Endocrine Abstracts

Glucocorticoids (Gcs) exert their pleiotropic effects through activation of the ubiquitously expressed glucocorticoid receptor (GR). The mechanism of transcriptional activation by the GR involves interaction with co-modulator proteins. Glucocorticoid sensitivity is mediated by the expression level of these co-modulator proteins and GR concentration. Therefore, the aim of this study was to identify additional GR interacting proteins that may influence glucocorticoid sensitivity by using a yeast two-hybrid genetic screen.

One of the cDNAs identified as interacting with the GR encoded a novel protein that showed an enhanced interaction with GR in the presence of antagonist, RU486. Database analysis identified 100% identity of this sequence with sequence on human chromosome 17. Bioinformatics analysis identified a 4-exon gene, with an open reading frame identical to the identified cDNA. This predicted a small protein of 121 amino acids, conserved in human, mouse and rat genomes. Northern blot analysis of a human multi-tissue filter identified a prominent transcript of 3kB with high-level expression in placenta, heart and muscle. Further bioinformatics analysis predicted a spliced transcript 3kB in length.

Using a GST pull down assay, this protein was found to interact constitutively with the full length GR with no additional effect of ligand binding. Transfection studies showed that the GR-interacting protein inhibited Gc transactivation of a simple reporter gene (MMTV) in human A549 cells. Immunocytochemistry revealed a mainly nuclear location of the protein when expressed with a cmyc-tag as expected of a transcriptional modulator.

We have identified a novel GR interacting protein that can modulate glucocorticoid sensitivity. We propose that this novel protein may play an important role in the mechanism of Gc-resistance in certain disease states and hence shows potential as a possible therapeutic target.