Cortisol resistance and the guinea pig glucocorticoid receptor

The guinea pig has been employed as a model to study the structure/function relationships of the glucocorticoid receptor (GR), and to determine the regions of the receptor important for binding hormone and antihormone. Guinea pigs have high levels of circulating cortisol and GR with a approximately 20-fold lower affinity for dexamethasone than mouse GR. Cloning and sequencing of guinea pig GR has identified 24 amino acid changes in the ligand-binding domain (LBD) compared to the human GR. By substituting the guinea pig GR LBD for the human LBD in a human GR expression vector we have shown in cotransfection studies that guinea pig GR LBD confers glucocorticoid resistance as observed in vivo. In initial studies guinea pig GR LBD appeared constitutively active; in subsequent studies to determine which of the 24 amino acid changes present in the guinea pig GR LBD conferred resistance, it became apparent that the guinea pig LBD (LBD delta), amplified by PCR for subcloning into the human GR expression vector, contained a single adenine deletion in the hinge region within ten bases of the PCR primer. This single base deletion resulted in a frameshift bringing a stop codon into frame one codon after the deletion. While this now clearly accounts for the observed constitutive activity, since it is known that C-terminally truncated steroid receptors exhibit constitutive activation such a truncation is more difficult to reconcile with the repeatedly demonstrable hormone dose-response curves obtained with this guinea pig GR LBD delta.(ABSTRACT TRUNCATED AT 250 WORDS)