Receptor interacting protein RIP140 inhibits both positive and negative gene regulation by glucocorticoids

Recent development in the field of gene regulation by nuclear receptors (NR s) have identified a role for cofactors in transcriptional control. While s ome of the NR-associated proteins serve as coactivators, the effect of the receptor interacting protein 140 (RIP140) on NR transcriptional responses i s complex. In this report we have studied the effect of RIP140 on gene regu lation by the glucocorticoid receptor (GR). We demonstrate that RIP140 anta gonized all GR-mediated responses tested, which included activation through classical GRE, the synergistic effects of glucocorticoids on AP-1 and Pbx1 /HOXB1 responsive elements, as well as gene repression through a negative G RE and cross-talk with NF-kappa B (RelA). This involved the ligand-binding domain of the GR and did not occur when the GR was bound to the antagonist RU486, The strong repressive effect of RIP140 was restricted to glucocortic oid-mediated responses in as much as it slightly increased signaling throug h the RelA and the Pit-1/Pbx proteins and only slightly repressed signaling through the Pbx1/HOXB1 and AP-1 proteins, excluding general squelching as a mechanism. Instead, this suggests that RIP140 acts as a direct inhibitor of GR function. In line with a direct effect of RIP140 on the GR, we demons trate a GR-RIP140 interaction in vitro by a glutathione S-transferase-pull down assay. Furthermore, the repressive effect of RIP140 could partially be overcome by overexpression of the coactivator TIF2, which involved a compe tition between TIF2 and RIP140 for binding to the GR.