Multifactor cis-dominant negative regulation of IL-2 gene expression in anergized T cells.
The molecular mechanism underlying IL-2 transcriptional blockade in anergic T cell clones is not fully understood. To examine whether an active negative regulatory process occurs, we created a reporter construct containing as an enhancer four copies of the NF-AT site and one copy of the octamer site (4X NF-AT-Oct). This construct was only slightly reduced (1.3-fold) in its expression when stimulated under anergic conditions, while a whole mouse IL-2 enhancer construct showed a reduction of 4.3-fold. Addition of the -176 to -96 sequence to the 4X NF-AT-Oct construct did not impart the ability to be affected by anergy, but addition of the -236 to -96 sequence did, demonstrating that anergy is an active inhibitory process and that more than the presence of the -150 AP-1 binding site (-152 to -147) is required to mediate the effect. Mutational studies of the -236 to -96 sequence indicated that the presence of both the -130 AP-1-like site (-187 to -181) and the -150 proximal AP-1 site were necessary to observe anergy. Because the -180 site is not required for trans-activation, it was possible to confirm by mutation in the normal mouse IL-2 enhancer that this site is absolutely essential for anergy induction. The simplest model to explain these results is that anergy is mediated by a complex of multiple transcription factors that exert a cis-acting dominant negative regulatory effect on the trans-activation of the IL-2 gene.
Kitagawa-Sakakida
S
S
Laboratory of Cellular and Molecular Immunology, National Institute of Allergy and Infectious Diseases, Bethesda, MD 20892, USA.
Schwartz
R H
RH
D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D017635:Clonal Anergy / Q000235:genetics*; D002999:Clone Cells / Q000276:immunology; D004742:Enhancer Elements, Genetic / Q000276:immunology; D005786:Gene Expression Regulation / Q000276:immunology*; D017930:Genes, Reporter / Q000276:immunology; D006801:Humans; D007376:Interleukin-2 / Q000235:genetics*; D051379:Mice; D009154:Mutation / Q000276:immunology; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000276:immunology; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism*; D018808:Transcription Factor AP-1 / Q000235:genetics; D014158:Transcription, Genetic / Q000276:immunology
D000911:Antibodies, Monoclonal; D007376:Interleukin-2; D016693:Receptors, Antigen, T-Cell, alpha-beta; D018808:Transcription Factor AP-1