New Mechanisms of Flucytosine Resistance in C. glabrata Unveiled by a Chemogenomics Analysis in S. cerevisiae.
New Mechanisms of Flucytosine Resistance in C. glabrata Unveiled by a Chemogenomics Analysis in S. cerevisiae.
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DOI：
10.1371/journal.pone.0135110
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Teixeira MC
中科院分区：
文献类型：
--
作者：
Costa C;Ponte A;Pais P;Santos R;Cavalheiro M;Yaguchi T;Chibana H;Teixeira MC
5-氟胞嘧啶目前在联合治疗中用作抗真菌药物，但真菌病原体很快就能对该药物产生耐药性，从而损害其治疗作用。了解潜在的抵抗机制对于解决这个问题至关重要。在这项工作中，筛选了酿酒酵母缺失突变体集合，以提高对氟胞嘧啶的抗性。通过化学基因组学分析，发现 183 个基因赋予这种抗真菌药物耐药性。与其在 DNA、RNA 和蛋白质合成中的已知作用一致，5-氟胞嘧啶抗性决定因素列表中过多出现的最重要的基因本体术语与 DNA 修复、RNA 和蛋白质代谢有关。其他功能类别包括碳水化合物和氮（尤其是精氨酸）代谢、脂质代谢和细胞壁重塑。基于以酿酒酵母作为模型系统获得的结果，对致病性酵母光滑念珠菌进行了进一步的研究。研究发现补充精氨酸可以减轻 5-氟胞嘧啶对光滑念珠菌的抑制作用。研究发现，在接触 5-氟胞嘧啶后的前 30 分钟内，裂解酶敏感性增加，表明这种抗真菌药物可充当细胞壁损伤剂。在 5-氟胞嘧啶存在下恢复指数生长后，细胞壁表现出更高的对裂解酶的抵抗力，表明细胞壁重塑是响应 5-氟胞嘧啶而发生的。此外，来自光滑念珠菌的水甘油孔蛋白编码基因 CgFPS1 和 CgFPS2 被鉴定为 5-氟胞嘧啶抗性的决定因素。研究发现 CgFPS1 和 CgFPS2 通过减少光滑念珠菌细胞中 5-氟胞嘧啶的积累来介导 5-氟胞嘧啶抗性。
5-Flucytosine is currently used as an antifungal drug in combination therapy, but fungal pathogens are rapidly able to develop resistance against this drug, compromising its therapeutic action. The understanding of the underlying resistance mechanisms is crucial to deal with this problem. In this work, the S. cerevisiae deletion mutant collection was screened for increased resistance to flucytosine. Through this chemogenomics analysis, 183 genes were found to confer resistance to this antifungal agent. Consistent with its known effect in DNA, RNA and protein synthesis, the most significant Gene Ontology terms over-represented in the list of 5-flucytosine resistance determinants are related to DNA repair, RNA and protein metabolism. Additional functional classes include carbohydrate and nitrogen—particularly arginine—metabolism, lipid metabolism and cell wall remodeling. Based on the results obtained for S. cerevisiae as a model system, further studies were conducted in the pathogenic yeast Candida glabrata. Arginine supplementation was found to relieve the inhibitory effect exerted by 5-flucytosine in C. glabrata. Lyticase susceptibility was found to increase within the first 30min of 5-flucytosine exposure, suggesting this antifungal drug to act as a cell wall damaging agent. Upon exponential growth resumption in the presence of 5-flucytosine, the cell wall exhibited higher resistance to lyticase, suggesting that cell wall remodeling occurs in response to 5-flucytosine. Additionally, the aquaglyceroporin encoding genes CgFPS1 and CgFPS2, from C. glabrata, were identified as determinants of 5-flucytosine resistance. CgFPS1 and CgFPS2 were found to mediate 5-flucytosine resistance, by decreasing 5-flucytosine accumulation in C. glabrata cells.