The Role of Protein-Protein Interactions in Sucrose Transport

植物有机体的一个决定性特征是蔗糖的光合作用生产。从本质上讲，大气中的碳被吸收成蔗糖，然后被输送到整个植物中，在那里它被用作能源。植物的生产力或产量直接取决于碳转化为蔗糖的净吸收，以及最终有多少蔗糖被输送到植物的可收获部分，如种子。了解蔗糖是如何在植物中运输的，以及这种运输是如何被调节的，对于理解基本的植物生物学和优化植物生产力都是至关重要的。大豆子叶中的蔗糖转运蛋白似乎与植物细胞质中的另一种蛋白相互作用。这种相互作用的蛋白包含至少三个锚蛋白重复基序，这些基序通常参与将膜转运蛋白连接到细胞骨架。由于蔗糖转运体与其他蛋白质相互作用，这种相互作用可能在细胞表面蔗糖转运体的组装或蛋白质复合物的形成/分解中很重要，这些蛋白质复合物既进口蔗糖又进一步代谢蔗糖。因此，蔗糖转运可能部分受到转运体和这种相互作用伙伴之间相互作用的性质的调节。本研究将试图阐明这种蛋白-蛋白相互作用在蔗糖转运中的功能意义。三个主要目标如下。目标# 1。鉴定与锚蛋白样蛋白相互作用的蛋白质。为了了解这种相互作用的本质，确定其他蛋白质是否也可能与植物蔗糖转运蛋白接触是很重要的。例如，如果蔗糖代谢酶是蔗糖转运体复合体的一部分，那么蔗糖的进口可能与其随后的代谢直接相关。这些实验将解决这种可能性。目标# 2。评估蛋白质-蛋白质相互作用对蔗糖摄取的作用。如果蔗糖转运蛋白与锚蛋白样蛋白的相互作用对植物的蔗糖摄取有影响，那么锚蛋白样蛋白功能的破坏应该以不同的方式破坏蔗糖转运。使用基因沉默技术和评估蔗糖摄取参数来破坏锚蛋白样蛋白的功能。目标# 3。检查蔗糖转运蛋白和锚蛋白样蛋白的亚细胞定位。几个重要的问题是解决表征性质的蔗糖转运：锚蛋白样蛋白（ST:ALP）在细胞中的相互作用。ST:ALP复合体在子叶发育过程中是否发生变化？ST:ALP复合物在什么时候形成相对于蔗糖转运体和ankryin样蛋白的合成？我们的mRNA分析表明，在子叶发育过程中，锚蛋白样蛋白比蔗糖转运蛋白积累得更早，这就提出了一个问题，即蔗糖转运蛋白在翻译过程中是否与锚蛋白样蛋白结合。免疫细胞化学分析的蔗糖转运蛋白，锚蛋白样蛋白，和其他潜在的相互作用蛋白共聚焦和电子显微镜将解决这些问题。

One defining feature of plant organisms is the photosynthetic production of sucrose. Essentially, carbon from the atmosphere is assimilated into sucrose which is then transported throughout the plant where it is used as an energy source. The productivity, or yield, of plants directly depends on the net assimilation of carbon into sucrose and ultimately how much of this sucrose is transported to the harvestable components of plants, such as the seed. Understanding how sucrose is transported through the plant and how this transport is regulated is critical for both for understanding fundamental plant biology and optimization of plant productivity.The sucrose transport protein from soybean cotyledons appears to interact with another protein in the cytosol of plant cells. This interacting protein contains at least three ankyrin repeat motifs which often are involved in linking membrane transport proteins to the cellular cytoskeleton. Since the sucrose transporter interacts with other proteins, it may be that this interaction is important in the assembly of sucrose transporters on the cell surface or in the formation/disassembly of protein complexes that both import sucrose and metabolize it further. Thus, sucrose transport may be regulated in part by the nature of interactions between the transporter and this interacting partner.This research will attempt to elucidate the functional significance of this protein-protein interaction in sucrose transport. The three primary objectives are as follows.Objective #1. Identify proteins that interact with the ankyrin-like protein. To understand the nature of this interaction, it is important to determine if other proteins might also be brought into contact with the plant sucrose transporter. For instance, if sucrose metabolizing enzymes are part of a complex with the sucrose transporter, then sucrose import may be directly linked to its subsequent metabolism. These experiments will address this possibility.Objective #2. Assess the role of protein-protein interaction on sucrose uptake. If the interaction of the sucrose transporter with the ankyrin-like protein has an impact on sucrose uptake in the plant, then disruption of the ankyrin-like protein function should disrupt sucrose transport in distinct ways. The ankyrin-like protein function will be disrupted using gene-silencing technology and sucrose uptake parameters assessed.Objective #3. Examine the subcellular localization of the sucrose transporter and the ankyrin-like protein. Several important questions are addressed by characterizing the nature of the sucrose transporter:ankyrin-like protein (ST:ALP) interaction in the cell. Does the ST:ALP complex change during cotyledon development? When does the ST:ALP complex form relative to synthesis of the sucrose transporter and the ankryin-like protein? Our mRNA analysis suggests that the ankyrin-like protein accumulates earlier in cotyledon development than does the sucrose transporter and this raises the question as to whether sucrose transporter is bound to the ankyrin-like protein as it is translated. Immunocytochemical analysis of the sucrose transporter, the ankyrin-like protein, and other potential interacting proteins confocal and electron microscopy will address these issues.