Role of nitrogen-responsive plant-type phosphoenolpyruvate carboxylase in the accumulation of seed storage protein in ancient wheat (spelt and kamut)

Abstract

ABSTRACTAncient wheat such as Triticum spelta L. and T. turanicum is a possible genetic material for improving wheat seed quality due to its grain characteristics, which differ from those of common wheat. Ancient wheat grains accumulate proteins with fewer allergens and higher levels of essential amino acids, antioxidants, minerals and vitamins. The identification of a genetic factor that controls seed protein content would be useful for breeding wheat varieties in accordance with various purposes. Previous studies implied that phosphoenolpyruvate carboxylase (PEPC) is involved in nitrogen accumulation in crop seeds. However, multiple types of PEPC genes are present in plant genomes, and it has still not been clarified yet which type of PEPC is relevant to seed nitrogen metabolism. In this study, to determine which PEPC is involved in seed protein accumulation in ancient wheat, we examined the effects of different levels of nitrogen application at the flowering stage on PEPC activity on developing seeds and on the protein content in mature seeds of three ancient wheat varieties. The responses of the PEPC activity and seed protein content were observed, but their patterns differed among the varieties. Among the experimental plots, PEPC activity and seed protein content showed a high correlation (r = 0.757, p value = 1.09E−3). Immunoblot analysis revealed that plant-type PEPC seemed to underlie the response of PEPC activity to nitrogen application, while no bacterial-type PEPC was detected by a peptide-antigen antibody. These results indicate that plant-type PEPC plays an important role in storage protein accumulation in ancient wheat seeds under nitrogen application at the flowering stage.Abbreviations: PEPC, phosphoenolpyruvate carboxylase; 2-ME, 2-mercapto ethanol; EDTA, ethylenediaminetetraacetic acid; DAF, days after flowering; SDS-PAGE, sodium dodecylsulfate-polyacrylamide gel electrophoresis.