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通过 ddPCR 对巨噬细胞吞噬白念珠菌早期阶段的碳代谢进行快照分析。

Carbon metabolism snapshot by ddPCR during the early step of Candida albicans phagocytosis by macrophages.

机构信息

Génétique Moléculaire des Levures, UMR-CNRS 5240 Microbiologie Adaptation et Pathogénie, Université de Lyon-Université Lyon1, Lyon, France.

DTAMB, FR 3728 Bio-Environnement et Santé, Université de Lyon-Université Lyon1, Lyon, France.

出版信息

Pathog Dis. 2020 Feb 1;78(1). doi: 10.1093/femspd/ftaa014.

Abstract

During Candida macrophage interactions, phagocytosed yeast cells feed in order to grow, develop hyphae and escape. Through numerous proteomic and transcriptomic studies, two metabolic phases have been described. A shift to a starvation mode is generally identified as early as one-hour post phagocytosis, followed by a glycolytic growth mode after C. albicans escaped from the macrophage. Healthy macrophages contain low amounts of glucose. To determine if this carbon source was sensed and metabolized by the pathogen, we explored the transcription level of a delimited set of key genes expressed in C. albicans cells during phagocytosis by macrophages, at an early stage of the interaction. This analysis was performed using a technical digital droplet PCR approach to quantify reliably the expression of carbon metabolic genes after 30 min of phagocytosis. Our data confirm the technique of digital droplet PCR for the detection of C. albicans transcripts using cells recovered after a short period of phagocytosis. At this stage, carbon metabolism is clearly oriented towards the use of alternative sources. However, the activation of high-affinity glucose transport system suggests that the low amount of glucose initially present in the macrophages is detected by the pathogen.

摘要

在念珠菌与巨噬细胞相互作用过程中,被吞噬的酵母细胞为了生长、发育菌丝并逃脱而进食。通过大量的蛋白质组学和转录组学研究,已经描述了两个代谢阶段。早在被吞噬后一小时,就会出现向饥饿模式的转变,然后在白色念珠菌从巨噬细胞中逃脱后,会出现糖酵解生长模式。健康的巨噬细胞中含有少量的葡萄糖。为了确定病原体是否感知并代谢了这种碳源,我们研究了在白色念珠菌被巨噬细胞吞噬的早期,一组在巨噬细胞中被吞噬的白色念珠菌细胞中表达的关键基因的转录水平。通过使用数字液滴 PCR 技术,在吞噬后 30 分钟内可靠地定量表达碳代谢基因,进行了这项分析。我们的数据证实了数字液滴 PCR 技术可用于检测短时间吞噬后回收的白色念珠菌转录物。在这个阶段,碳代谢明显偏向于使用替代来源。然而,高亲和力葡萄糖转运系统的激活表明,最初存在于巨噬细胞中的少量葡萄糖被病原体检测到。

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