Hormonal regulation of a cysteine proteinase gene, EPB-1, in barley aleurone layers: cis- and trans-acting elements involved in the co-ordinated gene expression regulated by gibberellins and abscisic acid.

The synthesis of EPB, a cysteine proteinase responsible for the degradation of seed endosperm storage proteins in barley (Hordeum vulgare), is induced by gibberellins (GA) and repressed by abscisic acid (ABA). The EPB gene family consists of two very similar members, EPB-1 and EPB-2, with the former being more highly induced by GA. We have functionally characterized the cis-acting elements in the EPB-1 promoter and determined that a gibberellin response element (GARE), a pyrimidine box and an upstream element are necessary for GA induction. By comparison with the promoters of alpha-amylase genes, which are also induced by GA, we suggest that GARE is coupled with the upstream element and the pyrimidine box to form a GA response complex. In addition, we have shown that the 3'-untranslated/untranscribed region of the EPB-1 gene is required for a low background expression in the absence of GA. Constitutive expression of a transcription factor, GAMyb, in the absence of GA leads to the transactivation of EPB-1 expression in a dosage dependent manner with the highest level comparable to that in fully GA-induced tissue. Co-expression of a truncated version of GAMyb containing only the DNA binding domain blocks the GA-induction of EPB-1, further supporting the role of GAMyb in the regulation of gene expression. Although ABA is very effective in blocking the GA induction of EPB-1, it has no effect on the GAMyb-mediated expression of EPB-1. We suggest that ABA acts upstream of the formation of functional GAMyb which co-ordinates the hormonal regulation of a diverse group of genes in cereal aleurone layers, including those encoding EPB and alpha-amylases.