Kaida Rumi, Satoh Yumi, Bulone Vincent, Yamada Yohko, Kaku Tomomi, Hayashi Takahisa, Kaneko Takako S
Department of Chemical and Biological Sciences, Japan Women's University, Tokyo 112-8681, Japan.
Plant Physiol. 2009 Aug;150(4):1822-30. doi: 10.1104/pp.109.139287. Epub 2009 Jun 3.
Wall-bound purple acid phosphatases have been shown to be potentially involved in the regulation of plant cell growth. The aim of this work was to further investigate the function of one of these phosphatases in tobacco (Nicotiana tabacum), NtPAP12, using transgenic cells overexpressing the enzyme. The transgenic cells exhibited a higher level of phosphatase activity in their walls. The corresponding protoplasts regenerating a cell wall exhibited a higher rate of beta-glucan synthesis and cellulose deposition was increased in the walls of the transgenic cells. A higher level of plasma membrane glucan synthase activities was also measured in detergent extracts of membrane fractions from the transgenic line, while no activation of Golgi-bound glycan synthases was detected. Enzymatic hydrolysis and methylation analysis were performed on the products synthesized in vitro by the plasma membrane enzymes from the wild-type and transgenic lines extracted with digitonin and incubated with radioactive UDP-glucose. The data showed that the glucans consisted of callose and cellulose and that the amount of each glucan synthesized by the enzyme preparation from the transgenic cells was significantly higher than in the case of the wild-type cells. The demonstration that callose and cellulose synthases are activated in cells overexpressing the wall-bound phosphatase NtPAP12 suggests a regulation of these carbohydrate synthases by a phosphorylation/dephosphorylation process, as well as a role of wall-bound phosphatases in the regulation of cell wall biosynthesis.
细胞壁结合型紫色酸性磷酸酶已被证明可能参与植物细胞生长的调控。这项工作的目的是利用过表达该酶的转基因细胞，进一步研究烟草（Nicotiana tabacum）中其中一种磷酸酶NtPAP12的功能。转基因细胞在其细胞壁中表现出更高水平的磷酸酶活性。再生细胞壁的相应原生质体表现出更高的β-葡聚糖合成速率，并且转基因细胞的细胞壁中纤维素沉积增加。在转基因系膜组分的去污剂提取物中也检测到更高水平的质膜葡聚糖合酶活性，而未检测到高尔基体结合型聚糖合酶的激活。对用洋地黄皂苷提取并用放射性UDP-葡萄糖孵育的野生型和转基因系的质膜酶体外合成的产物进行了酶促水解和甲基化分析。数据表明，葡聚糖由胼胝质和纤维素组成，并且转基因细胞的酶制剂合成的每种葡聚糖的量明显高于野生型细胞。在过表达细胞壁结合型磷酸酶NtPAP12的细胞中胼胝质和纤维素合酶被激活的证明表明，这些碳水化合物合酶通过磷酸化/去磷酸化过程受到调控，以及细胞壁结合型磷酸酶在细胞壁生物合成调控中的作用。
