Cryptococcus neoformans is rich in polysaccharides of the cell wall and capsule. Dectin-2 recognizes high-mannose polysaccharides and plays a central role in the immune response to fungal pathogens. Previously, we demonstrated Dectin-2 was involved in the activation of dendritic cells upon stimulation with C. neoformans, suggesting the existence of a ligand recognized by Dectin-2. In the present study, we examined the cell wall structures of C. neoformans contributing to the Dectin-2-mediated activation of immune cells. In a NFAT-GFP reporter assay of the reported cells expressing Dectin-2, the lysates, but not the whole yeast cells, of an acapsular strain of C. neoformans (Cap67) delivered Dectin-2-mediated signaling. This activity was detected in the supernatant of β-glucanase-treated Cap67 and more strongly in the semi-purified polysaccharides of this supernatant using ConA-affinity chromatography (ConA-bound fraction), in which a large amount of saccharides, but not protein, were detected. Treatment of this supernatant with periodic acid and the addition of excessive mannose, but not glucose or galactose, strongly inhibited this activity. The ConA-bound fraction of the β-glucanase-treated Cap67 supernatant was bound to Dectin-2-Fc fusion protein in a dose-dependent manner and strongly induced the production of interleukin-12p40 and tumour necrosis factor-α by dendritic cells; this was abrogated under the Dectin-2-deficient condition. Finally, 98 kDa mannoprotein (MP98) derived from C. neoformans showed activation of the reporter cells expressing Dectin-2. These results suggested that a ligand with mannose moieties may exist in the cell walls and play a critical role in the activation of dendritic cells during infection with C. neoformans.

中文翻译：

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Dectin-2介导的信号由新隐球菌的细胞壁多糖触发。

新型隐球菌富含细胞壁和胶囊的多糖。Dectin-2识别高甘露糖多糖，并在对真菌病原体的免疫反应中发挥重要作用。以前，我们证明Dectin-2在用新孢梭菌刺激后参与树突状细胞的活化，这表明存在由Dectin-2识别的配体。在本研究中，我们检查了新形成梭状芽胞杆菌的细胞壁结构，这些细胞壁结构有助于Dectin-2介导的免疫细胞活化。在报道的表达Dectin-2的细胞的NFAT-GFP报告基因分析中，新孢梭菌（Cap67）荚膜菌株的裂解物而非整个酵母细胞传递了Dectin-2介导的信号。使用ConA亲和色谱法（ConA结合级分）在β-葡聚糖酶处理过的Cap67的上清液中检测到此活性，并在该上清液的半纯化多糖中更强地检测到了该活性，其中有大量的糖而不是蛋白质，被检测到。用高碘酸处理该上清液并添加过量的甘露糖而不是葡萄糖或半乳糖，强烈抑制了该活性。β-葡聚糖酶处理的Cap67上清液的ConA结合部分与Dectin-2-Fc融合蛋白以剂量依赖性方式结合，并强烈诱导树突状细胞产生白介素12p40和肿瘤坏死因子-α。在Dectin-2缺乏的情况下将其取消。最后，源自新孢子虫的98 kDa甘露糖蛋白（MP98）显示表达Dectin-2的报告细胞的激活。