Identification of cis-acting elements important for expression of the starch-branching enzyme I gene in maize endosperm

The genes encoding the starch-branching enzymes (SBE) SBEI, SBEIIa, and SBE IIb in maize (Zea mays) are differentially regulated in tissue specificity and during kernel development. To gain insight into the regulatory mechanis ms controlling their expression, we analyzed the 5'-flanking sequences of S be1 using a transient gene expression system. Although the 2.2-kb 5'-flanki ng sequence between -2,190 and +27 relative to the transcription initiation site was sufficient to promote transcription, the addition of the transcri bed region between +28 and +228 containing the first exon and intron result ed in high-level expression in suspension-cultured maize endosperm cells. A series of 5' deletion and linker-substitution mutants identified two criti cal positive cis elements, -314 to -295 and -284 to -255. An electrophoreti c mobility-shift assay showed that nuclear proteins prepared from maize ker nels interact with the 60-bp fragment containing these two elements. Expres sion of the Sbe1 gene is regulated by sugar concentration in suspension-cul tured maize endosperm cells, and the region -314 to -145 is essential for t his effect. Interestingly, the expression of mEmBP-1, a bZIP transcription activator, in suspension-cultured maize endosperm cells resulted in a 5-fol d decrease in Sbe1 promoter activity, suggesting a possible regulatory role of the C-box present in the Sbe1? promoter from -227 to -220.