ABSTRACT

Aspergillus fumigatus is the leading cause of life-threatening invasive mold infections in immunocompromised individuals. This ubiquitous saprophyte possesses several natural attributes allowing it to evade the immune system, including the ability to withstand high toxic Cu concentrations within the phagosomes of macrophages and neutrophils. We previously established that at high levels, Cu binds and activates the A. fumigatus transcription factor AceA, which upregulates the expression of the Cu exporter CrpA to expel excess Cu. Deletion of aceA or crpA result in extreme Cu sensitivity and attenuated virulence.

To identify other elements participating in resistance to Cu, we performed a genome-wide analysis of the transcriptome by RNAseq to analyze the AceA-dependent response of A. fumigatus to excess Cu. We deleted key genes whose transcription was strongly upregulated by high Cu, including those encoding homologs of the three Cu chaperones cox17, atx1 and ccs1. Detailed analysis of these genes indicates that in A. fumigatus, cox17 is an essential gene with a possible role in respiration, the atxA gene product participates in reductive iron uptake and ccsA encodes the Cu chaperone activating A. fumigatus Sod1. Interestingly, although the ccsA-null strain was extremely sensitive to high Cu and oxidative stress, it was not attenuated in virulence in a mouse model of invasive pulmonary aspergillosis.

Our work provides (i) a detailed view of the genome-wide transcriptional response of A. fumigatus to excess Cu, (ii) identification of the AceA-dependent transcriptome and (iii) analysis of the roles of the three Cu chaperones cox17, atxA and ccsA.

摘要

烟曲霉是免疫功能低下个体中威胁生命的侵袭性霉菌感染的主要原因。这种无处不在的腐生菌具有使其能够逃避免疫系统的多种自然属性，包括能够承受巨噬细胞和中性粒细胞吞噬体内高毒性铜浓度的能力。我们之前确定，在高水平时，Cu 结合并激活烟曲霉转录因子 AceA，它上调 Cu 输出蛋白 CrpA 的表达以排出过量的 Cu。aceA或crpA的缺失导致极度的铜敏感性和减弱的毒力。

为了确定参与对铜的抗性的其他元素，我们通过 RNAseq 对转录组进行了全基因组分析，以分析烟曲霉对过量铜的 AceA 依赖性反应。我们删除了转录被高铜强烈上调的关键基因，包括那些编码三个铜伴侣cox17、atx1和ccs1的同源物的基因。对这些基因的详细分析表明，在烟曲霉中，cox17是可能在呼吸作用中发挥作用的必需基因，atxA基因产物参与还原性铁的吸收，ccsA编码激活烟曲霉Sod1的Cu 伴侣蛋白。有趣的是，虽然ccsA-null 菌株对高铜和氧化应激极为敏感，在侵袭性肺曲霉病小鼠模型中其毒力未减弱。

我们的工作提供（i）烟曲霉对过量铜的全基因组转录反应的详细视图，（ii）AceA依赖性转录组的鉴定和（iii）三种铜伴侣cox17，atxA的作用分析和ccsA。