A rice functional transcriptional activator, RISBZ1, responsible for endosperm-specific expression of storage protein genes through GCN4 motif

The GCN4 motif, a cis-element that is highly conserved in the promoters of cereal seed storage protein genes, plays a central role in controlling endo sperm-specific expression. This motif is the recognition site for a basic l eucine zipper transcriptional factor that belongs to the group of maize Opa que-2 (O2)-like proteins. Five different basic leucine zipper cDNA clones, designated RISBZ1-5, have been isolated from a rice seed cDNA library. The predicted gene products can be divided into two groups based on their amino acid sequences. Although all the RISBZ proteins are able to interact with the GCN4 motif, only RISBZ1 is capable of activating (more than 100-fold ex pression) the expression of a reporter gene under a minimal promoter fused to a pentamer of the GCN4 motif, Loss-of-function and gain-of-function expe riments using the yeast GAL4 DNA binding domain revealed that the proline-r ich N-terminal domain (27 amino acids in length) is responsible for transac tivation, The RISBZ1 protein is capable of forming homodimers as well as he terodimers with other RISBZ subunit proteins. RISBZ1 gene expression is res tricted to the seed, where it precedes the expression of storage protein ge nes. When the RISBZ1 promoter was transcriptionally fused to the beta -gluc uronidase reporter gene and the chimeric gene was introduced into rice, the beta -glucuronidase gene is specifically expressed in aleurone and subaleu rone layer of the developing endosperm, These findings suggest that the spe cific expression of transcriptional activator RISBZ1 gene may determine the endosperm specificity of the storage protein genes.