Neutrophil extracellular traps contain calprotectin, a cytosolic protein complex involved in host defense against Candida albicans.
Neutrophil extracellular traps contain calprotectin, a cytosolic protein complex involved in host defense against Candida albicans.
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DOI：
10.1371/journal.ppat.1000639
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发表时间：
2009-10
期刊：
影响因子：
6.7
通讯作者：
Zychlinsky A
中科院分区：
文献类型：
--
作者：
Urban CF;Ermert D;Schmid M;Abu-Abed U;Goosmann C;Nacken W;Brinkmann V;Jungblut PR;Zychlinsky A
中性粒细胞是感染部位的第一道防线。它们在吞噬作用后在细胞内或通过抗菌蛋白的脱粒和释放中性粒细胞胞外陷阱（NET）在细胞外遇到并杀死微生物。NET被证明可以诱捕和杀死微生物。然而，其完整的蛋白质组成和抗菌机制还不清楚。使用蛋白质组学方法，我们确定了24 NET相关蛋白。对这些蛋白质的定量分析和高分辨率电子显微镜显示，NET由修饰的核小体和严格选择的其他蛋白质组成。与以前的结果相反，我们发现几个NET蛋白质是在未刺激的中性粒细胞的细胞质。我们证明了这些蛋白质中，抗菌异源二聚体钙卫蛋白作为主要的抗真菌成分在NET中释放。钙卫蛋白在NET中的缺乏导致体外抗真菌活性的完全丧失。三种不同的白色念珠菌在体内感染模型的分析表明，NET的形成是一个迄今为止尚未认识到的钙卫蛋白释放的途径。通过比较野生型和钙卫蛋白缺陷动物，我们发现钙卫蛋白对于清除感染至关重要。综上所述，本研究证实了钙卫蛋白的体外抗真菌活性，并进一步证明了它有助于有效的宿主防御C。体内白色念珠菌。我们第一次表明，一定比例的钙卫蛋白结合到NET在体外和体内。中性粒细胞是吞噬细胞，通过吞噬细菌来解除和杀死它们。中性粒细胞的吞噬杀伤机制还没有被很好地描述，它是如何科普那些太大而不能内化的微生物的。值得注意的是，嗜中性粒细胞还可以通过释放嗜中性粒细胞胞外陷阱（NET）来细胞外杀伤或抑制。NET是由染色质（组蛋白和DNA）制成的纤维，装饰有抗菌蛋白。NET诱捕并杀死细菌、真菌和寄生虫等微生物。我们想知道NET是否以及如何控制可以形成大丝状体的致病真菌，如白色念珠菌。我们纯化了所有NET结合蛋白，并鉴定了其中的24个。我们发现钙卫蛋白是主要的抗真菌NET结合蛋白。钙卫蛋白是已知的抗菌，但在这里，我们证明，NET的形成是一种新的释放机制，这种细胞质蛋白。NET基质与真菌密切接触，NET中高局部浓度的钙卫蛋白支持抗真菌活性。此外，在小鼠中，钙卫蛋白对于皮肤、肺和全身感染中对白色念珠菌的有效抗真菌反应是必不可少的。在这些动物的组织切片中，我们检测到NET和NET相关的钙卫蛋白。因此，我们的研究为免疫系统如何应对真菌病原体的机制提供了更多的见解。
Neutrophils are the first line of defense at the site of an infection. They encounter and kill microbes intracellularly upon phagocytosis or extracellularly by degranulation of antimicrobial proteins and the release of Neutrophil Extracellular Traps (NETs). NETs were shown to ensnare and kill microbes. However, their complete protein composition and the antimicrobial mechanism are not well understood. Using a proteomic approach, we identified 24 NET-associated proteins. Quantitative analysis of these proteins and high resolution electron microscopy showed that NETs consist of modified nucleosomes and a stringent selection of other proteins. In contrast to previous results, we found several NET proteins that are cytoplasmic in unstimulated neutrophils. We demonstrated that of those proteins, the antimicrobial heterodimer calprotectin is released in NETs as the major antifungal component. Absence of calprotectin in NETs resulted in complete loss of antifungal activity in vitro. Analysis of three different Candida albicans in vivo infection models indicated that NET formation is a hitherto unrecognized route of calprotectin release. By comparing wild-type and calprotectin-deficient animals we found that calprotectin is crucial for the clearance of infection. Taken together, the present investigations confirmed the antifungal activity of calprotectin in vitro and, moreover, demonstrated that it contributes to effective host defense against C. albicans in vivo. We showed for the first time that a proportion of calprotectin is bound to NETs in vitro and in vivo. Neutrophils are phagocytes that disarm and kill microbes by engulfing them. Less well characterized than their phagocytic killing mechanisms is how neutrophils cope with microbes that are too large to be internalized. Notably, neutrophils may also kill or inhibit extracellularly by releasing Neutrophil Extracellular Traps (NETs). NETs are fibers made of chromatin (histones and DNA) decorated with antimicrobial proteins. NETs ensnare and kill microbes, such as bacteria, fungi and parasites. We wanted to find out if and how NETs control pathogenic fungi that can form large filaments such as Candida albicans. We purified all NET-bound proteins and identified 24 of them. We found that calprotectin is the major antifungal NET-bound protein. Calprotectin was known to be antimicrobial but here we demonstrate that NET formation is a novel release mechanism for this cytoplasmic protein. The NET matrix comes in close contact with the fungi and the high local concentration of calprotectin in the NETs supports the antifungal activity. Furthermore, in mice calprotectin is essential for an efficient antifungal response to Candida albicans in skin, lung and systemic infections. In tissue sections from these animals we detected NETs and NET-associated calprotectin. Thus, our study gives more insights into mechanisms how the immune system copes with fungal pathogens.
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影响因子：
15.9
作者：
DIAMOND, RD;CLARK, RA;HAUDENSCHILD, CC
通讯作者：
HAUDENSCHILD, CC
影响因子：
6.4
作者：
Christin, L;Wysong, DR;Diamond, RD
通讯作者：
Diamond, RD
DOI：
10.1073/pnas.0900226106
发表时间：
2009-04-21
影响因子：
11.1
作者：
Guimaraes-Costa, Anderson B.;Nascimento, Michelle T. C.;Saraiva, Elvira M.
通讯作者：
Saraiva, Elvira M.
影响因子：
5.3
作者：
Ermert, David;Urban, Constantin F.;Brinkmann, Volker
通讯作者：
Brinkmann, Volker
影响因子：
82.9
作者：
Clark, Stephen R.;Ma, Adrienne C.;Kubes, Paul
通讯作者：
Kubes, Paul