IDENTIFICATION OF AMINO-ACIDS IN THE TAU-2-REGION OF THE MOUSE GLUCOCORTICOID RECEPTOR THAT CONTRIBUTE TO HORMONE-BINDING AND TRANSCRIPTIONAL ACTIVATION

The tau(2)-region of steroid hormone receptors is a highly conserved r egion located at the extreme N-terminal end of the hormone-binding dom ain. A protein fragment encoding tau 2 has been shown to function as a n independent transcriptional activation domain; however, because this region is essential for hormone binding, it has been difficult to det ermine whether the tau(2)-region also contributes to the transactivati on function of intact steroid receptors. In this study a series of ami no acid substitutions were engineered at conserved positions in the ta u 2-region of the mouse glucocorticoid receptor (mGR, amino acids 533- 562) to map specific amino acid residues that contribute to the hormon e-binding function, transcriptional activation, or both. Substitution of alanine or glycine for some amino acids (mutations E546G, P547A, an d D555A) reduced or eliminated hormone binding, but the transactivatio n function of the intact GR and/or the minimum tau 2-fragment was unaf fected for each of these mutants. Substitution of alanine for amino ac id S561 reduced transactivation activity in the intact GR and the mini mum tau 2-fragment but had no effect on hormone binding. The single mu tation L550A and the double amino acid substitution L541G+L542G affect ed both hormone binding and transactivation. The fact that the S561A a nd L550A substitutions each caused a loss of transactivation activity in the minimum tau 2-fragment and the full-length GR indicated that th e tau 2-region does contribute to the overall transactivation function of the full-length GR. Overall, the N-terminal portion of the tau 2-r egion (mGR 541-547) was primarily involved in hormone binding, whereas the C-terminal portion of the tau 2-region (mGR 548-561) was primaril y involved in transactivation.