes, and then stained with PE Mouse anti-IRF-7 (pS477/pS479). Flow cytometry was performed on a BD FACSCalibur™ flow cytometry system.
BD™ Phosflow PE Mouse anti-IRF-7 (pS477/pS479)
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- This reagent has been pre-diluted for use at the recommended Volume per Test. We typically use 1 × 10^6 cells in a 100-µl experimental sample (a test).
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Interferon regulatory factor 7 (IRF-7) is a transcription factor that regulates anti-viral defenses by controlling the induction of type-I interferon (IFN) responses. IRF-7 expression is induced in lymphoid cells by virus infection, as well as by IFN, lipopolysaccharide, and TNF-α. IRF-7 responses are initiated by Toll-like receptors (TLR) or the cytoplasmic protein retinoic acid inducible gene I (RIG-I). Upon TLR activation, it forms cytoplasmic complexes with MyD88, an adaptor in the TLR signaling pathways. The TLR-dependent and RIG-I-dependent pathways activate kinases, such as IKK-ε and TBK1, that phosphorylate IRF-7 and induce movement of IRF-7-containing complexes to the nucleus, where it preferentially activates IFN-α promoters.
The K47-671 monoclonal antibody recognizes human IRF-7 phosphorylated at serines 477 and 479 (pS477/pS479). Our in-house testing is performed on a cell line that has been co-transfected with TBK1 and IRF-7. Phosphorylation of IRF-7 in the transfectants requires virus infection or over-expression of a signaling molecule of the RIG-I pathway, such as TBK1. Phosphorylation of endogenous IRF-7 in untransfected cells has not yet been detected. We confirmed that mAb K47-671 does not cross-react with TBK1 by Western blot analysis using the purified antibody.
研发参考 (8)
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Honda K, Yanai H, Mizutani T, et al. Role of a transductional–tanscriptional processor complex involving MyD88 and IRF-7 in Toll-like receptor signaling. Proc Natl Acad Sci U S A. 2004; 101(43):15416-15421. (Biology). 查看参考
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Honda K, Yanai H, Negishi H, et al. IRF-7 is the master regulator of type-I interferon-dependent immune responses. Nature. 2005; 434:772-777. (Biology). 查看参考
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Hoshino K, Sugiyama T, Matsumoto M, et al. IκB kinase-α is critical for interferon-α production induced by Toll-like receptors 7 and 9. Nature. 2006; 440(7086):949-953. (Biology). 查看参考
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Kawai T, Sato S, Ishii KJ, et al. Interferon-α induction through Toll-like receptors involves a direct interaction of IRF7 with MyD88 and TRAF6. Nat Immunol. 2004; 5(10):1061-1068. (Biology). 查看参考
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Lin R, Mamane Y, Hiscott J. Multiple regulatory domains control IRF-7 activity in response to virus infection. J Biol Chem. 2000; 275(44):34320-34327. (Biology). 查看参考
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Matikainen S, Sirén J, Tissari J, et al. Tumor necrosis factor alpha enhances influenza A virus-induced expression of antiviral cytokines by activating RIG-I gene expression. J Virol. 2006; 80(7):3515-3522. (Biology). 查看参考
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Paz S, Sun Q, Nakhaei P, et al. Induction of IRF-3 and IRF-7 phosphorylation following activation of the RIG-I pathway. Cell Mol Biol (Noisy-le-grand). 2006; 52(1):17-28. (Clone-specific). 查看参考
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Sharma S, tenOever BR, Grandvaux N, Zhou G-P, Lin R, Hiscott J. Triggering the interferon antiviral response through an IKK-related pathway. Science. 2003; 300:1148-1151. (Biology). 查看参考
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