The promoter of the wheat puroindoline-a gene (PinA) exhibits a more complex pattern of activity than that of the PinB gene and is induced by wounding and pathogen attack in rice

Abstract

Puroindolines form the molecular basis of wheat grain hardness. However, little is known about puroindoline gene regulation. We previously reported that the Triticum aestivum puroindoline-b gene (PinB) promoter directs beta-glucuronidase gene (uidA) seed-specific expression in transgenic rice. In this study, we isolated a puroindoline-a gene (PinA), analyzed PinA promoter activity by 5' deletions and compared PinA and PinB promoters in transgenic rice. Seeds of PinA-1214 and PinB-1063 transgenic plants strongly expressed uidA in endosperm, in the aleurone layer and in epidermis cells in a developmentally regulated manner. The GUS activity was also observed in PinA-1214 embryos. Whereas the PinB promoter is seed specific, the PinA promoter also directed, but to a lower level, uidA expression in roots of seedlings and in the vascular tissues of palea and pollen grains of dehiscent anthers during flower development. In addition, the PinA promoter was induced by wounding and by Magnaporthe grisea. By deletion analysis, we showed that the "390-bp" PinA promoter drives the same expression pattern as the "1214-bp" promoter. Moreover, the "214-bp" PinA promoter drives uidA expression solely in pollen grains of dehiscent anthers. The presence of putative cis-regulatory elements that may be related to PinA expression is discussed from an evolutionary point of view. By electrophoretic mobility shift assay, we showed that putative cis-elements (WUN-box, TCA motifs and as-1-like binding sites) whose presence in the PinA promoter may be related to wounding and/or the pathogen response form complexes with nuclear extracts isolated from wounded wheat leaves.