A novel metal-starvation candidacidal mechanism for Histatin 5

项目摘要： 口咽和食管念珠菌病主要由条件致病菌白色念珠菌引起 并继续成为免疫功能低下人群的健康问题。反复感染C.白色念珠菌是 通常对多种主要的抗真菌药物类型具有耐药性。在这些情况下，很少有替代疗法，因为 临床上可用的抗真菌剂靶向有限数量的真菌途径。唾液组胺素5（Hst 5）是 由人的大唾液腺分泌，具有高的体外杀真菌活性。白色念珠菌和其他 念珠菌属。Hst 5的作用机制与目前使用的抗真菌药物不同， 使其成为一种有吸引力的替代治疗;然而，最终的杀死机制尚不清楚。HST 5 处理对C.白色念珠菌细胞，包括活性氧（ROS）积累， 高渗透压甘油（HOG 1）途径的激活和G1期细胞周期阻滞，然而其潜在的 其作用机制仍然是未知的。重要的是，Hst 5能够结合锌（Zn 2+）和铜（Cu 2+）。 具有高亲和力的离子和这些金属的结合已经显示出影响对未知生物的杀伤活性。 原因Hst 5进入真菌细胞是其高杀伤活性和游离金属结合位点所必需的 似乎是摄取肽所必需的;因此，该项目的中心假设是Hst 5进入真菌细胞耗尽细胞内的Zn 2+和Cu 2+，这是Hst 5 诱导C.白色念珠菌作为工具 为了探测Hst 5的金属依赖性效应，Hst 5是一种突变型Hst 5肽， 将使用绑定。在目标1中，将验证Hst 5耗尽细胞内Zn 2+和Cu 2+的能力，并且 ROS解毒超氧化物歧化酶蛋白在Hst 5介导的金属代谢过程中的表达和活性 将表征耗尽。目的2：探讨Zn ~（2+）对HOG ~（1+）信号通路和细胞周期的调控作用 将测试Hst 5抑制，并将证实Hst 5治疗通过以下途径使这些过程失调： 细胞内Zn 2+耗竭。这一项目的完成将是HST 5研究向前迈出的宝贵一步 作用机制，并提供了新的C.白色念珠菌应激反应。

Project Summary: Oropharyngeal and esophageal candidiasis are mainly caused by the opportunistic pathogen Candida albicans and continue to be a health issue in immunocompromised populations. Recurrent infections of C. albicans are often resistant to multiple major antifungal drug types. In these cases there are few alternative therapies since clinically available antifungals target a limited number of fungal pathways. Salivary Histatin 5 (Hst 5) is secreted by human major salivary glands and has high in vitro fungicidal activity against C. albicans and other Candida species. Hst 5 has a mechanism of action that is distinct from antifungal drugs currently in use, making it an attractive alternative treatment; however the ultimate mechanism of killing is unclear. Hst 5 treatment induces several effects on C. albicans cells, including reactive oxygen species (ROS) accumulation, activation of the high osmolarity glycerol (HOG1) pathway and cell cycle arrest in G1, however its underlying mechanism of action is still mostly unknown. Importantly, Hst 5 is able to bind zinc (Zn2+) and copper (Cu2+) ions with high affinity and binding of these metals has been shown to impact killing activity for unknown reasons. Entry of Hst 5 into fungal cells is required for its high killing activity, and free metal binding sites appear to be necessary for uptake of the peptide; therefore the central hypothesis of this project is that Hst 5 enters fungal cells to deplete intracellular Zn2+ and Cu2+, which is the mechanism by which Hst 5 induces oxidative stress, changes in HOG1 signaling, and G1 cell cycle arrest in C. albicans. As a tool to probe the metal-dependent effects of Hst 5, a mutant Hst 5 peptide that is designed to be deficient in metal binding will be used. In Aim 1, the ability of Hst 5 to deplete intracellular Zn2+ and Cu2+ will be verified, and the expression and activity of ROS-detoxifying superoxide dismutase proteins during Hst 5 mediated metal depletion will be characterized. In Aim 2 the ability of Zn2+ to regulate HOG1 pathway signaling and cell cycle arrest will be tested, and it will be confirmed that Hst 5 treatment dysregulates these processes through intracellular Zn2+ depletion. Completion of this project will be a valuable step forward in research into the Hst 5 mechanism of action, and provide new characterization of C. albicans stress responses.