CAREER: Cellular Mechanisms of Killer Toxin Resistance in Yeasts

该奖项全部或部分由2021年美国救援计划法案（公法117-2）资助。真菌细胞通过构建由碳水化合物和蛋白质组成的坚固的细胞壁与底层膜将自己与环境分离。这使得代谢过程能够区室化，并保护真菌细胞免受外部压力，包括广泛用于控制不必要的真菌生长的商业杀真菌剂。由不同种类的真菌产生的抗真菌蛋白可以攻击细胞壁和细胞膜，导致易感真菌细胞的损伤和死亡。这些抗真菌剂已被引用为潜在地可用于控制不期望的真菌的生长。本研究将探讨真菌抵抗抗真菌蛋白中毒的基本细胞机制。具体来说，该项目将测试用于细胞壁和膜构建和稳定性的基因突变如何导致抗真菌蛋白的耐药性。抗真菌蛋白由相当大比例的真菌产生，特别是由与昆虫、水果和发酵（例如酿造和烘焙）相关的酵母产生。公众对酵母的普遍熟悉将使学童和当地农贸市场的顾客能够参与分离产生新型抗真菌蛋白的酵母。这些推广活动将通过本科研究生的参与得到加强，以鉴定新型抗真菌蛋白及其应用，进一步研究真菌膜和细胞壁的功能和组织。来自爱达荷州大学代表性不足群体的学生也将被纳入研究活动。这种方法的核心目标是提高大学生的保留率，中学后和学士后学生的高等教育入学率，并改善公共STEM教育。本研究的目标是发现真菌对抗真菌“杀手”毒素的耐药性的重要细胞机制。这将揭示对细胞表面功能和弹性至关重要的新途径。研究假设是Ace 2和细胞形态发生（RAM）信号网络的调节器在细胞壁和膜组织中起着未描述的作用，并且杀伤毒素受体Kre 1 p的定位和多样性对于杀伤毒素中毒至关重要。这些假设是根据初步数据制定的，这些数据已经确定了RAM网络中的突变，导致杀伤毒素抗性和膜和细胞壁完整性的丧失。为了检验这些假设，将确定导致杀伤毒素抗性的RAM网络中的突变如何改变与胞质分裂和细胞极性相关的网络组织、定位和功能。具体而言，将测试这些突变对杀伤毒素膜受体（Kre 1 p）定位的影响。这种方法将通过深入研究Kre 1 p多样性对杀手毒素抗性的影响以及防止自我中毒的杀手酵母的免疫机制来补充。这项研究的结果将证实RAM网络在细胞表面组织中的新作用。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

This award is funded in whole or in part under the American Rescue Plan Act of 2021 (Public Law 117-2).Fungal cells separate themselves from their environment by constructing a robust cell wall made of carbohydrates and proteins with an underlying membrane. This enables the compartmentalization of metabolic processes and protects fungal cells from external stressors, including commercial fungicides that are used extensively to control unwanted fungal growth. Antifungal proteins produced by different species of fungi can attack the cell wall and membrane causing injury and the death of susceptible fungal cells. These antifungals have been cited as potentially being useful to control the growth of undesirable fungi. This research will explore the fundamental cellular mechanisms that are employed by fungi to resist intoxication by antifungal proteins. Specifically, the project will test how mutations in genes used for cell wall and membrane construction and stability result in resistance to antifungal proteins. Antifungal proteins are produced by a significant proportion of fungi, especially by yeasts that are associated with insects, fruits, and fermentation (e.g. brewing and baking). The general familiarity of the public with yeasts will enable schoolchildren and patrons of the local farmer’s market to participate in the isolation of yeasts that produce novel antifungal proteins. These outreach activities will be enhanced by the participation of undergraduate research students to enable the identification of novel antifungal proteins and their application to further investigate the function and organization of the fungal membrane and cell wall. Students from underrepresented groups at the University of Idaho will also be included in research activities. The central goal of this approach is to increase retention of university students, enrollment in higher education of postsecondary and postbaccalaureate students, and to improve public STEM education.The goal of this research is to discover the cellular mechanisms that are important for fungal resistance to antifungal “killer” toxins. This will uncover novel pathways that are important for cell surface function and resilience. The research hypotheses are that the Regulator of Ace2 and Cell Morphogenesis (RAM) signaling network plays an undescribed role in cell wall and membrane organization, and that the localization and diversity of the killer toxin receptor Kre1p is essential for intoxication by killer toxins. These hypotheses were formulated based on preliminary data that has identified mutations in the RAM network that result in killer toxin resistance and the loss of membrane and cell wall integrity. To test these hypotheses, it will be determined how mutations in the RAM network that cause killer toxin resistance alter the networks organization, localization, and function relating to cytokinesis and cell polarity. Specifically, the effect of these mutations on the localization of the killer toxin membrane receptor (Kre1p) will be tested. This approach will be complemented by an in-depth study of the effect of Kre1p diversity on killer toxin resistance and the immunity mechanisms of killer yeasts that prevent self-intoxication. The outcome of this research will be to confirm the novel role of the RAM network in cell surface organization. This will be complemented by the investigation of the RAM network and Kre1p and their role in killer toxin resistance.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.