Dynamics and Structure of Arabinogalactan-Proteins at the Plasma Membrane-Cell Wall Interface in Plants

阿拉伯半乳糖蛋白（AGPs）和其他糖缀合物在玫瑰细胞质膜和细胞壁之间的界面上的丰度表明，这些分子在这两种不同结构的表面之间起着中介作用。这一假设将在整个细胞和模型系统上进行验证，通过分析AGPs的扰动对质膜组分的横向扩散和质膜与细胞壁之间的线性运动的影响。这些运动类型的特征将被确定为膨胀或施加压力的函数。AGPs的扰动包括聚集、酶促裂解和合成改变。这样的表征将提供关于膨胀和AGPs对膜动力学的影响的进一步信息。我们将进一步纯化能够在活的原生质体上切割质膜AGPs的酶，并阐明它们的作用模式。还将分析质膜AGPs的生化结构以及在分离过程中与质膜AGPs共硫化的分子。质膜AGPs的结构分析将包括确定糖基连接组成，主要的端粒构型和氨基末端序列。这些生化研究将为理解AGPs在发育过程中作为中介剂或/和特异性识别分子的作用提供重要信息。植物细胞壁是复杂的细胞外基质结构，含有蛋白质和多糖成分，其详细组织结构在很大程度上是未知的。由于细胞的高水压状态（称为膨涨），质膜必须以大约7个大气压的压力紧紧地压在细胞壁上。然而，膜的组成部分必须能够自由地进行快速的横向扩散，即使是贴在壁上。在细胞生长过程中尤其如此。在质膜中发现了许多蛋白质连接的糖缀合物，其中的碳水化合物部分位于细胞外部，并延伸到膜和细胞壁之间。其中一些类似于动物细胞膜的蛋白多糖。它们的功能是完全未知的。该项目将探索其中一组糖缀合物阿拉伯半乳聚糖的功能和结构方面，并假设阿拉伯半乳聚糖可能作为植物细胞膜和细胞壁之间的物理介质或缓冲。

The abundance of arabinogalactan-proteins (AGPs) and other glycoconjugates at the interface between the plasma membrane and the cell wall of rose cells suggests that these molecules serve as mediating agents between the surfaces of these two distinct structures. This hypothesis will be tested on whole cells and model systems by analyzing the effect of perturbations of AGPs on both lateral diffusion of plasma membrane components and on linear motion between the plasma membrane and cell wall. Characteristics of these types of motions will be determined as a function of turgor or applied pressure. Perturbations of AGPs will include aggregation, enzymatic cleavage, and altered synthesis. Such characterization will provide further information on the effects of both turgor and AGPs on membrane dynamics. Enzymes capable of cleaving plasma membrane AGPs on live protoplasts will be further purified and their mode of action elucidated. Analysis of the biochemical structure of plasma membrane AGPs and molecules that copurify with plasma membrane AGPs during isolation will also be conducted. Structural analysis of plasma membrane AGPs will include determination of glycosyl linkage composition, predominant anomeric configurations, and amino terminal sequence. The biochemical studies will provide important information towards understanding the role of AGPs as mediating agents or/and as specific recognition molecules during development. The cell walls of plants are complex extracellular matrix structures containing protein and polysaccharide components, the detailed organization of which is largely unknown. As a result of the cellular high water pressure state known as turgor, the plasma membrane must be tightly appressed to the wall by about 7 atmospheres of pressure. However, the components of the membrane must be free to undergo rapid lateral diffusion, even while appressed against the wall. This is particularly true during cell growth. A number of protein-linked glycoconjugates have been found in the plasma membrane, the carbohydrate portion of which is external to the cell and extends between the membrane and the wall. Some of these are analogous to proteoglycans of animal cell membranes. The functions of these are totally unknown. This project will explore functional and structural aspects of one group of these glycoconjugates, the arabinogalactans, with the hypothesis in mind that the arabinogalactans may serve as physical mediators or cushions between the membrane and the wall of plant cells.