Ability of the glucocorticoid modulatory element to modify glucocorticoid receptor transactivation indicates parallel pathways for the expression of glucocorticoid modulatory element and glucocorticoid response element activities.

The glucocorticoid modulatory element (GME) of the rat tyrosine aminotransferase gene is located at -3.6 kb and 1 kb upstream of the glucocorticoid response elements (GREs). The GME has the unique transcriptional properties of modulating both the dose-response curve of agonists bound to the glucocorticoid receptor (GR) and the residual agonist activity of GR-bound antisteroids. The expression of GME activity involves the binding of two novel proteins (GMEB-1 and GMEB-2) that we have recently cloned. However, the mechanistic details are limited. The DNA sequence requirements for GME activity (CGTC) also remain poorly defined, which restricts efforts to identify other GME modulated genes. To help understand the mechanism for the unusual activities of the GME and to identify permissive gene environments for GME activity, we compared the changes in GME activity and GRE action (i.e. the fold induction by GR) caused by modifying several parameters. Phasing between the GME and downstream tandem GREs was unimportant, in contrast to other cis-acting elements like the GRE, while GME activity decreased rapidly when placed at increasingly larger distances 3' to a tandem GRE. A minimal promoter was less effective in supporting GME than GRE activity. Although CREB binds to the GME, overexpression of CREB reduced GRE, but not GME, activity and a CRE was inactive when substituted for the GME. No effect of the GME was observed on the binding of GRs to a single GRE. However, the GME upstream of a single GRE was also unable to produce a left shift in the Dex dose-response curve under conditions where the GME was active with two GREs. In the absence of any GREs, the GME displayed intrinsic activity by elevating basal level expression. Collectively, these results indicate that an optimal position for a functional GME is within 250 bp upstream of a tandem GRE driving a complex promoter. Furthermore, as the changes in GME activity did not correlate with those for fold induction from the GRE, the mechanisms for expression of GME and GRE activities appear to utilize parallel, as opposed to common pathways.