Biological Fusions Conjugations in Yeast

描述(由申请人提供)：本提案的目的是了解酵母到丝状物的转换和真菌毒力之间的联系。这种开关与包围真菌细胞的分子、β-葡聚糖和相连的甘露糖蛋白(粘附素)紧密相连。对酿酒酵母模型体系中丝状菌的分析将指导对较难驯化的白色念珠菌的研究。这两种真菌的基因组都编码了许多甘露糖蛋白，这些蛋白具有独特的粘附性。这些粘附素是真菌细胞相互作用(絮凝和丝状化)、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IME4 RNA在控制粘附素表达方面的作用将在这两种生物中得到确定。酪醇是一种自动调节分子，它触发酵母形式和丝状形式之间的转换，以及它在真菌与吞噬细胞相互作用中的作用，将通过对其生物合成、感知和在信号转导中的作用的分析来解决。巨噬细胞区分酵母菌和念珠菌的能力可能是由于每个生物体表面的β-葡聚糖对Dectin-1的不同可及性，Dectin-1是巨噬细胞上关键的非调理真菌受体。初步实验表明，真菌细胞表面呈现的不同的β-葡聚糖导致真菌与Dectin-1的不同结合，以及从巨噬细胞中不同地激发炎症细胞因子。是否揭开β-葡聚糖或β-葡聚糖的结构是关键，这一问题将通过系统地使用全基因组突变文库来识别具有改变的β-葡聚糖提呈的突变体并使用这些突变体来识别免疫反应来解决。此外，还设计了一些实验，以确定甘露糖蛋白转运到真菌细胞壁所需的所有基因。我们的发现是低氧诱导假丝酵母菌的丝状化，这一发现将被用于分析在低氧压下对毒力重要的基因。在我们的分析中发现的基因是开发抗真菌新疗法的潜在目标，真菌对那些正在接受化疗并患有艾滋病的人来说是如此具有破坏性。

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.