Regulation of Alginate Gelation During Wall Development in Fucus Embryos

褐藻的受精卵将褐藻酸盐合成并重新组装成细胞壁。墨角藻为研究植物细胞壁形成提供了一个简单的系统。海藻酸盐是两种酸性糖甘露糖酸盐和谷氨酰糖酸盐的线性聚合物，排列在非凝胶和凝胶亚基中。最近的工作确定了胶凝亚基合成的细胞内位点为高尔基体。在特定酶裂解后，通过电泳检测细胞内和细胞外海藻酸盐的分子大小和亚基长度分布。通过已知链长荧光杂交探针标记受精卵片段，研究海藻酸盐受精卵在海水环境中的凝胶化能力。通过酶微分析，两种酶的活性，一种调节海藻酸盐凝胶（epimase将非凝胶亚基转化为凝胶亚基）和一种引起其裂解（裂解酶切割海藻酸盐链以扩大壁）的活性将与壁发育相关。制备纯化酶的抗血清，用于壁发育过程中的免疫金超微结构定位。在发育过程中，由受精卵制备的信使RNA的体外翻译产物将被免疫沉淀。本项目将提供有关胶凝壁碳水化合物的合成、组装、结构和修饰的酶和mRNA控制的信息。本项目将在碳水化合物、蛋白质和mRNA水平上对胶凝植物细胞壁聚合物的控制提供详细的信息。这种凝胶化/非凝胶化使细胞壁成分能够适应生长，同时保持生物体的完整性。

Alginate synthesis and de novo assembly into a cell wall by the fertilized egg of the brown alga. Fucus provides a simple system for studying plant cell wall formation. Alginate is a linear polymer of the two acidic sugars, mannuronate and guluronate, arranged in nongelling and gelling subunits. Recent work identified the intracellular site of synthesis of the gelling subunits as the Golgi apparatus. Alginate molecular size and the subunit length distributions of intracellular and extracellular alginate will be examined by electrophoresis after cleavage by specific enzymes. The ability of zygote alginate to gel in a seawater environment will be studied by labelling zygote sections with fluorescent hybridization probes of known chain length. The activities of two enzymes, one regulating alginate gelation (epimerase converts nongelling subunits to gelling subunits) and one causing its cleavage (lyase cuts alginate chains for wall expansion) will be correlated with wall development by enzyme microassays. Antisera to the purified enzymes will be prepared for immunogold ultrastructural localization during wall development. In vitro translation products of messenger RNA prepared form zygotes during development will be immunoprecipitated. This project will provide information on enzyme and mRNA controls of the synthesis, assembly, structure and modification of a gelling wall carbohydrate. %%% This project will provide detailed information on the control of a gelling plant cell wall polymer at the carbohydrate, protein and mRNA levels of control. This gelling/nongelling allows the cell wall components to adapt to growth while maintaining the integrity of the organism.