Antifungal azoles are the most widely used antifungal drugs in clinical and agricultural practice. Fungi can mount adaptive responses to azole stress by modifying the transcript levels of many genes, and the responsive mechanisms to azoles are the basis for fungi to develop azole resistance. In this study, we identified a new Zn(II)₂Cys₆ transcription factor, ADS-1, with a positive regulatory function in transcriptional responses to azole stress in the model filamentous fungal species Neurospora crassa. Under ketoconazole (KTC) stress, the ads-1 transcript level was significantly increased in N. crassa. Deletion of ads-1 increased susceptibility to different azoles, while its overexpression increased resistance to these azoles. The cdr4 gene, which encodes the key azole efflux pump, was positively regulated by ADS-1. Deletion of ads-1 reduced the transcriptional response by cdr4 to KTC stress and increased cellular KTC accumulation under KTC stress, while ads-1 overexpression had the opposite effect. ADS-1 also positively regulated the transcriptional response by erg11, which encodes the azole target lanosterol 14α-demethylase for ergosterol biosynthesis, to KTC stress. After KTC treatment, the ads-1 deletion mutant had less ergosterol but accumulated more lanosterol than the wild type, while ads-1 overexpression had the opposite effect. Homologs of ADS-1 are widely present in filamentous fungal species of Ascomycota but not in yeasts. Deletion of the gene encoding an ADS-1 homolog in Aspergillus flavus also increased susceptibility to KTC and itraconazole (ITZ). Besides, deletion of A. flavus ads-1 (Afads-1) significantly reduced the transcriptional responses by genes encoding homologs of CDR4 and ERG11 in A. flavus to KTC stress, and the deletion mutant accumulated more KTC but less ergosterol. Taken together, these findings demonstrate that the function and regulatory mechanism of ADS-1 homologs among different fungal species in azole responses and the basal resistance of azoles are highly conserved.

中文翻译：

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真菌Zn（II）2Cys6转录因子ADS-1调节抗真菌唑反应下的药物外流和麦角固醇代谢。

抗真菌唑类是临床和农业实践中使用最广泛的抗真菌药物。真菌可以通过修饰许多基因的转录水平来提高对吡咯胁迫的适应性反应，并且对吡咯的响应机制是真菌发展对吡咯抗性的基础。在这项研究中，我们确定了一种新的Zn（II）₂ Cys ₆转录因子ADS-1，在模型丝状真菌物种Neurospora crassa中对唑胁迫的转录响应中具有正调节功能。在酮康唑（KTC）胁迫下，景天猪笼草的ads-1转录水平显着增加。删除广告1增加了对不同唑的敏感性，而其过表达增加了对这些唑的抗性。编码关键的唑外排泵的cdr4基因受到ADS-1的正调控。ads-1的缺失减少了cdr4对KTC胁迫的转录反应，并增加了在KTC胁迫下细胞KTC的积累，而ads-1的过表达具有相反的作用。ADS-1还通过erg11积极调节转录反应，该基因编码对麦角固醇进行生物合成的吡咯目标羊毛甾醇14α-脱甲基酶对KTC胁迫。经过KTC处理后，ads-1缺失突变体的麦角固醇较少，但积累的羊毛脂比野生型多，而ads-1过度表达产生相反的效果。ADS-1的同源物广泛存在于子囊菌的丝状真菌物种中，但在酵母中不存在。黄曲霉中编码ADS-1同源物的基因的缺失也增加了对KTC和伊曲康唑（ITZ）的敏感性。此外，黄曲霉 ads-1（Af ads-1）的缺失显着降低了编码黄曲霉CDR4和ERG11同源基因的转录反应。对KTC胁迫，缺失突变体积累了更多的KTC，但麦角固醇较少。综上所述，这些发现表明不同真菌物种之间的ADS-1同源物在唑响应和唑的基础抗性中的功能和调节机制是高度保守的。