V-ATPase, which comprises 13-14 subunits, is essential for pH homeostasis in all eukaryotes, but its proper function requires a regulator to assemble its subunits. While RAVE (regulator of H+-ATPase of vacuolar and endosomal membranes) and Raboconnectin-3 complexes assemble V-ATPase subunits in Saccharomyces cerevisiae and humans, respectively, the function of the RAVE complex in fungal pathogens remains largely unknown. In this study, we identified two RAVE complex components, Rav1 and Wdr1, in the fungal meningitis pathogen Cryptococcus neoformans, and analyzed their roles. Rav1 and Wdr1 are orthologous to yeast RAVE and human Rabconnectin-3 counterparts, respectively, forming the hybrid RAVE (hRAVE) complex. Deletion of RAV1 caused severe defects in growth, cell cycle control, morphogenesis, sexual development, stress responses, and virulence factor production, while the deletion of WDR1 resulted in similar but modest changes, suggesting that Rav1 and Wdr1 play central and accessary roles, respectively. Proteomics analysis confirmed that Wdr1 was one of the Rav1-interacting proteins. Although the hRAVE complex generally has V-ATPase-dependent functions, it also has some V-ATPase-independent roles, suggesting a unique role beyond conventional intracellular pH regulation in C. neoformans. The hRAVE complex played a critical role in the pathogenicity of C. neoformans, and RAV1 deletion attenuated virulence and impaired blood-brain barrier crossing ability. This study provides comprehensive insights into the pathobiological roles of the fungal RAVE complex and suggests a novel therapeutic strategy for controlling cryptococcosis.

中文翻译：

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杂种 RAVE 复合物在新型隐球菌中发挥 V-ATP 酶依赖性和非依赖性病理生物学作用。

V-ATP 酶由 13-14 个亚基组成，对于所有真核生物的 pH 稳态至关重要，但其正常功能需要调节器来组装其亚基。虽然 RAVE（液泡和内体膜的 H+-ATP 酶调节剂）和 Raboconnectin-3 复合物分别在酿酒酵母和人类中组装 V-ATP 酶亚基，但 RAVE 复合物在真菌病原体中的功能仍然很大程度上未知。在本研究中，我们在真菌性脑膜炎病原体新型隐球菌中鉴定了两个 RAVE 复杂成分 Rav1 和 Wdr1，并分析了它们的作用。Rav1 和 Wdr1 分别与酵母 RAVE 和人 Rabconnectin-3 对应物直系同源，形成杂合 RAVE (hRAVE) 复合物。RAV1 的缺失导致生长、细胞周期控制、形态发生、性发育、应激反应和毒力因子产生的严重缺陷，而 WDR1 的缺失导致类似但适度的变化，表明 Rav1 和 Wdr1 分别发挥核心和辅助作用。蛋白质组学分析证实 Wdr1 是 Rav1 相互作用蛋白之一。虽然 hRAVE 复合物通常具有 V-ATP 酶依赖性功能，但它也具有一些 V-ATP 酶独立作用，这表明在新型隐球菌中，hRAVE 复合物具有超越传统细胞内 pH 调节的独特作用。hRAVE 复合物在新型隐球菌的致病性中发挥着关键作用，RAV1 缺失会减弱毒力并损害血脑屏障穿越能力。这项研究提供了对真菌 RAVE 复合体病理生物学作用的全面见解，并提出了一种控制隐球菌病的新治疗策略。