Biological Fusions Conjugations in Yeast

描述（由申请方提供）：本提案的目的是了解酵母菌丝转换与真菌毒力之间的联系。这个开关与包围真菌细胞的分子、β-葡聚糖和连接的甘露糖蛋白（粘附素）密切相关。对酿酒酵母模型系统中表达的分析将指导对较难处理的病原体白念珠菌的研究。这两种真菌的基因组编码许多甘露糖蛋白，赋予独特的粘附特性。这些粘附素是真菌细胞相互作用（絮凝和沉淀）、与惰性表面（琼脂和塑料）以及与哺乳动物细胞相互作用所必需的。一种新的反义IME 4 RNA在控制粘附素的表达中的作用将在两种生物体中确定。酪醇是一种自动调节分子，它在触发酵母形态和丝状体形态之间的转换中的作用，以及它在真菌与吞噬细胞相互作用中的作用，将通过分析其生物合成、感知和信号转导作用来解决。巨噬细胞区分酵母菌属和念珠菌属的能力可能是由于每种生物体表面上的β-葡聚糖对巨噬细胞上的关键非调理素真菌受体Dectin-1的可及性不同。初步实验表明，真菌细胞表面上的差异β-葡聚糖呈递导致真菌与Dectin-1的差异结合，并导致从巨噬细胞中不同地诱发炎性细胞因子。通过系统地使用全基因组突变体文库来鉴定β-葡聚糖呈递改变的突变体，并使用这些突变体来鉴定免疫应答，将解决β-葡聚糖的暴露或β-葡聚糖的结构是否是关键的问题。设计额外的实验以鉴定甘露糖蛋白转运到真菌细胞壁所需的所有基因。我们的发现，缺氧诱导假丝酵母菌的毒力将被用来分析基因的重要性，在低氧张力。在我们的分析中发现的基因是开发针对真菌的新疗法的潜在靶点，真菌对那些接受化疗和患有艾滋病的人来说是如此的毁灭性。

DESCRIPTION (provided by the applicant): The goal of this proposal is to understand the connection between the yeast to filament switch and fungal virulence. This switch is intimately connected to the molecules that encircle the fungal cell, beta-glucan and the linked mannoproteins (adhesins). The analysis of filamentation in the model system, Saccharomyces cerevisiae will guide the studies in the less tractable pathogen, Candida albicans. The genomes of both fungi encode many mannoproteins that confer unique adherence properties. These adhesins are required for interactions of fungal cells with each other {flocculation and filamentation), with inert surfaces (agar and plastic) and with mammalian cells. The role of a novel antisense IME4 RNA in controlling the expression of the adhesins will be determined in both organisms. The role of tyrosol, an autoregulatory molecule in triggering the switch between the yeast form and the filament form as well as its role in the interaction of fungi with phagocytic cells will be resolved by analysis of its biosynthesis, perception and role in signal transduction. The ability of macrophages to distinguish between Saccharomyces and Candida is likely to result from differential accessibility of beta-glucan on the surface of each organism to Dectin-1, the key non-opsonic fungal receptor on macrophages. Preliminary experiments show that differential beta-glucan presentation on the fungal cell surface leads to differential binding of fungi to Dectin-1 and to different elicitation of inflammatory cytokines from macrophages. The question of whether the unmasking of beta-glucan or the structure of beta-glucan is key will be resolved by systematic use of whole genome mutant libraries to identify mutants with altered beta-glucan presentation and the use of those mutants to identify the immune response. Additional experiments are designed to identify all the genes required for the mannoproteins to transit to the fungal cell wall. Our finding that hypoxia induces filamentation in Candida will be used to analyze the genes important for virulence under low oxygen tensions. The genes uncovered in our analyses are potential targets for the development of novel therapeutics against fungi, which are so devastating to those undergoing chemotherapy and afflicted with AIDS.