New elements of the Target of Rapamycin pathway in Candida albicans

描述（由申请人提供）：该提案有两个目标：有助于我们理解真菌条件致病菌C中Tor信号传导的生物学。白色念珠菌，并确定与雷帕霉素类似物协同抗真菌活性的潜在药物靶标。Tor途径促进真核生物的生长和增殖，以响应营养物质的可用性和有害应激物的缺乏。这一途径对许多基本细胞过程的中心作用已经从模型酵母和后生动物的工作中显现出来。In C.白念珠菌发病机制中，Tor信号传导被预测发挥关键作用，原因有三个。首先，Tor通路共调节C.白色念珠菌其次，真菌在宿主吞噬细胞中遇到饥饿条件;第三，宿主免疫系统施加有害应激。Tor途径也很重要，因为它被原核产物雷帕霉素特异性抑制，为开发用于治疗念珠菌感染的真菌特异性雷帕霉素类似物打开了大门。C.白色念珠菌和模式酵母的差异已经变得明显。为此，提出了一种无偏的正向遗传方法来发现新的C。白色念珠菌Tor信号传导的模式生物未知，并确定新的目标抑制小分子，可以增强雷帕霉素类似物的杀伤活性。我们建议筛选我们的杂合转座子突变体集合，因为使用这种试剂，可以鉴定参与Tor信号传导的必需基因。该集合的另一个优点是杂合子显示的单倍不足表型在相应的纯合子中更可能是严重的，从而使筛选偏向于仅与Tor间接相关的具有轻度影响的突变。使用该突变体集合的试验性筛选产生了我们两个目标中的每一个的代表：分离出激酶调节剂的同源物，其在后生动物免疫系统的发育中发出信号，因为它是雷帕霉素敏感性所必需的。其次，在一个假定的质膜离子泵的突变体被发现是精致的雷帕霉素过敏。该离子泵没有人类同源物。其保守的S.酿酒酵母同源基因被两种不同的小分子所抑制，这支持了可以发现更多抑制剂的想法。白念珠菌Tor信号传导，并可能为协同杀念珠菌药物的小分子筛选奠定基础。 公共卫生相关性：白色念珠菌是侵袭性真菌感染的最常见原因，即使治疗得当，死亡率也很高。这项工作将确定一个新的组成部分，中央信号通路所需的生长和增殖的C。白色念珠菌，并表征协同抗真菌治疗的潜在药物靶点。

DESCRIPTION (provided by applicant): This proposal has two goals: contribute to our understanding of the biology of Tor signaling in the fungal opportunistic pathogen C. albicans, and identify potential drug targets for synergistic antifungal activity with rapamycin analogs. The Tor pathway promotes growth and proliferation in eukaryotes in response to availability of nutrients and absence of noxious stressors. The centrality of this pathway to many essential cellular processes has emerged from work in model yeasts and in metazoans. In C. albicans pathogenesis, Tor signaling is predicted to play a crucial role for three reasons. Firstly, the Tor pathway co-regulates morphogenesis in C. albicans. Secondly, the fungus encounters starvation conditions in host phagocytes; and thirdly, noxious stresses are imposed by the host immune system. The Tor pathway is also important because it is specifically inhibited by the prokaryotic product rapamycin, opening the door to development of fungal-specific rapamycin analogs for treatment of Candida infections. Partial divergence between the Tor pathway of C. albicans and that of model yeasts has already become apparent. For this reason, an unbiased forward genetic approach is proposed to discover novel components of C. albicans Tor signaling not known from model organisms, and to identify new targets for inhibition by small molecules that can potentiate the cidal activity of rapamycin analogs. We propose a screen of our heterozygous transposon mutant collection because with this reagent, essential genes involved in Tor signaling can be identified. Another advantage of this collection is that haploinsufficiency phenotypes, displayed by heterozygotes, are more likely to be severe in the corresponding homozygotes, biasing the screen against mutations with mild effects only indirectly linked to Tor. A pilot screen using this mutant collection yielded representatives of each of our two goals: a homolog of a kinase regulator, which signals in development of the metazoan immune system, was isolated because it is required for rapamycin sensitivity. Secondly, a mutant in a putative plasma membrane ion pump was found to be exquisitely rapamycin hypersensitive. This ion pump has no human homolog. Its conserved S. cerevisiae homolog has been shown to be inhibited by two different small molecules, supporting the idea that further inhibitors can be found. Characterizing these genes will contribute to our basic understanding of C. albicans Tor signaling and possibly lay the groundwork for a small molecule screen for synergistic candidacidal drugs. PUBLIC HEALTH RELEVANCE: Candida albicans is the most common cause of invasive fungal infections, which even treated appropriately, carry a high mortality. This work will identify new components of a central signaling pathway required for growth and proliferation of C. albicans, and characterize a potential drug target for synergistic antifungal therapy.