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. 1994 Dec;14(12):8007–8017. doi: 10.1128/mcb.14.12.8007

A functional glucocorticoid-responsive unit composed of two overlapping inactive receptor-binding sites: evidence for formation of a receptor tetramer.

M Garlatti 1, M Daheshia 1, E Slater 1, J Bouguet 1, J Hanoune 1, M Beato 1, R Barouki 1
PMCID: PMC359339  PMID: 7969140

Abstract

An unusual glucocorticoid-responsive element (called GRE A) was found to mediate the induction of the cytosolic aspartate aminotransferase gene 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 heterologous 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 electrophoretic mobility shift assays, purified rat liver glucocorticoid receptor (GR) formed a low-mobility complex with GRE A, presumably containing a GR tetramer. When purified bacterially expressed DBD was used, low-mobility complexes as well as dimer and monomer complexes were formed. In inactive mutated oligonucleotides, no GR tetramer formation was detected. Modification of the imperfect half-sites in order to increase their affinity for GR gave a DNA sequence that bound a GR tetramer in a highly cooperative manner. This activated unit consisting of two overlapping consensus GREs mediated glucocorticoid induction with a higher efficiency than consensus GRE.

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