Mapping of glucocorticoid receptor DNA binding domain surfaces contributing to transrepression of NF-kappa B and induction of apoptosis.
Glucocorticoids (GCs) function, in part, through the ability of the glucocorticoid receptor (GR) to activate gene expression and in part through the transrepression of AP-1 and NF-kappaB. Here we characterize the effect of GR DNA binding domain (DBD) mutations, previously analyzed for changes in the ability to activate gene expression or transrepress AP-1. We have identified a GR mutant capable of distinguishing between transrepression of NF-kappaB and AP-1. Using circular dichroism spectroscopy, we show that this mutation does not appreciably alter GR DBD conformation, suggesting that functional differences between the mutant and wild type protein are the result of an alteration of a specific interaction surface. These data suggest that transrepression of NF-kappaB and AP-1 occurs through distinct protein-protein interactions and argue against the hypothesis that transrepression occurs through competition for a single coactivator protein. Introduction of these mutations into GC-resistant CEM lymphoblastic T cells restored dexamethasone (DEX)-mediated apoptosis as did wild type GR regardless of whether these mutants were transrepression or activation defective. Thus, DEX-mediated apoptosis in transformed T cells is more complex than originally appreciated.
Tao
Y
Y
Department of Pharmaceutical Sciences, University of Colorado Health Sciences Center, Denver 80262, USA.
Williams-Skipp
C
C
Scheinman
R I
RI
D017209:Apoptosis; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D003907:Dexamethasone / Q000494:pharmacology; D005938:Glucocorticoids / Q000494:pharmacology; D006801:Humans; D016328:NF-kappa B / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D011965:Receptors, Glucocorticoid / Q000235:genetics / Q000378:metabolism*; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism*; D018808:Transcription Factor AP-1 / Q000378:metabolism
D004268:DNA-Binding Proteins; D005938:Glucocorticoids; D016328:NF-kappa B; D011965:Receptors, Glucocorticoid; D012097:Repressor Proteins; D018808:Transcription Factor AP-1; D003907:Dexamethasone