DEVELOPMENTAL CONTROL OF THE BETA-PHASEOLIN GENE REQUIRES POSITIVE, NEGATIVE, AND TEMPORAL SEED-SPECIFIC TRANSCRIPTIONAL REGULATORY ELEMENTS AND A NEGATIVE ELEMENT FOR STEM AND ROOT EXPRESSION

To identify cis-acting regulatory elements responsible for developmental control of the common bean seed storage protein beta-phaseolin, a series of 5'-deletion mutants of the 782 bp upstream sequence together with the coding and 3'-regions of the beta-phaseolin gene were used to transform tobacco. One major positive regulatory element (-295/-228) and a putative minimal promoter (-64/-14) were indicated by large reductions in phaseolin mRNA and protein concentrations in seeds of plants deficient for these regions. In addition, minor negative (-422/-296) and positive (-782/-423) elements also influenced the level of phaseolin mRNA expression in seeds. One temporal element (-295/-107) governed late expression of phaseolin mRNA in seeds, and possibly a second (-64/-14) regulated early expression. The -64/-14 promoter containing two TATA boxes conferred spatially regulated gene expression, and was sufficient for a low level of expression in root and stem. Significant levels of phaseolin mRNA and protein were detected in stem cortices and secondary roots of plants lacking the -295/-107 negative element. No significant expression in leaf tissue was detected. Results demonstrate that developmental control of beta-phaseolin requires a minimal promoter, one element for the suppression of expression in root and stem tissue, three elements governing quantitative expression in seeds, and at least one temporal element controlling expression in seeds.