A FUNCTIONAL GLUCOCORTICOID-RESPONSIVE UNIT COMPOSED OF 2 OVERLAPPINGINACTIVE RECEPTOR-BINDING SITES - EVIDENCE FOR FORMATION OF A RECEPTOR TETRAMER

An unusual glucocorticoid-responsive element (called GRE A) was found to mediate the induction of the cytosolic aspartate aminotransferase g ene by glucocorticoids and was bound by the glucocorticoid receptor in a DNase I footprinting assay. GRE A consists of two overlapping GREs, each comprising a conserved half-site and an imperfect half-site. The complete unit was able to confer glucocorticoid inducibility to a het erologous promoter (Delta MTV-CAT). Mutation of any of the half-sites, including the imperfect ones, abolished inducibility by the hormone, demonstrating that each of the isolated GREs was inactive. In electrop horetic mobility shift assays, purified rat liver glucocorticoid recep tor (GR) formed a low-mobility complex with GRE A, presumably containi ng a GR tetramer. When purified bacterially expressed DBD was used, lo w-mobility complexes as well as dimer and monomer complexes were forme d. In inactive mutated oligonucleotides, no GR tetramer formation was detected. Modification of the imperfect half-sites in order to increas e their affinity for GR gave a DNA sequence that bound a GR tetramer i n a highly cooperative manner. This activated unit consisting of two o verlapping consensus GREs mediated glucocorticoid induction with a hig her efficiency than consensus GRE.