Tissue-specific expression and temporal regulation of the rice glutelin Gt3 gene are conferred by at least two spatially separated cis-regulatory elements.

The rice glutelin Gt3 promoter was fused to a beta-glucuronidase (GUS) reporter gene and its expression evaluated in transgenic tobacco plants. Histochemical analysis revealed that the expression of the introduced Gt3 promoter/GUS (beta-glucuronidase) chimeric gene was confined to endosperm tissue of developing seeds. 5'-promoter deletion analysis revealed that two domains of the Gt3 promoter, -346 to -263 bp (domain I) and -945 to -726 bp (domain II) from the transcriptional start site, were essential for optimum expression of the GUS reporter gene. Removal of 5' sequences upstream of -726 resulted in a reduction in overall promoter activity and a shift in temporal expression from a maximum of 16-20 days after flowering to 24 days. Removal of DNA sequences from the 5' end to -346 yielded a promoter fragment that was still able to confer endosperm-specific expression, although a further deletion to -263 abolished promoter activity. These data suggest that at least two cis-regulatory elements are required for endosperm specificity and temporal regulation of glutelin Gt3 gene expression.