RNAi-mediated knockdown of the XIP-type endoxylanase inhibitor gene, OsXIP, has no effect on grain development and germination in rice.

OsXIP (Oryza sativa xylanase inhibitor protein) is a XIP-type xylanase inhibitor which was identified as a protein encoded by a wound stress-responsive gene in rice. Although the OsXIP gene was specifically expressed in mature grains under basal conditions, recombinant OsXIP had no effect on rice endogenous xylanases, and OsXIP-suppressed transgenic rice plants did not exhibit any change in grain development and germination, suggesting that rice development may be independent of OsXIP. Analysis using an OsXIP-specific antibody revealed that OsXIP is markedly accumulated in apoplast in rice root cells by wounding. These results reinforced the possibility that OsXIP is involved in plant defense mechanisms against phytopathogens.

Induction of a novel XIP-type xylanase inhibitor by external ascorbic acid treatment and differential expression of XIP-family genes in rice.

Rice microarray analysis showed that a number of stress-related genes are induced by external addition of L-ascorbic acid (AsA). The gene designated as AK073843 which is homologous to class capital SHA, Cyrillic chitinase was found to exhibit the highest induction among these genes. However, its crucial residues within the chitinase active site are substituted with other residues, suggesting that the protein has no chitinase activity. The recombinant protein which is encoded by the AK073843 gene produced in Escherichia coli has xylanase inhibitor activity, indicating that the gene encodes a novel rice XIP-type xylanase inhibitor protein (OsXIP). The expression of OsXIP was enhanced not only by exogenous AsA treatment but also by various stresses such as citrate and sodium chloride treatments, and wounding; however, it was not influenced by increasing endogenous AsA content. External AsA treatment caused a significant increase in electrolyte leakage from rice root. These results suggested that OsXIP was induced by stress which is caused by external AsA treatment. Rice XIP-family genes, OsXIP, riceXIP and RIXI, showed differential organ-specific expression. Also, these genes were differentially induced by stress and stress-related phytohormones. The transcripts of OsXIP and riceXIP were undetectable under normal conditions, and were drastically induced by wounding and methyl jasmonate (MeJA) treatment in the root. RIXI was constitutively expressed in the shoot but not induced by wounding and stress-related phytohormones. Thus, XIP-type xylanase inhibitors were suggested to be specialized in their function and involved in defense mechanisms in rice.

Generation and properties of ascorbic acid-overproducing transgenic tobacco cells expressing sense RNA for l-galactono-1,4-lactone dehydrogenase.

L-Galactono-1,4-lactone dehydrogenase (GalLDH; EC 1.3.2.3) is the last enzyme in the putative L-ascorbic acid (AsA) biosynthetic pathway of plants. Here, we show for the first time that the overexpression of GalLDH can increase AsA content in tobacco (Nicotiana tabacum L.) BY-2 cells. To see the effect, we analyzed the properties of these AsA-overproducing transgenic cell lines, especially in relation to AsA content of cells, cell division, senescence and resistance to oxidative stress. The mitotic index in AsA-overproducing cells was higher than in wild-type cells. Moreover, the browning of these cells was markedly restrained, and the proportion of dead cells was reduced, especially in the later period of culture. These AsA-overproducing cells also acquired resistance to paraquat (methyl viologen), which produces active oxygen species. These results contribute to the previous insights about AsA and raise the possibility of the generation of plants that have resistance to environmental stresses by increasing their AsA content.