Eugene A. Nothnagel - US grants

A class of macromolecules known as arabinogalactan-proteins (AGPs), that are present both at the outside surface of the plasma membrane and in the cell wall space of plant cells, will be investigated. Because AGPs have the ability to bind carbohydrates and are localized on the surface of the plasma membrane, they may play an important role at the membrane- cell wall interface in terms of cellular responses. The main objectives of the investigation are to compare the structure of AGPs on the cell wall with that of AGPs in plasma membrane, to study AGP interaction with some antigens, to characterize an enzyme that destroys the carbohydrate binding properties of plasma membrane AGPs, and to characterize the interaction between the AGPs and the plasma membrane with those in the cell wall. The project will increase our knowledge about the structural components of plasma membrane in plant cells and interaction of those components with the cell wall. Knowledge gained from this could eventually lead to a better understanding of cell adhesion, water retention, disease resistance and cell-cell recognition in plant cells.

Financial support is requested for acquisition of a mass selective detector to be used with an existing gas chromatograph. The resulting gas chromatograph-mass spectrometer combination would be used in structure determination of complex biological molecules in plant extracts. Applications in structure determination include studies of the complex carbohydrate portion of plasma membrane arabinogalatan-proteins and studies of possible non-amino acid constituents of oil body proteins. Applications in detection and quantitation include measurements of endogenous plant hormones in plants exposed to drought or chilling stress and in seeds during germination. These projects require capabilities of sensitive and quantitative chemical identification that cannot be achieved without a mass selective detector.

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.

9808309 Nothnagel This is the 20th Annual Symposium in Plant Physiology entitled "Cell and Developmental Biology of Arabinogalactan-Proteins," to be held on January 21-23, 1999 at the University of California, Riverside. This conference will provide researchers the opportunity to share research findings on Arabinogalactan molecules that are a class of complex macromolecules found on plant cell surfaces. Recent research indicates that these molecules serve important roles in plant growth and development. This meeting will bring together researchers in the field and to provide researchers the opportunity to establish future collaborative research projects. The meeting will involve a combination of oral and poster presentations. This meeting is unique in that it is the first meeting that will be focused specifically on plant arabinogalactan molecules.