Crucial role for nuclear factor of activated T cells in T cell receptor-mediated regulation of human interleukin-17.
The biological activities of the inflammatory cytokine interleukin (IL)-17 have been widely studied. However, comparatively little is known about how IL-17 expression is controlled. Here, we examined the basis for transcriptional regulation of the human IL-17 gene. IL-17 secretion was induced in peripheral blood mononuclear cells following anti-CD3 cross-linking to activate the T cell receptor (TCR), and costimulatory signaling through CD28 strongly enhanced CD3-induced IL-17 production. To define cis-acting elements important for IL-17 gene regulation, we cloned 1.25 kb of genomic sequence upstream of the transcriptional start site. This putative promoter was active in Jurkat T cells following CD3 and CD28 cross-linking, and its activity was inhibited by cyclosporin A and MAPK inhibitors. The promoter was also active in Hut102 T cells, which we have shown to secrete IL-17 constitutively. Overexpression of nuclear factor of activated T cells (NFAT) or Ras enhanced IL-17 promoter activity, and studies in Jurkat lines deficient in specific TCR signaling pathways provided supporting evidence for a role for NFAT. To delineate the IL-17 minimal promoter, we created a series of 5' truncations and identified a region between -232 and -159 that was sufficient for inducible promoter activity. Interestingly, two NFAT sites were located within this region, which bound to NFATc1 and NFATc2 in nuclear extracts from Hut102 and Jurkat cells. Moreover, mutations of these sites dramatically reduced both specific DNA binding and reporter gene activity, and chromatin immunoprecipitation assays showed occupancy of NFAT at this region in vivo. Together, these data show that NFAT is the crucial sensor of TCR signaling in the IL-17 promoter.
Department of Oral Biology, School of Dental Medicine, University at Buffalo, SUNY, Buffalo, New York 14214, USA.
Xikui K
XK
Liu
Xin
X
Lin
Sarah L
SL
Gaffen
D001483:Base Sequence; D004247:DNA / Q000235:genetics; D004268:DNA-Binding Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D066298:In Vitro Techniques; D020381:Interleukin-17 / Q000235:genetics*; D019169:Jurkat Cells; D008969:Molecular Sequence Data; D050778:NFATC Transcription Factors; D009687:Nuclear Proteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism*; D015398:Signal Transduction; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*
D004268:DNA-Binding Proteins; D020381:Interleukin-17; D050778:NFATC Transcription Factors; C494003:NFATC1 protein, human; C494006:NFATC2 protein, human; D009687:Nuclear Proteins; D011948:Receptors, Antigen, T-Cell; D014157:Transcription Factors; D004247:DNA