Peroxisome Proliferator-Activated Receptor γ-Dependent Repression of the Inducible Nitric Oxide Synthase Gene

Abstract

The peroxisome proliferator-activated receptor γ (PPARγ) is a member of the nuclear receptor superfamily that activates target gene transcription in a ligand-dependent manner. In addition, liganded PPARγ can inhibit transcription of genes induced by gamma interferon (IFN-γ) and/or lipopolysaccharides (LPSs), including the inducible nitric oxide synthase (iNOS) gene. Inhibition of the iNOS promoter is achieved partially through antagonizing the activities of NF-κB, AP-1, and STAT1, which are known to mediate effects of LPS and IFN-γ. Previous studies have suggested that transrepression of these factors by nuclear receptors involves competition for limiting amounts of the general coactivators CREB-binding protein (CBP) and p300. CBP and p300 are thought to be recruited to nuclear receptors through bridging factors that include SRC-1, although CBP also interacts directly with PPARγ through its amino terminus. These observations have raised questions concerning the involvement of SRC-1-like factors in CBP recruitment and transrepression. We here provide evidence that PPARγ's ability to repress iNOS transcription requires the ligand-dependent charge clamp that mediates interactions with CBP and SRC-1. Single amino acid mutations in PPARγ that abolished ligand-dependent interactions with SRC-1 and CBP not only resulted in complete loss of transactivation activity but also abolished transrepression. Conversely, a CBP deletion mutant containing the SRC-1 interaction domain but lacking the N-terminal PPARγ interaction domain was inactive as a PPARγ coactivator and failed to rescue transrepression. Together, these findings are consistent with a model in which transrepression by PPARγ is achieved by targeting CBP through direct interaction with its N-terminal domain and via SRC-1-like bridge factors.

ACKNOWLEDGMENTS

This work was supported by an institutional fellowship from the National Heart, Lung, and Blood Institute to M.L. and NIH grants to C.K.G. C.K.G. is an Established Investigator of the American Heart Association.