Tonoplast transport as a determinant of tomato fruit chemical composition

液泡膜运输是番茄果实化学成分的决定因素

基本信息

批准号：
BB/H00338X/1
负责人：
Lee Sweetlove
金额：
$ 74.51万
依托单位：
University of Oxford
依托单位国家：
英国
项目类别：
Research Grant
财政年份：
2010
资助国家：
英国
起止时间：
2010 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=BB%2FH00338X%2F1
关键词：
Tonoplast transport determinant tomato fruit

项目摘要

The aim of this proposal is to investigate factors controlling the chemical composition of tomato fruit, a crop of major economic importance worldwide. Both the flavour and nutritional quality of tomatoes are determined by the chemicals that accumulate during fruit ripening, yet we have only a limited understanding of how this process is controlled. In mature fruit, the cells are dominated by a compartment called the central vacuole, which contains most of the sap in fleshy fruit. This compartment can occupy as much as 95 % of the cell's volume, the remaining 5 % being taken up by the cell cytoplasm and outlying cell wall. As the tomato fruit grows, chemicals such as sugars, organic acids and amino acids are produced in the cytoplasm. They are then removed from their site of synthesis by transport into the central vacuole across the bounding membrane surrounding this compartment, called the tonoplast. But this traffic is not all one-way. As the fruit ripens, some solutes leave the vacuole to be re-metabolized in the cytoplasm, with other solutes moving back into the vacuole to compensate. Thus, the composition of the mature fruit is a complex outcome of metabolic events in the cytoplasm combined with transport of solutes across the tonoplast membrane. Whereas the pathways of basic metabolism in fruit cells are well understood, we have much less knowledge of the transport proteins that reside in the tonoplast membrane. In fact, we have indirect evidence that these proteins may play a much more important role in determining fruit composition than previously suspected. As the first part of this project, therefore, we shall isolate the tonoplast membrane from tomato fruit at defined stages during their development and analyse its protein content by mass spectrometry. This will provide a valuable inventory of proteins residing in the tonoplast membrane, and of their changes in abundance during the ripening process. By correlating these changes with the chemical composition of the fruit, we should obtain the first clues as to which tonoplast proteins are important in regulating transport across the vacuolar membrane. In another strand of the project, we will use a genetic approach to obtain independent information on factors controlling fruit composition. A powerful resource for this purpose is provided by the natural genetic variation found between cultivated tomatoes and their close relatives in the wild. Indeed, several of these species are sufficiently closely related that they can be hybridized. By analysing the characteristics of the progeny of such crosses (e.g. with respect to fruit composition), it is possible to make deductions about which genes may be contributing to particular traits. Using this approach, we will investigate whether any of the genes correlated with differences in fruit composition encode likely tonoplast membrane proteins. If they do, we will cross-reference this list against the information on tonoplast proteins obtained by mass spectrometry. This will allow us to focus on a limited number of the most promising candidates for more detailed characterization. In the final part of the project, we will test the function of the selected candidate proteins directly to determine, first, whether they indeed reside in the tonoplast membrane in intact cells, and second, what solutes they are capable of transporting into and out of the vacuole. We will focus on candidate transporters of organic acids and amino acids, as these are important determinants of fruit flavour and acidity that have been little investigated to date. The combination of the protein identification and genetic approaches promises to yield important new information on the factors determining fruit composition. This will also be valuable for directing future breeding strategies towards the selection of new elite lines with improved fruit traits, without the need for intervention using genetic modification techniques.

该提议的目的是调查控制番茄水果化学成分的因素，番茄水果的化学成分是全球重要的经济重要性。西红柿的风味和营养质量都取决于果实成熟过程中积累的化学物质，但我们对如何控制该过程的理解只有有限的了解。在成熟的果实中，这些细胞由一个称为中央液泡的隔室支配，其中包含大部分的肉质水果中的汁液。该隔室可占用该细胞体积的95％，其余5％被细胞细胞和外围细胞壁吸收。随着番茄果的生长，糖，有机酸和氨基酸等化学物质会在细胞质中产生。然后，通过将其转运到该隔室周围的边界膜中的中央液泡中，将它们从合成部位移出，称为托管。但是这种流量并非全部。随着果实成熟的成熟，一些溶液会使液泡在细胞质中重新代谢，而其他溶质移回了液泡中以补偿。因此，成熟果实的组成是细胞质中代谢事件的复杂结果，结合溶质在整个块状膜上的转运。尽管人们对水果细胞中碱性代谢的途径的了解很清楚，但我们对居住在块状膜中的转运蛋白的了解要少得多。实际上，我们有间接的证据表明，这些蛋白质在确定果实中可能起着比以前怀疑的重要作用更为重要。因此，作为该项目的第一部分，我们将在开发过程中在定义的阶段与番茄果实分离骨头膜，并通过质谱分析其蛋白质含量。这将提供驻留在块状膜上的蛋白质的宝贵库存，并在成熟过程中的丰度变化。通过将这些变化与果实的化学组成相关联，我们应该获得第一个线索，即在调节整个液泡膜的转运中，哪种块状蛋白很重要。在该项目的另一个方面，我们将使用一种遗传方法来获取有关控制水果组成的因素的独立信息。为此目的的有力资源是由在野外培养的西红柿及其近亲之间发现的自然遗传变异提供的。实际上，其中几个物种足够密切，可以杂交它们。通过分析此类十字的后代的特征（例如，关于果实成分），可以对哪些基因可能有助于特定性状进行推论。使用这种方法，我们将研究任何基因是否与果实成分的差异相关，并可能编码可能的块状膜蛋白。如果这样做，我们将根据质谱法获得的有关块状蛋白的信息进行交叉引用。这将使我们能够专注于有限数量的最有前途的候选人，以进行更详细的特征。在项目的最后一部分中，我们将直接测试所选候选蛋白的功能，首先确定它们是否确实驻留在完整细胞中的Tonoplast膜中，其次，它们能够将它们能够运输到液泡中。我们将重点关注有机酸和氨基酸的候选转运蛋白，因为这些是迄今为止尚未研究的水果风味和酸度的重要决定因素。蛋白质鉴定和遗传方法的组合有望产生有关决定果实组成的因素的重要新信息。这对于将未来的育种策略引导到具有改进的水果特征的新精英线的选择也将是有价值的，而无需使用遗传修饰技术进行干预。