Analysis of starch-to-sucrose transition mechanism in germinating rice seed and its application

本研究旨在阐明水稻种子萌发过程中淀粉向蔗糖的转化机制，建立种子萌发的调控体系。1.水稻种子萌发过程中α-淀粉酶和UDP-葡萄糖焦磷酸化酶(UGPase)在萌发过程中的时态、特异性和激素调控表达：UGPase在水稻种子萌发的盾片和糊粉组织中为管家酶。α-淀粉酶I-1和II-4在糊粉层中的表达在时间和空间上基本相同，尽管它们在萌发早期的胚胎组织中存在差异。研究发现，GA对α-淀粉酶II-4基因启动子区域缺乏赤霉素顺式元件的转录后调控作用发生在发芽水稻种子糊粉层，该层由钙离子和钙调素介导。组蛋白乙酰化在α-淀粉酶基因转录中的作用：曲古菌素A是组蛋白脱乙酰酶的有效抑制剂，可使未加赤霉素组的α-淀粉酶基因表达水平升高，提示组蛋白乙酰化参与了α-淀粉酶基因表达的调控。盾片组织的蛋白质组学分析：在萌发的水稻种子的盾片组织中，列出了几种具有时间依赖性和脱落酸(ABA)诱导表达的蛋白质。在这些蛋白质中发现了糖酵解酶，如磷酸甘油酸变位酶、甘油醛-3P脱氢酶等。GS-129蛋白具有如下特征：(A)GS-129与细胞壁物质紧密结合，(B)GS-129是一种同时含有N-连接和O-连接糖链的糖蛋白，(C)GS-129可能是一种木聚糖水解酶。

Aim of this research project is to clarify the starch-to-sucrose transition mechanism in germinating rice seed and to establish control system of seed germination. The results obtained in this research period are as follow:1. Temporal, special and hormone-regulated expression of α-amylase and UDP-glucose pyrophosphorylase (UGPase) in germinating rice seeds: UGPase was concluded to be house-keeping enzyme in the scutellar and aleurone tissues. Temporal and spatial expressions of α-amylase I-1 and II-4 were essentially identical in the aleurone layer, although these were distinguishable in the embryo tissues at the early stage of germinatioa It was found that the post-transcriptional regulation of expression of α-amylase II-4, that lacks the gibberellin (GA) response cis-element in the promoter region of the gene, by GA operates in the aleurone layer of germinating rice seed, which is mediated by Ca^<2+> and calmodulin.2. Involvement of histone acetylation in transcription of α-amylase genes: A potent inhibitor for histone deacetylase, Trichostatin A increased the level of α-amylase mRNA in the aleuome without GA, suggesting that the histone acetylation is involved in the regulation of α-amylase gene expression.3. Proteome analysis of scuteller tissues: In the scutellar tissues of germinating rice seeds, several proteins that exhibited the time-dependent and abscisic acid (ABA)-inducible expression were listed up. Glycolitic enzymes, such as phosphoglycerate mutase, glyceraldehydes-3P dehydrogenase, etc., were identified in these proteins. GS-129, one of the listed proteins exhibited the following characteristics: (a) GS-129 tightly associated with cell wall materials, (b) GS-129 was glycoprotein bearing both N-linked and O-linked saccharide chains, (c) GS-129 was suggested to be a possible xylan hydrolyzing enzyme.