DNA-dependent cofactor selectivity of the glucocorticoid receptor
Final Report Abstract
Our lab has a long standing research interest in GR-mediated transcriptional repression on the one hand, and in histone deacetylases on the other hand. In addition, we discovered that acetylation of STAT1 affects its transcriptional activity as well as its crosstalk with NF-kB. Both proteins are central regulators of immune functions. Furthermore, we found that the mutual modulation of different modifications, such as acetylation, phosphorylation, ubiquitylation and sumoylation, is an important aspect of the cross-talk and fine-tuning of signaling pathways. When we extended our analysis to the acetylation of GR, we were surprised that we were unable to detect this modification in response to treatment of cells with inhibitors of class I/II HDACs. Unexpectedly, acetylation of GR was only detectable in response to class III HDAC inhibitors such as NAM or Ex-527. SIRT1-specific inhibitors as well as siRNA knockdown experiments suggest that SIRT1 is indeed the primary GR deacetylase. Our mass spectrometry analysis of post-translational modifications of GR identified novel acetylation sites in the N-terminal region whereas previously published sites are located within the GR hinge region. The reduced activity of the GR in reporter gene assays suggests that the acetylation status is indeed relevant for GR function. SIRT1 is a cellular energy sensor, responsive to metabolic fluctuations of NAD+ levels. Metabolic signaling by the AMP kinase (AMPK) has been shown to modulate SIRT1 activity by stimulating the NAD+ synthesis. According to our results AMPK signaling reduces GR acetylation, indicating that energy metabolism regulates GR in an acetylation-dependent manner.
