AtrR Is an Essential Determinant of Azole Resistance in Aspergillus fumigatus
AtrR Is an Essential Determinant of Azole Resistance in Aspergillus fumigatus
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DOI：
10.1128/mbio.02563-18
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发表时间：
2019-03-01
期刊：
影响因子：
6.4
通讯作者：
Moye-Rowley, W. Scott
中科院分区：
文献类型：
--
作者：
Paul, Sanjoy;Stamnes, Mark;Moye-Rowley, W. Scott
与唑类耐药烟曲霉菌相关的曲霉病在某些患者群体中的死亡率可接近90%。唑类药物耐药的最佳途径涉及cyp 51 A基因的变化，该基因编码唑类药物的靶点羊毛甾醇α-14去甲基酶。目前描述的最常见的唑类耐药等位基因是一个连锁变化，对应于cyp 51 A编码序列的变化和启动子中34 bp区域的重复，导致串联重复序列（TR）。我们以前的研究确定了一个积极作用的转录因子称为AtrR，结合cyp 51 A的启动子，以及一个重要的膜转运蛋白基因称为abcG 1。在这项工作中，我们描述了两种不同的突变体等位基因的atrR，无论是生产过剩或表位标记的形式，导致组成激活这个因素。使用atrR的表位标记的等位基因进行染色质免疫沉淀结合高通量测序（ChIP-seq），确定AtrR的基因组结合位点。近900个基因被发现在其启动子区域中具有AtrR反应元件（ATRE）。通过RNA测序（RNA-seq）进行的转录组评估表明，两种等位基因均导致靶基因子集的转录升高。电泳迁移率变动分析和DNase I保护定位ATRE在abcG 1和cyp 51 A启动子。cyp 51 A中的ATRE位于34-bp重复元件内。当AtrR缺失或过量产生时，小鼠模型中的毒力受到损害，表明该因子的适当剂量是致病的关键。重要信息烟曲霉是人类主要的丝状真菌病原体。与A.烟曲霉通常用唑类药物治疗，但对这些抗真菌剂的耐药性正在增加。与唑类耐药真菌相关的曲霉病的死亡率极高。以前的工作已经确定唑类药物靶向编码基因cyp 51 A的转录控制是A.烟熏。在这里，我们证明了转录因子AtrR结合到cyp 51 A启动子中的一个区域，该区域与该基因的等位基因相关，从而赋予临床上重要的唑类耐药。利用高通量基因组技术，我们还发现了一个由AtrR控制的靶基因的大套件。这些数据表明，AtrR协调调节涉及耐药性，代谢和毒力的许多不同的过程。我们对AtrR功能的新认识为A.烟熏。
Aspergillosis associated with azole-resistant Aspergillus fumigatus has a mortality rate that can approach 90% in certain patient populations. The best-understood avenue for azole resistance involves changes in the cyp51A gene that encodes the target of azole drugs, lanosterol alpha-14 demethylase. The most common azole resistance allele currently described is a linked change corresponding to a change in the coding sequence of cyp51A and a duplication of a 34-bp region in the promoter leading to a tandem repeat (TR). Our previous studies identified a positively acting transcription factor called AtrR that binds to the promoter of cyp51A as well as that of an important membrane transporter protein gene called abcG1. In this work, we characterize two different mutant alleles of atrR, either an overproducing or an epitope-tagged form, causing constitutive activation of this factor. Using an epitope-tagged allele of atrR for chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-seq), the genomic binding sites for AtrR were determined. Close to 900 genes were found to have an AtrR response element (ATRE) in their promoter regions. Transcriptome evaluation by RNA sequencing (RNA-seq) indicated that both alleles led to elevated transcription of a subset of target genes. An electrophoretic mobility shift assay and DNase I protection mapping localized the ATREs in both the abcG1 and cyp51A promoters. The ATRE in cyp51A was located within the 34-bp repeat element. Virulence in a murine model was compromised when AtrR was either deleted or overproduced, indicating that the proper dosage of this factor is key for pathogenesis.IMPORTANCE Aspergillus fumigatus is the major filamentous fungal pathogen in humans. Infections associated with A. fumigatus are often treated with azole drugs, but resistance to these antifungal agents is increasing. Mortality from aspergillosis associated with azole-resistant fungi is extremely high. Previous work has identified transcriptional control of the azole drug target-encoding gene cyp51A as an important contributor to resistance in A. fumigatus. Here, we demonstrate that the transcription factor AtrR binds to a region in the cyp51A promoter that is associated with alleles of this gene conferring clinically important azole resistance. Using high-throughput genomic technologies, we also uncover a large suite of target genes controlled by AtrR. These data indicate that AtrR coordinately regulates many different processes involved in drug resistance, metabolism, and virulence. Our new understanding of AtrR function provides important new insight into the pathogenesis of A. fumigatus.