Multifactorial Role of Mitochondria in Echinocandin Tolerance Revealed by Transcriptome Analysis of Drug-Tolerant Cells.
Multifactorial Role of Mitochondria in Echinocandin Tolerance Revealed by Transcriptome Analysis of Drug-Tolerant Cells.
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DOI：
10.1128/mbio.01959-21
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发表时间：
2021-08-31
期刊：
影响因子：
6.4
通讯作者：
Perlin DS
中科院分区：
文献类型：
--
作者：
Garcia-Rubio R;Jimenez-Ortigosa C;DeGregorio L;Quinteros C;Shor E;Perlin DS
真菌感染在世界范围内引起严重的死亡率和发病率，并且有限的现有抗真菌库因对棘白菌素耐药的菌株的出现而进一步削弱，棘白菌素是抗真菌治疗的一线。光假丝酵母是一种机会性真菌病原体，其棘白菌素药物靶点β-1,3-葡聚糖合成酶（GS）快速发生突变，与耐药和临床失败有关。尽管棘球白菌素被认为是假丝酵母的杀菌剂，但有一部分秃毛假丝酵母细胞能在接触棘球白菌素后存活下来，形成一个耐药细胞库，由此产生耐药突变。尽管它们很重要，但人们对罕见的耐药细胞的生理学知之甚少。我们使用荧光激活的细胞分选富集棘白菌素耐受细胞，然后使用改良的单细胞RNA测序来检查它们的转录景观。该分析确定了不同于典型酵母环境胁迫反应的转录特征，其特征是染色体结构和DNA拓扑通路上调，氧化应激反应下调，其中氧化应激反应在活性氧水平增加的情况下仍然存在。进一步的分析表明线粒体与棘球白素耐受性有关，其中线粒体复合物I和IV的抑制剂减少了棘球白素介导的细胞杀伤，但缺乏各种线粒体成分的突变体都表现出棘球白素耐受性低的表型。最后，从线粒体突变体中纯化的GS酶复合物表现出正常的体外抑制动力学，表明线粒体缺陷影响药物-靶标相互作用下游的细胞存活。综上所述，这些结果为裸棘球藻对棘白菌素的反应提供了新的见解，并揭示了线粒体在棘白菌素耐受中的多因素作用。
Fungal infections cause significant mortality and morbidity worldwide, and the limited existing antifungal reservoir is further weakened by the emergence of strains resistant to echinocandins, a first line of antifungal therapy. Candida glabrata is an opportunistic fungal pathogen that rapidly develops mutations in the echinocandin drug target β-1,3-glucan synthase (GS), which are associated with drug resistance and clinical failure. Although echinocandins are considered fungicidal in Candida sp., a subset of C. glabrata cells survive echinocandin exposure, forming a drug-tolerant cell reservoir, from which resistant mutations are thought to emerge. Despite their importance, the physiology of rare drug-tolerant cells is poorly understood. We used fluorescence-activated cell sorting to enrich for echinocandin-tolerant cells, followed by modified single-cell RNA sequencing to examine their transcriptional landscape. This analysis identified a transcriptional signature distinct from the stereotypical yeast environmental stress response and characterized by upregulation of pathways involved in chromosome structure and DNA topology and downregulation of oxidative stress responses, of which the latter was observed despite increased levels of reactive oxygen species. Further analyses implicated mitochondria in echinocandin tolerance, wherein inhibitors of mitochondrial complexes I and IV reduced echinocandin-mediated cell killing, but mutants lacking various mitochondrial components all showed an echinocandin hypotolerant phenotype. Finally, GS enzyme complexes purified from mitochondrial mutants exhibited normal in vitro inhibition kinetics, indicating that mitochondrial defects influence cell survival downstream of the drug-target interaction. Together, these results provide new insights into the C. glabrata response to echinocandins and reveal a multifactorial role of mitochondria in echinocandin tolerance.