Analyzing the action of evolutionarily conserved modules on HMW-GS 1Ax1 promoter activity.

key messageThe specific and high-level expression of 1Ax1 is determined by different promoter regions. HMW-GS synthesis occurs in aleurone layer cells. Heterologous proteins can be stored in protein bodies. High-molecular-weight glutenin subunit (HMW-GS) is highly expressed in the endosperm of wheat and relative species, where their expression level and allelic variation affect the bread-making quality and nutrient quality of flour. However, the mechanism regulating HMW-GS expression remains elusive. In this study, we analyzed the distribution of cis-acting elements in the 2659-bp promoter region of the HMW-GS gene 1Ax1, which can be divided into five element-enriched regions. Fragments derived from progressive 5' deletions were used to drive GUS gene expression in transgenic wheat, which was confirmed in aleurone layer cells, inner starchy endosperm cells, starchy endosperm transfer cells, and aleurone transfer cells by histochemical staining. The promoter region ranging from - 297 to - 1 was responsible for tissue-specific expression, while fragments from - 1724 to - 618 and from - 618 to - 297 were responsible for high-level expression. Under the control of the 1Ax1 promoter, heterologous protein could be stored in the form of protein bodies in inner starchy endosperm cells, even without a special location signal. Our findings not only deepen our understanding of glutenin expression regulation, trafficking, and accumulation but also provide a strategy for the utilization of wheat endosperm as a bioreactor for the production of nutrients and metabolic products.