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Molecular biological approach to improve the sink and source function in Rosaceae fruit tree

改善蔷薇科果树库源功能的分子生物学方法

基本信息

批准号：
09306002
负责人：
YAMAKI Shohei
金额：
$ 22.34万
依托单位：
NAGOYA UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (A).
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 2000
项目状态：
已结题

项目摘要

The purpose of this research is to make clear the sink-source relation of fruit by investigating the properties and gene expression of enzymes which play an important role for sugar metabolism and the vacuolar function to accumulate sugars.1. Sorbitol dehydrogenase was purified from Japanese pear fruit, and its cDNA was also cloned from fruits of apple, loquat and peach. The gene expression was investigated throughout the development of their fruits.2. The gene of sorbitol-6-phosphate dehydrogenase expressed on source leaves unfolded enough, but not on leaves folded yet.3. Sucrsoe synthase isoform appearing with sugar accumulation was purified and the reaction inclined to sucrose synthesis rather than sucrose cleavage. The direction of reaction changes easily by phosphorylation and non-phosphorylation.4. Two soluble acid invertases were purified from Japanese pear fruit, and there was no difference between properties of both isoforms.5. We isolated the cDNA clone of fructokinase which is closely relating to fructose accumulation and examined the relations between the gene expression and fructose metabolism.6. H^+-ATPase and H^+-pyrophosphatase which supply the energy needed to uptake sugars into vacuole were purified from pear fruit, and the gene expression throughout fruit development were investigated using cDNA clone of their subunits.7. We could prove the presence of sugar transporters across the tonoplast by sugar uptake experiment, although could not isolate their protein and cDNA.As described above, we could obtain the proteins and cDNA of some enzymes needed to accumulate sugars into fruit, analyzed their gene expressions and clarified partially the melecular mechanism of sugar accumulation.

本研究的目的是通过对果实糖代谢和液泡积累功能中起重要作用的酶的特性和基因表达的研究，明确果实的库源关系。从日本梨果中分离纯化了山梨醇脱氢酶，并从苹果、枇杷和桃子果实中克隆了其cDNA。在其果实发育过程中对基因表达进行了研究。山梨糖醇-6-磷酸脱氢酶基因在源叶上的表达充分展开，但在折叠叶上表达不足。蔗糖合成酶同工型出现糖积累，反应倾向于蔗糖合成而不是蔗糖裂解。磷酸化和非磷酸化很容易改变反应方向。从日本梨果中纯化了两种可溶性酸转化酶，两种酶的性质没有差异。我们分离了与果糖积累密切相关的果糖激酶cDNA克隆，并研究了该基因表达与果糖代谢的关系。从梨果实中纯化出了向液泡吸收糖提供能量的H^+- atp酶和H^+-焦磷酸酶，并利用其亚基cDNA克隆研究了它们在果实发育过程中的表达。通过糖摄取实验，我们可以证实糖转运体的存在，但无法分离出糖转运体的蛋白和cDNA。如上所述，我们可以获得果实积累糖分所需的一些酶的蛋白质和cDNA，分析它们的基因表达，部分阐明糖积累的分子机制。