ROLE OF PROTEIN GLYCOSYLATION IN DEVELOPMENT

盘基网柄菌细胞黏菌modB基因突变 产生蛋白质连接的聚糖，这些聚糖似乎缺乏大部分原始的 马萨诸塞州 在该基因座突变等位基因纯合的二倍体菌株中， 还观察到：1）假定的接触位点A糖蛋白（gp80） 减少到ca。正常水平的10%，2）分离的 聚集期细胞明显减少，3）子实体变小 形成。 另外两种糖蛋白的表达， 缺失的碳水化合物表位似乎在 免疫荧光和SDS凝胶分析的分辨率。 基于这些 研究结果，我们假设减少gp80在细胞表面的表达， 导致解离后细胞再结合减少， 子实体大小。 我们将在以下背景下研究这一假设： 更大的问题是聚糖在体内的积累和定位 发育调节糖蛋白。 我们将首先调查 gp80水平下降的依据，使用合成和周转研究 和EM定位方法。 然后我们会产生新的突变， 改变modB依赖性聚糖，检查这些修饰， 单糖组成、聚糖大小分布和互补 分析然后测试对gp80表达、细胞 再结合和子实体大小（孢子数）。 我们推测， 恢复一部分缺失的聚糖，或通过仅除去一部分聚糖， 通过突变的方法，我们应该能够分离modB的这些影响， 对gp80积累和细胞再结合的影响， 结构基础。 如果我们无法将这些影响与 碳水化合物结构的细微变化，我们将推断它们在 由于共同的机制（采用共同的识别过程）而产生的事实 其防止GP 80到达小区 表面（或在到达时将其移除）。 最后，聚糖参数 携带碳水化合物表位的其它两种糖蛋白将被 与gp80的这些比较，以寻求对不同的解释， modB对其积累的影响。 我们将比较modB依赖性聚糖与另一种聚糖的作用， 表达在孢子外壳蛋白上，其携带被 另一组单克隆抗体。 使用为 modB依赖性聚糖，我们将研究这种独特的聚糖的作用， 在将孢子外壳蛋白分配到前孢子囊泡中（其 胞吐作用受到发育调节）并且，分泌后， 将孢子外壳蛋白定位到孢子外壳、粘液鞘中， 茎管。

The modB mutation in the cellular slime mold Dictyostelium discoideum produces protein-linked glycans which appear to lack most of their original mass. In diploid strains homozygous for mutant alleles at this locus, it is also observed that 1) the putative contact sites A glycoprotein (gp80) is reduced to ca. 10% of its normal level, 2) reassociation of dissociated aggregation stage cells is markedly reduced, and 3) smaller fruiting bodies are formed. The expression of two other glycoproteins which normally bear the missing carbohydrate epitope do not appear to be affected at the level of resolution of immunofluorescence and SDS gel analysis. Based on these findings, we hypothesize that reduced gp80 expression on the cell surface results in reduced cell reassociation following dissociation and in reduced fruiting body size. We will investigate this hypothesis in the context of the larger question of glycan function in the accumulation and localization of developmentally-regulated glycoproteins. We will first investigate the basis for the decreased level of gp80, using synthesis and turnover studies and EM localization methods. We will then generate new mutations which alter the modB-dependent glycan, examine these modifications in terms of monosaccharide composition, glycan size distribution, and complementation analysis and then test for coordinate effects on gp80 expression, cell reassociation and fruiting body size (spore number). We speculate that by restoring a part of the missing glycan, or by removing only a part of it, by mutational means, we should be able to dissociate these effects of modB on gp80 accumulation and cell reassociation if they have a different structural basis. If we are unable to to dissociate the effects with even subtle changes in carbohydrate structure, we will infer that they are in fact due to a common mechanism (employing a common recognition process based on common structure) which prevents gp80 from arriving at the cell surface (or removes it once it arrives). Finally, glycan parameters for the other two glycoproteins carrying the carbohydrate epitope will be compared with these for gp80 to seek an explanation for the different effects of modB on their accumulation. We will compare the role of the modB-dependent glycan with another glycan, expressed on spore coat proteins, which carries an epitope recognized by another group of monoclonal antibodies. Using methods worked out for the modB-dependent glycan, we will investigate the role of this distinct glycan in distributing the spore coat proteins into prespore vesicles (whose exocytosis is developmentally regulated) and, following secretion, localizing the spore coat proteins into the spore coat, slime sheath and stalk tube.