Deciphering the Intracellular Niche of Cryptococcus neoformans in Alveolar Macrophages

Cryptococcus neoformans is a fungal pathogen responsible for ~200,000 deaths yearly. This ubiquitously present yeast affects immunocompromised individuals, which left untreated is invariably fatal. Once C. neoformans particles are inhaled and make their way into the lungs they encounter alveolar macrophages. Here, interactions influence whether the infection is controlled or disseminates to cause disease. Since C. neoformans can survive and replicate inside host macrophages, they represent a safe haven, hidden from the immune response of the body. The mechanisms C. neoformans use to mediate intracellular parasitism are not fully understood. Even less is known about how immune signals impact this niche. Therefore, we aim to gain a better understanding of the macrophage–cryptococcal interactions and decipher the properties of the cryptococcal-containing phagosome (CCP), which may be key to determining infection outcome. We identified three populations of CCPs with different behaviors: one that gains acidification after phagocytosis but subsequently loses it; some that acidify and remain acidic; and some that never acidify. We also directly observed phagosomal membrane damage, suggesting a possible mechanism for pH manipulation. This contrasts with phagosomes containing S. cerevisiae, which rapidly acidify, stay acidic, and have low levels of membrane damage. Moreover, we have identified a population of CCPs that display both early endosomal lipid and lysosomal protein characteristics, a combination not normally observed. This suggests C. neoformans can alter its phagosome and provides a potential mechanism for intracellular survival that may be driving pathogenesis. We have demonstrated a delay in acquisition of lysosomal markers to CCPs. When macrophages are stimulated with interferon-?, acquisition of lysosomal markers occurs more rapidly. Host Rab20 has been shown to be interferon-? inducible and appears to facilitate this promotion of phago-lysosomal fusion and may be key in influencing the infection, thereby linking host immune status and phagosomal properties during cryptococcal infection.

新型隐球菌（Cryptococcus neoformans）是一种致病原真菌，每年导致约20万人死亡。这种普遍存在的酵母菌主要影响免疫功能低下的人群，若未得到及时治疗，则往往导致死亡。一旦新型隐球菌颗粒被吸入并进入肺部，它们便会遭遇肺泡巨噬细胞。在此，这些相互作用将决定感染是被控制还是扩散至全身引起疾病。鉴于新型隐球菌能够在宿主巨噬细胞内存活和繁殖，它们构成了一个安全的避难所，免受机体免疫反应的侵害。新型隐球菌用于介导细胞内寄生的机制尚不完全明了。关于免疫信号如何影响这一生态位，了解更是甚少。因此，我们的目标是深入理解巨噬细胞与新型隐球菌之间的相互作用，并解析包含新型隐球菌的噬胞体（CCP）的特性，这可能是决定感染结果的关键。我们已鉴定出三种具有不同行为的CCP群体：一种在吞噬后酸化但随后失去酸化能力；一些酸化并保持酸性；还有一些从未酸化。我们还直接观察到噬胞体膜损伤，这表明可能存在pH操纵的机制。这与含有酿酒酵母（S. cerevisiae）的噬胞体形成对比，后者迅速酸化、保持酸性并具有低水平的膜损伤。此外，我们还发现了一群同时表现出早期内质网脂质和溶酶体蛋白特性的CCP，这种组合通常不会观察到。这表明新型隐球菌可以改变其噬胞体，并提供了一种可能促进细胞内存活的机制，这可能是致病过程驱动的因素。我们已经证明了溶酶体标记在CCP上的获取存在延迟。当巨噬细胞受到干扰素-γ的刺激时，溶酶体标记的获取速度会更快。宿主Rab20已被证明是干扰素-γ诱导的，并且似乎有助于促进噬胞-溶酶体融合，这可能在影响感染过程中发挥关键作用，从而将宿主免疫状态与新型隐球菌感染过程中的噬胞体特性联系起来。