Glucocorticoids repress NF-kappa B-driven genes by disturbing the interaction of p65 with the basal transcription machinery, irrespective of coactivator levels in the cell

Glucocorticoids (GCs) are used to combat inflammatory diseases. Their benef icial effect relies mainly on the inhibition of NF-kappa B-and/or AP-1-driv en proinflammatory gene expression. Previously, we have shown that GCs repr ess tumor necrosis factor-induced IL-6 gene expression by an NF-kappa B-dep endent nuclear mechanism without changing the DNA-binding capacity of NF-ka ppa B or the expression levels of the cytoplasmic inhibitor of NF-kappa B ( 1 kappa B-alpha), in the present work, we investigate the effect of GC repr ession on different natural and/or recombinant NF-kappa B-driven reporter g ene constructs in the presence of increasing amounts of various coactivator molecules, such as CREB-binding protein (CBP), p300, and SRC-1. We found t hat GCs maintain their repressive capacities, irrespective of the amount of cofactor present in the cell. Similar results were obtained for the recipr ocal transrepression of a GC receptor (GR) element-driven reporter gene by p65, We demonstrate that neither the expression levels of p65 and CBP nor t heir physical association are affected by activated CR, Using Gal4 chimeras , we show that repression by GCs is specific for p65-mediated transactivati on, ruling out competition for limiting nuclear factors as the major underl ying mechanism of gene repression. In addition, the transactivation potenti al of a point-mutated Cal4-p65 variant with a decreased CBP interaction cap ability is still repressed by GR. Finally, we present evidence that the spe cificity of GC repression on p65-driven gene expression is codetermined by the TATA box context.