Hara Yoshinao, Yokoyama Ryusuke, Osakabe Keishi, Toki Seiichi, Nishitani Kazuhiko
Department of Developmental Biology and Neurosciences, Graduate School of Life Sciences, Tohoku University, Sendai, 980-8578, Japan.
Center for Collaboration among Agriculture, Industry and Commerce, University of Tokushima, Tokushima, 770-8501 Japan.
Ann Bot. 2014 Oct;114(6):1309-18. doi: 10.1093/aob/mct292. Epub 2013 Dec 19.
Although xyloglucans are ubiquitous in land plants, they are less abundant in Poales species than in eudicotyledons. Poales cell walls contain higher levels of β-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.
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.
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.
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.
尽管木葡聚糖在陆地植物中普遍存在，但在禾本目物种中的含量低于双子叶植物。禾本目植物细胞壁中β-1,3/1,4混合连接葡聚糖和阿拉伯木聚糖的含量高于木葡聚糖。尽管禾本目中木葡聚糖的含量相对较低，但水稻（Oryza sativa）中的木葡聚糖内转糖基酶/水解酶（XTH）基因家族在大小上与双子叶植物拟南芥的相当。这就提出了一个问题，即木葡聚糖是否是水稻XTH基因产物的底物，而这些基因产物的酶活性在很大程度上仍未得到表征。
本研究聚焦于OsXTH19（属于XTH家族的IIIA组，在水稻地上部生长组织中特异性表达），以及另外两个XTH，即作为参照的OsXTH11（I/II组）和OsXTH20（IIIA组），并对三种重组水稻XTH，即OsXTH11、OsXTH20和OsXTH19的酶活性进行了测定。
所有三种OsXTH基因产物都具有木葡聚糖内切水解酶（XEH，EC 3·2·1·151）活性，且OsXTH11同时具有XEH和木葡聚糖内转糖基酶（XET，EC 2·4·1·207）活性。然而，当以葡糖醛酸阿拉伯木聚糖或混合连接葡聚糖作为底物时，这些蛋白质既没有水解酶活性也没有转糖基酶活性。这些结果与组织学观察结果一致，后者表明pOsXTH19::GUS在木葡聚糖免疫反应性存在的组织附近特异性表达。过表达OsXTH19（携带花椰菜花叶病毒35S启动子::OsXTH19 cDNA构建体）或OsXTH19表达受抑制（携带pOsXTH19 RNAi构建体）的转基因水稻品系未表现出明显的表型变化，这表明XTH家族成员之间存在功能冗余和协作，正如在拟南芥中所观察到的那样。
OsXTH20和OsXTH19仅对木葡聚糖起水解酶作用，而OsXTH11仅对木葡聚糖同时表现出水解酶和XET活性。对OsXTH19表达改变的转基因品系的表型分析表明，OsXTH19和相关XTH在水稻生长中发挥冗余作用。
