The nuclear-cytoplasmic trafficking of matrix proteins (M) is essential for henipavirus budding, with M protein ubiquitination playing a pivotal role in this dynamic process. Despite its importance, the intricacies of the M ubiquitination cascade have remained elusive. In this study, we elucidate a novel mechanism by which Nipah virus (NiV), a highly pathogenic henipavirus, utilizes a ubiquitination complex involving the E2 ubiquitin-conjugating enzyme RAD6A and the E3 ubiquitin ligase RAD18 to ubiquitinate the virus's M protein, thereby facilitating its nuclear-cytoplasmic trafficking. We demonstrate that RAD18 interacts with RAD6A, enabling the latter to supply ubiquitins for the RAD18-mediated transfer of ubiquitin to M through RAD18-M interactions. Specifically, M is ubiquitinated by the RAD6A-RAD18 complex at lysine (K) 258 through a K63-linked ubiquitination, a modification crucial for M's function. This ubiquitination drives M's relocation to the cytoplasm, directing it to plasma membranes for effective viral egress. Conversely, disrupting the RAD6A-RAD18-M axis, mutating RAD18's E3 ligase activity, or inhibiting RAD6A activity with TZ9 (a RAD6-ubiquitin thioester formation inhibitor) impairs M ubiquitination, resulting in defective nuclear export and budding of NiV. Significantly, live NiV and Hendra virus infection is attenuated in RAD18 knockout cells or in cells treated with TZ9, highlighting the critical physiological role of RAD6A-RAD18-mediated M ubiquitination in the henipavirus life cycle. Our findings not only reveal how NiV manipulates a nucleus-localized ubiquitination complex to promote virus's M protein ubiquitination and nuclear export, but also suggest that the small molecule inhibitor TZ9 could serve as a potential therapeutic against henipavirus infection.

中文翻译：

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E3 连接酶 RAD18 介导的亨尼帕病毒基质蛋白泛素化促进其核质运输和病毒出口。


基质蛋白 （M） 的核细胞质运输对于亨尼帕病毒出芽至关重要，M 蛋白泛素化在这个动态过程中起着关键作用。尽管 M 泛素化级联反应很重要，但它的复杂性仍然难以捉摸。在这项研究中，我们阐明了一种新的机制，即尼帕病毒 （NiV） 是一种高致病性亨尼帕病毒，利用涉及 E2 泛素结合酶 RAD6A 和 E3 泛素连接酶 RAD18 的泛素化复合物来泛素化病毒的 M 蛋白，从而促进其核质运输。我们证明 RAD18 与 RAD6A 相互作用，使后者能够通过 RAD18-M 相互作用为 RAD18 介导的泛素向 M 的转移提供泛素。具体来说，M 通过 K63 连接的泛素化被赖氨酸 （K） 258 位点的 RAD6A-RAD18 复合物泛素化，这是一种对 M 功能至关重要的修饰。这种泛素化驱动 M 重新定位到细胞质，将其引导到质膜以有效传播病毒。相反，破坏 RAD6A-RAD18-M 轴、突变 RAD18 的 E3 连接酶活性或用 TZ9（一种 RAD6-泛素硫酯形成抑制剂）抑制 RAD6A 活性会损害 M 泛素化，导致核输出缺陷和 NiV 出芽。值得注意的是，在 RAD18 敲除细胞或用 TZ9 处理的细胞中，活 NiV 和 Hendra 病毒感染减弱，突出了 RAD6A-RAD18 介导的 M 泛素化在亨尼帕病毒生命周期中的关键生理作用。 我们的研究结果不仅揭示了 NiV 如何操纵位于细胞核的泛素化复合物以促进病毒的 M 蛋白泛素化和核输出，而且还表明小分子抑制剂 TZ9 可以作为针对亨尼帕病毒感染的潜在治疗方法。