Development of monoclonal antibodies against GBF1 and their use in studying its functions.
Interferon-gamma (IFN-gamma) regulates a number of cellular genes using a variety of cellular signaling pathways. Previously, we identified a novel IFN-regulated element, IFN-gamma-activated transcriptional element (GATE), in the promoter of the murine IFN regulatory factor-9 (IRF-9) gene. This element binds to novel factors. We have recently characterized a novel regulatory factor, GATE binding factor 1 (GBF1), which promoted IFN-gamma-induced transcription. Although GBF1 was a potent inducer of transcription, it did not bind to DNA well in vitro. To understand its role in IFN-gamma-induced actions, we raised monoclonal antibodies (mAb) against GBF1. These antibodies are highly useful in Western, immunoprecipitation, and immunocytochemical analyses. Employing these antibodies, we show that GBF1 is recruited to the endogenous IRF-9 promoter. We also show GBF1 interacts with CAAAT/enhancer binding protein-beta (C/EBP-beta), the other GATE binding factor. Furthermore, other cytokines, such as interleukin-1 (IL-1) and IL-6, induced the expression of GBF1. These antibodies may be useful tools for investigating the role of GBF1 in cytokine-induced responses.
Roy
Sanjit K
SK
Greenebaum Cancer Center, Department of Microbiology and Immunology, 655 W. Baltimore Street, University of Maryland School of Medicine, Baltimore, MD 21201, USA.
Hu
Junbo
J
Meng
Qingjun
Q
Hu
Jiadi
J
Reddy
Sekhar P
SP
Kalvakolanu
Dhananjaya V
DV
D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D015153:Blotting, Western; D022782:CCAAT-Enhancer-Binding Protein-beta / Q000378:metabolism*; D002460:Cell Line; D047369:Chromatin Immunoprecipitation; D016207:Cytokines / Q000378:metabolism; D004247:DNA / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay; D020662:Guanine Nucleotide Exchange Factors / Q000737:chemistry* / Q000276:immunology; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry; D047468:Immunoprecipitation; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit / Q000235:genetics*; D007371:Interferon-gamma / Q000378:metabolism*; D007372:Interferons / Q000378:metabolism; D007375:Interleukin-1 / Q000378:metabolism*; D051379:Mice; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry; D015398:Signal Transduction; D013997:Time Factors; D014158:Transcription, Genetic
D000911:Antibodies, Monoclonal; D022782:CCAAT-Enhancer-Binding Protein-beta; D016207:Cytokines; C120445:GBF1 protein, human; D020662:Guanine Nucleotide Exchange Factors; C494227:IRF9 protein, human; D050834:Interferon-Stimulated Gene Factor 3, gamma Subunit; D007375:Interleukin-1; D011994:Recombinant Proteins; D007371:Interferon-gamma; D004247:DNA; D007372:Interferons