Cryptococcus neoformans is a human-pathogenic fungal pathogen that causes life-threatening meningoencephalitis in immunocompromised individuals. To investigate the roles of N-glycan core structure in cryptococcal pathogenicity, we constructed mutant strains of C. neoformans with defects in the assembly of lipid-linked N-glycans in the luminal side of the endoplasmic reticulum (ER). Deletion of ALG3 (alg3Δ), which encodes dolichyl-phosphate-mannose (Dol-P-Man)-dependent α-1,3-mannosyltransferase, resulted in the production of truncated neutral N-glycans carrying five mannose residues as a major species. Despite moderate or nondetectable defects in virulence-associated phenotypes in vitro, the alg3Δ mutant was avirulent in a mouse model of systemic cryptococcosis. Notably, the mutant did not show defects in early stages of host cell interaction during infection, including attachment to lung epithelial cells, opsonic/nonopsonic phagocytosis, and manipulation of phagosome acidification. However, the ability to drive macrophage cell death was greatly decreased in this mutant, without loss of cell wall remodeling capacity. Furthermore, deletion of ALG9 and ALG12, encoding Dol-P-Man-dependent α-1,2-mannosyltransferases and α-1,6-mannosyltransferases, generating truncated core N-glycans with six and seven mannose residues, respectively, also displayed remarkably reduced macrophage cell death and in vivo virulence. However, secretion levels of interleukin-1β (IL-1β) were not reduced in the bone marrow-derived dendritic cells obtained from Asc- and Gsdmd-deficient mice infected with the alg3Δ mutant strain, excluding the possibility that pyroptosis is a main host cell death pathway dependent on intact core N-glycans. Our results demonstrated N-glycan structures as a critical feature in modulating death of host cells, which is exploited by as a strategy for host cell escape for dissemination of C. neoformans.

中文翻译：

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通过调节宿主细胞死亡，核心N-聚糖结构对于新型隐球菌的致病性至关重要。

新型隐球菌是一种人类致病性真菌病原体，可在免疫功能低下的个人中引起威胁生命的脑膜脑炎。为了研究N-聚糖核心结构在隐球菌致病性中的作用，我们构建了内含网状结构（ER）腔侧脂质连接的N-聚糖组装中具有缺陷的新隐球菌突变株。删除编码二聚磷酸磷酸甘露糖（Dol-P-Man）依赖的α-1,3-甘露糖基转移酶的ALG3（alg3Δ），导致产生截短的中性N-带有五个甘露糖残基的聚糖是主要种类。尽管在体外与毒力相关的表型存在中度或不可检测的缺陷，但在系统性隐球菌病小鼠模型中，alg3Δ突变体无毒。值得注意的是，该突变体在感染期间宿主细胞相互作用的早期阶段未显示缺陷，包括附着于肺上皮细胞，调理/非调理吞噬作用和吞噬体酸化操作。但是，在这种突变体中，驱动巨噬细胞死亡的能力大大降低，而没有细胞壁重塑能力的丧失。此外，缺失编码Dol-P-Man依赖的α-1,2-甘露糖基转移酶和α-1,6-甘露糖基转移酶的ALG9和ALG12，生成截短的核心具有六个和七个甘露糖残基的N-聚糖也分别显示出显着降低的巨噬细胞死亡和体内毒力。但是，在感染了alg3Δ突变株的Asc和Gsdmd缺陷小鼠的骨髓来源的树突状细胞中，白细胞介素1β（IL-1β）的分泌水平并未降低，但不包括发烧是主要宿主的可能性细胞死亡途径依赖于完整的核心N-聚糖。我们的结果证明Ñ聚糖结构如在调节宿主细胞中，这是由作为宿主细胞逃避的策略用于传播利用死亡的关键特征隐球菌。