Function of xyloglucan endotransglucosylase/hydrolases in rice.

BACKGROUND AND AIMS: Although xyloglucans are ubiquitous in land plants, they are less abundant in Poales species than in eudicotyledons. Poales cell walls contain higher levels of ß-1,3/1,4 mixed-linked glucans and arabinoxylans than xyloglucans. Despite the relatively low level of xyloglucans in Poales, the xyloglucan endotransglucosylase/hydrolase (XTH) gene family in rice (Oryza sativa) is comparable in size to that of the eudicotyledon Arabidopsis thaliana. This raises the question of whether xyloglucan is a substrate for rice XTH gene products, whose enzyme activity remains largely uncharacterized. METHODS: This study focused on OsXTH19 (which belongs to Group IIIA of the XTH family and is specifically expressed in growing tissues of rice shoots), and two other XTHs, OsXTH11 (Group I/II) and OsXTH20 (Group IIIA), for reference, and measurements were made of the enzymatic activities of three recombinant rice XTHs, i.e. OsXTH11, OsXTH20 and OsXTH19. KEY RESULTS: All three OsXTH gene products have xyloglucan endohydrolase (XEH, EC 3·2·1·151) activity, and OsXTH11 has both XEH and xyloglucan endotransglycosylase (XET, EC 2·4·1207) activities. However, these proteins had neither hydrolase nor transglucosylase activity when glucuronoarabinoxylan or mixed-linkage glucan was used as the substrate. These results are consistent with histological observations demonstrating that pOsXTH19::GUS is expressed specifically in the vicinity of tissues where xyloglucan immunoreactivity is present. Transgenic rice lines over-expressing OsXTH19 (harbouring a Cauliflower Mosaic Virus 35S promoter::OsXTH19 cDNA construct) or with suppressed OsXTH19 expression (harbouring a pOsXTH19 RNAi construct) did not show dramatic phenotypic changes, suggesting functional redundancy and collaboration among XTH family members, as was observed in A. thaliana. CONCLUSIONS: OsXTH20 and OsXTH19 act as hydrolases exclusively on xyloglucan, while OsXTH11 exhibits both hydrolase and XET activities exclusively on xyloglucans. Phenotypic analysis of transgenic lines with altered expression of OsXTH19 suggests that OsXTH19 and related XTH(s) play redundant roles in rice growth.

Involvement of plasma membrane H+-ATPase in anoxic elongation of stems in pondweed (Potamogeton distinctus) turions.

• Pondweed (Potamogeton distinctus) turions can elongate in the absence of O(2). Alcoholic fermentation serves to produce energy for anoxic elongation via the breakdown of starch stored in cells. However, the mechanism of cell growth during anoxic elongation is not fully understood. • Changes in pH, H(+) equivalent and lactate content of the incubation medium were measured during anoxic elongation. The effects of fusicoccin (FC), indole-3-acetic acid (IAA), vanadate, erythrosine B and K(+) channel blockers on anoxic elongation were examined. Cytoplasmic pH and vacuolar pH were measured by (31)P nuclear magnetic resonance (NMR) spectroscopy. • Acidification of the incubation medium occurred during anoxic elongation. The contribution of CO(2) and lactic acid was not sufficient to explain the acidification. FC and IAA enhanced the elongation of stem segments. Vanadate and erythrosine B inhibited anoxic elongation. Acid growth of notched segments was observed. The activity of plasma membrane H(+)-ATPase extracted from pondweed turions was increased slightly in anoxic conditions, but that from pea epicotyls sensitive to anoxic conditions was decreased by incubation in anoxic conditions. Both the cytoplasmic pH and vacuolar pH of pondweed turion cells chased by (32)P NMR spectroscopy were stabilized during a short period < 3 h after anoxic conditions. • We propose that the enhancement of H(+) extrusion by anoxic conditions induces acidification in the apoplast and may contribute to the stabilization of pH in the cytoplasm.

Cloning, characterization, and expression of xyloglucan endotransglucosylase/hydrolase and expansin genes associated with petal growth and development during carnation flower opening.

Growth of petal cells is a basis for expansion and morphogenesis (outward bending) of petals during opening of carnation flowers (Dianthus caryophyllus L.). Petal growth progressed through elongation in the early stage, expansion with outward bending in the middle stage, and expansion of the whole area in the late stage of flower opening. In the present study, four cDNAs encoding xyloglucan endotransglucosylase/hydrolase (XTH) (DcXTH1-DcXTH4) and three cDNAs encoding expansin (DcEXPA1-DcEXPA3) were cloned from petals of opening carnation flowers and characterized. Real-time reverse transcription-PCR analyses showed that transcript levels of XTH and expansin genes accumulated differently in floral and vegetative tissues of carnation plants with opening flowers, indicating regulated expression of these genes. DcXTH2 and DcXTH3 transcripts were detected in large quantities in petals as compared with other tissues. DcEXPA1 and DcEXPA2 transcripts were markedly accumulated in petals of opening flowers. The action of XTH in growing petal tissues was confirmed by in situ staining of xyloglucan endotransglucosylase (XET) activity using a rhodamine-labelled xyloglucan nonasaccharide as a substrate. Based on the present findings, it is suggested that two XTH genes (DcXTH2 and DcXTH3) and two expansin genes (DcEXPA1 and DcEXPA2) are associated with petal growth and development during carnation flower opening.