Target genes and regulatory domains of the GAMYB transcriptional activator in cereal aleurone

Abstract

GAMYB is an MYB transcription factor which is expressed in cereal aleurone cells in response to gibberellin (GA). HvGAMYB binds to the TAACAAA box of a high-pl α-amylase gene promoter and transcriptionally activates its expression. In this study, we examined the role of HvGAMYB in activating expression of other GA-regulated genes encoding hydrolytic enzymes. In transient expression experiments, HvGAMYB transactivated expression of reporter genes fused to a low-pl α-amylase gene promoter, an EII(1-3,1-4)-β-glucanase gene promoter and a cathepsin B-like protease promoter. HvGAMYB DNA binding specificity was determined using a PCR-based random site selection using HvGAMYB fusion protein isolated from E. coli. The deduced consensus closely resembled gibberellin response elements in α-amylase promoters. Functional analysis of HvGAMYB transient expression of C terminal HvGAMYB deletions in barley aleurone cells identified two transcriptional activation domains (TADs) which function in transcriptional regulation of both high- and low-pl α-amylase promoters. The same TADs were identified using a heterologous yeast expression system. Together, these results indicate that HvGAMYB has two TADs. These domains are C-terminal to its DNA-binding domain.