Caza Mélissa, Hu Guanggan, Price Michael, Perfect John R, Kronstad James W
Department of Microbiology and Immunology, Michael Smith Laboratories, University of British Columbia, Vancouver, Canada.
Department of Medicine and Molecular Genetics and Microbiology, Duke University Medical Center, Duke University, Durham, North Carolina, USA.
mSphere. 2016 Jan 13;1(1). doi: 10.1128/mSphere.00080-15. eCollection 2016 Jan-Feb.
The opportunistic pathogen Cryptococcus neoformans causes fungal meningoencephalitis in immunocompromised individuals. In previous studies, we found that the Hap complex in this pathogen represses genes encoding mitochondrial respiratory functions and tricarboxylic acid (TCA) cycle components under low-iron conditions. The orthologous Hap2/3/4/5 complex in Saccharomyces cerevisiae exerts a regulatory influence on mitochondrial functions, and Hap4 is subject to glucose repression via the carbon catabolite repressor Mig1. In this study, we explored the regulatory link between a candidate ortholog of the Mig1 protein and the HapX component of the Hap complex in C. neoformans. This analysis revealed repression of MIG1 by HapX and activation of HAPX by Mig1 under low-iron conditions and Mig1 regulation of mitochondrial functions, including respiration, tolerance for reactive oxygen species, and expression of genes for iron consumption and iron acquisition functions. Consistently with these regulatory functions, a mig1Δ mutant had impaired growth on inhibitors of mitochondrial respiration and inducers of ROS. Furthermore, deletion of MIG1 provoked a dysregulation in nutrient sensing via the TOR pathway and impacted the pathway for cell wall remodeling. Importantly, loss of Mig1 increased susceptibility to fluconazole, thus further establishing a link between azole antifungal drugs and mitochondrial function. Mig1 and HapX were also required together for survival in macrophages, but Mig1 alone had a minimal impact on virulence in mice. Overall, these studies provide novel insights into a HapX/Mig1 regulatory network and reinforce an association between mitochondrial dysfunction and drug susceptibility that may provide new targets for the development of antifungal drugs. IMPORTANCE Fungal pathogens of humans are difficult to treat, and there is a pressing need to identify new targets for antifungal drugs and to obtain a detailed understanding of fungal proliferation in vertebrate hosts. In this study, we examined the roles of the regulatory proteins Mig1 and HapX in mitochondrial function and antifungal drug susceptibility in the fungus Cryptococcus neoformans. This pathogen is a particular threat to the large population of individuals infected with human immunodeficiency virus (HIV). Our analysis revealed regulatory interactions between Mig1 and HapX, and a role for Mig1 in mitochondrial functions, including respiration, tolerance for reactive oxygen species, and expression of genes for iron consumption and iron acquisition functions. Importantly, loss of Mig1 increased susceptibility to the antifungal drug fluconazole, which is commonly used to treat cryptococcal disease. These studies highlight an association between mitochondrial dysfunction and drug susceptibility that may provide new targets for the development of antifungal drugs.
机会性致病病原体新型隐球菌可在免疫功能低下的个体中引发真菌性脑膜脑炎。在之前的研究中，我们发现该病原体中的Hap复合物在低铁条件下会抑制编码线粒体呼吸功能和三羧酸（TCA）循环组分的基因。酿酒酵母中的直系同源Hap2/3/4/5复合物对线粒体功能发挥调控作用，且Hap4会受到碳分解代谢物阻遏物Mig1介导的葡萄糖阻遏作用。在本研究中，我们探究了Mig1蛋白的候选直系同源物与新型隐球菌中Hap复合物的HapX组分之间的调控联系。该分析揭示了在低铁条件下HapX对MIG1的抑制作用以及Mig1对HAPX的激活作用，还有Mig1对线粒体功能的调控，包括呼吸作用、对活性氧的耐受性以及铁消耗和铁获取功能相关基因的表达。与这些调控功能一致，mig1Δ突变体在呼吸链抑制剂和活性氧诱导剂存在的情况下生长受损。此外，MIG1的缺失引发了通过TOR途径进行的营养感知失调，并影响了细胞壁重塑途径。重要的是，Mig1的缺失增加了对氟康唑的敏感性，从而进一步确立了唑类抗真菌药物与线粒体功能之间的联系。Mig1和HapX对于在巨噬细胞内存活也是必需的，但单独的Mig1对小鼠毒力的影响极小。总体而言，这些研究为HapX/Mig1调控网络提供了新见解，并强化了线粒体功能障碍与药物敏感性之间的关联，这可能为抗真菌药物的开发提供新靶点。重要性人类真菌病原体难以治疗，迫切需要确定抗真菌药物的新靶点，并详细了解真菌在脊椎动物宿主体内的增殖情况。在本研究中，我们研究了调控蛋白Mig1和HapX在新型隐球菌线粒体功能和抗真菌药物敏感性中的作用。该病原体对大量感染人类免疫缺陷病毒（HIV）的个体构成了特殊威胁。我们的分析揭示了Mig1和HapX之间的调控相互作用，以及Mig1在线粒体功能中的作用，包括呼吸作用、对活性氧的耐受性以及铁消耗和铁获取功能相关基因的表达。重要的是，Mig1的缺失增加了对常用于治疗隐球菌病的抗真菌药物氟康唑的敏感性。这些研究突出了线粒体功能障碍与药物敏感性之间的关联，这可能为抗真菌药物的开发提供新靶点。
