Glucocorticoids are efficient antiinflammatory agents, and their effects include transcriptional repression of several cytokines and adhesion molecules. Whereas glucocorticoids down-regulate the expression of genes relevant during inflammation, nuclear factor (NF)-kappa B/Rel proteins function as important positive regulators of these genes. The expression of intercellular adhesion molecule-1 (ICAM-1), which plays an essential role in recruitment and migration of leukocytes to sites of inflammation, is also down-regulated by glucocorticoids. We found that a functional NF-kappa B site in the ICAM-1 promoter, which can be activated by either 12-O-tetradecanoylphorbol-13-acetate or tumor necrosis factor-alpha (TNF alpha), is also the target for glucocorticoids. In this report we present evidence that the ligand-activated glucocorticoid receptor (GR) is able to repress RelA-mediated activation of the ICAM-1 NF-kappa B site. Conversely, transcriptional activation by GR via a glucocorticoid response element is specifically repressed by RelA, but not by other NF-kappa B/Rel family members. Mutational analysis of GR demonstrates that the DNA binding domain and the ligand binding domain are required for the functional repression of NF-kappa B activation. Despite the importance of the DNA binding domain, we found that the transcriptional repression of NF-kappa B, mediated by GR, is not caused by binding of GR to the ICAM-1 NF-kappa B element, but by a physical interaction between the GR and RelA protein. The repressive effect of GR on NF-kappa B-mediated activation was not shared by other steroid/thyroid receptors. Only the progesterone receptor, which belongs to the same subfamily as GR and which possesses high homology with GR, was able to repress NF-kappa B-mediated transcription. These studies highlight a possible molecular mechanism that can explain the antiinflammatory effects of glucocorticoid treatment during inflammation.

Interactions between transcription factors are an important means of regulating gene transcription. The present study describes the mutual repression of two transcription factors, the RelA(p65) subunit of NF-B and the progesterone receptor (PR). This trans-repression is shown to occur independent of PR isoform, reporter construct, or cell type used. Together with the demonstration of an interaction between PR and RelA in vitro, these findings suggest that the mutual repression is due to a direct interaction between these proteins. Furthermore, activation of NF-B by tumor necrosis factor-␣ also results in repression of PR, while PR is able to repress tumor necrosis factor-␣-induced NF-B activity. Since NF-B-regulating cytokine receptors are expressed in progesterone target tissues, like breast and endometrium, the mutual repression of PR and RelA could play an important role in a wide variety of physiological processes in these tissues, including maintenance of pregnancy, immunosuppression, and tumorigenesis.The human progesterone receptor (PR) 1 belongs to the superfamily of steroid/thyroid hormone receptors (1-3). The transcription factors of this family share (at least) two structurally related functional domains, the DBD, which contains the socalled zinc finger motif, and the more C-terminally located HBD. Two transactivation domains have been mapped in the PR, of which one is located N-terminal to the DBD. This transactivation domain, named AF-1, functions autonomously, but the level of activity depends on the cell type and reporter construct used (4, 5). The second transactivation domain AF-2 lies within the HBD, and its activity is strictly dependent on the presence of ligand (4). Detailed analysis of the function of these transactivation domains of the PR has also led to more insight in anti-hormone action. Anti-progestins bind to the receptor without activating AF-2, and the partial agonistic activity evoked by the anti-progestin RU486 is therefore the result of its stimulation of AF-1 activity (4, 5).Within the superfamily of steroid receptors, PR is unique in that it exists in two isoforms, named A and B (6), which differ in their N terminus. Differences in transcriptional activity between the PR B (amino acids 1-933) and PR A (amino acids 164 -933) isoforms have been observed, depending on the cell type and the reporter construct used (4, 5).Functioning of transcription factors can be modified by interplay with transcription factors of a different type, resulting in either an inhibitory or stimulatory effect. In particular, interactions between steroid receptors and members of the AP1 family of transcription factors have been studied extensively. AP1 family members, like c-Jun and c-Fos, and GR have been shown to repress each others functioning (7-11), but the actual mechanism of repression is currently controversial. In vitro, a direct interaction between GR and AP1 was shown, resulting in impaired DNA binding (7, 9), while others failed to detect a direct interaction (10, 11) or found the two prot...

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The RelA subunit of NF-B and the glucocorticoid receptor mutually repress each others transcriptional activity, thus providing a mechanism for immunosuppression. Deletion analysis of the glucocorticoid receptor has shown that the DNA binding domain and the ligand binding domain are essential components for repression. Here, we show by deletions and point mutations that both the Rel homology domain and the transactivation domains of RelA are required for repression of the transcriptional activity of the glucocorticoid receptor in intact cells. However, only the Rel homology domain of RelA was found to associate with the glucocorticoid receptor in vitro. RelA mutants, not able to repress glucocorticoid receptor activity, but still able to dimerize, behaved as transdominant inhibitors of the repressive activity of wild type RelA. Furthermore, we show that the 13 S E1A protein is able to interfere with the transrepressive activity of RelA. We propose that negative cross-talk between the glucocorticoid receptor and RelA is due to direct interaction via the Rel homology domain of RelA and the DNA binding domain of the glucocorticoid receptor in combination with interference by the transactivation domains of RelA with the transcriptional activity of the glucocorticoid receptor.