9316897 West Dictyostelium spore coats contain about one -half polysaccharide, including cellulose and galuronan, and about one half protein, including at least 7 different protein species. The coat is assembled in two phases. First, preformed coat proteins and the galuran are secreted, forming a layer at the cell surface. Most of the proteins exchange between neighboring spores, indicating that assembly occurs extracellularly after secretion. One protein. is unique in that it is not exchanged, as if it anchors or initiates coat assembly. During the second phase cellulose accumulates, and the proteins organize further. Cellulose has traditionally been considered to be the primary determinant of structure and function. The PI proposes two hypotheses to investigate the relative contribution of proteins and polysaccharides. Hypothesis I suggests that proteins play a critical role in initiating assembly of the coat so that the absence of essential groups of proteins such as SP86, the protein part of the coat will not assemble or will not be attached to the plasma membrane, and what accumulates will not be positioned correctly. Hypothesis II suggests that in the absence of cellulose, a microscopically detectable and sedimentable protein pseudocoat will still form.. The proposal has three aims: 1) To disrupt the gene for the spore coat protein SP87; 2) To isolate the gene for the spore coat protein SP86 and disrupt it; and 3) To disrupt the gene for UDPGPP2, and to investigate the effect of this gene on coat structure. %%% Spore coats and other extracellular matrices protect enclosed cells from unfavorable environments. Although the constituents of a number of these matrices is partially known, knowledge of how this parts fit together, how their proportions are regulated, and how protection occurs is largely derived by speculation. The spore coat of the cellular slime mold Dictyostelium is a good system to study matrices at a molecular level, becaus e it is simple it can be isolated in bulk and in high purity, the genes for a number of its proteins and enzymes for polysaccharide synthesis have been cloned and can be disrupted, and good assays exist for the composition. structure and function of the coat. The results of this investigation will enhance our knowledge about the assembly of walls in plant and fungal cells. ***

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University of Florida
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