Modes of action and structure of host-selective pathogenicity factors produced by Phytophthora pathogens

本课题从疫霉菌（Phytophthora sp .）中分离纯化了具有活性的宿主选择性抑制因子-葡聚糖，并对其作用方式进行了分析，得到了以下结果：1)在含Ca2+的蒸馏水中，用接种了虫芽孢悬浮液和孢子发酵液的透析管内液分离得到了一种磷酸葡聚糖（P-SG）。2) P-SG的特征是葡萄糖在18-23处聚合，每分子含有1-4个磷酸，以及-1,3-半葡聚糖酶和磷酸酶敏感的葡聚糖。在不同ph的凝胶过滤柱上对P-SG进行了进一步的HPLC分析，分离得到的P-SG仍然是由几个葡聚糖分子组成的，对它们的结构进行了详细的分析。3)通过在测试组织的自照相上测试电刺激化学发光的减少，设计了一个抑制活性的分析系统。4)发现P-SG可以抑制激发剂刺激的Ca^<2+>从质膜上的运动，从而抑制超氧化物生成系统的激活。5)测定了P-SG与宿主细胞膜表面的分子相互作用，但其结果受到质膜悬浮液中葡聚糖酶和磷酸酶的干扰，需要进一步完善体外体系以分析其分子机制。

In this project, isolation and purification of the active host-selective suppressor-glucans from Phytophthora spp.and analyzes of the their mode of action were carried out and the following results were obtained.1) A phospho-glucan (P-SG) was isolated from inner solution of a dialysis tube on which zoospore-suspension of P.infestrans was inoculated as well as spore-germinated fluid in Ca2+ containing destilled water.2) The P-SG was characterized by glucans with polymerization of glucose at 18-23 and with 1-4 phosphate per molecule and beta-1,3-galucanase- and phosphatase sensitive glucans. The P-SG was still consisted of several glucan molecules since it was further separated through HPLC analyzes on gel filtration columns at different pH.The detailed analyzes of their structures were remained to be carried out.3) A assay system for suppressor activity was devised by testing the reduction of elicitorstimulated chemiluminescence on an autophotography of the test tissued.4) P-SG was found to inhibit the Ca^<2+> movement from plasmamembrane which was stimulated by elicitor treatment and resultantly inhibit the activation of the superoxide generating system.5) A molecular interaction between P-SG and surface of host plasmamembrane was tested, but the results was disturbed by the presence of glucanase and phosphatase in the plasmamembrane suspension, The further improvement of the in vitro systemn was requested for analyzes of the molecular mechanisms of the supperessor.