RNAi machinery regulates nutrient metabolism and fluconazole resistance in the pathogenic fungus Cryptococcus deneoformans
RNAi machinery regulates nutrient metabolism and fluconazole resistance in the pathogenic fungus Cryptococcus deneoformans
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RNAi机制调节病原真菌隐球菌的营养代谢和氟康唑耐药性
DOI：
10.1093/mmy/myac095
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发表时间：
2023-02-22
期刊：
影响因子：
2.9
通讯作者：
Zhu，Xudong
中科院分区：
文献类型：
--
作者：
Ma，Xiaoyu;Li，Chenxi;Zhu，Xudong
RNAi机制在植物和动物细胞中因其在基因组功能调节中的关键作用而被广泛研究。然而，RNAi在控制真菌生长发育中的潜在作用研究很少，特别是在担子菌酵母隐球菌中。为了研究RNAi在病原真菌中的生物学功能，采用高通量测序技术对野生型和RNAi突变体的mRNA进行了比较分析。新形动物。结果揭示了RNAi突变体中与代谢过程相关的基因表达的明显差异。此外，Ago 2 Δ突变体在营养限制条件下的生长显著降低，表明Ago 2在营养代谢中起重要作用。进一步的研究发现RNAi突变体中存在差异表达的转运蛋白，其中参与氟康唑外排的转运蛋白表达显著上调。更重要的是，由于转运蛋白的上调，RNAi突变株对氟康唑产生了耐药性。通过使用“自杀”CRISPR-Cas9系统破坏AFR 1基因，我们证实了RNAi突变体中上调的ABC转运蛋白Afr 1有助于氟康唑抗性。总之，我们的数据表明，inC。隐球菌RNAi途径参与营养代谢，并在抑制氟康唑耐药中发挥作用，为深入了解隐球菌的RNAi机制提供了重要线索，为隐球菌病的临床治疗提供了重要启示。
The RNAi machinery has been extensively studied in plant and animal cells for their crucial roles in the regulation of genome function. However, the potential roles of RNAi in controlling fungal growth and development have been poorly studied, especially in the basidiomycetous yeastCryptococcus deneoformans. To characterize the biological functions of RNAi in the pathogenic fungus, a comparative analysis of mRNA profiles using high-throughput sequencing technology was performed for the wild type and the RNAi mutants ofC. deneoformans. The results revealed a clear difference in the expression of genes associated with metabolic processes in the RNAi mutants. Besides, the growth under nutrient-limited conditions was significantly reduced in theago2Δmutant, suggesting the essential roles of Ago2 in nutrient metabolism. Further investigations revealed the differentially expressed transporters in the RNAi mutants, in which transporters involved in fluconazole efflux were significantly up-regulated. More importantly, on account of the upregulated transporters, RNAi mutant strains developed resistance to fluconazole. By disruptingAFR1gene using the ‘suicide’ CRISPR-Cas9 system, we verified that the upregulated ABC transporter Afr1 in the RNAi mutants contributed to the fluconazole resistance. In summary, our data demonstrate that inC. deneoformansthe RNAi pathway participates in nutrient metabolism and plays a role in the repression of fluconazole resistance, which provides a deep insight into RNAi mechanisms inCryptococcusand brings great hints for the clinical treatment of cryptococcosis.