While it is well appreciated that late domains in the viral matrix proteins are crucial to mediate efficient virus budding, little is known about roles of late domains in the viral nucleocapsid proteins. Here, we characterized the functional relevance of a YxxL motif with potential late-domain function in the Ebola virus nucleocapsid protein VP24. Mutations in the YxxL motif had two opposing effects on the functions of VP24. On the one hand, the mutation affected the regulatory function of VP24 in viral RNA transcription and replication, which correlated with an increased incorporation of minigenomes into released transcription- and replication-competent virus-like particles (trVLPs). Consequently, cells infected with those trVLPs showed higher levels of viral transcription. On the other hand, mutations of the YxxL motif greatly impaired the intracellular transport of nucleocapsid-like structures (NCLSs) composed of the viral proteins NP, VP35, and VP24 and the length of released trVLPs. Attempts to rescue recombinant Ebola virus expressing YxxL-deficient VP24 failed, underlining the importance of this motif for the viral life cycle.IMPORTANCE Ebola virus (EBOV) causes a severe fever with high case fatality rates and, so far, no available specific therapy. Understanding the interplay between viral and host proteins is important to identify new therapeutic approaches. VP24 is one of the essential nucleocapsid components and is necessary to regulate viral RNA synthesis and condense viral nucleocapsids before their transport to the plasma membrane. Our functional analyses of the YxxL motif in VP24 suggested that it serves as an interface between nucleocapsid-like structures (NCLSs) and cellular proteins, promoting intracellular transport of NCLSs in an Alix-independent manner. Moreover, the YxxL motif is necessary for the inhibitory function of VP24 in viral RNA synthesis. A failure to rescue EBOV encoding VP24 with a mutated YxxL motif indicated that the integrity of the YxxL motif is essential for EBOV growth. Thus, this motif might represent a potential target for antiviral interference.

中文翻译：

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埃博拉病毒VP24的YxxL母本的完整性对于像核衣壳的结构运输和病毒RNA合成的调控很重要。

尽管众所周知病毒基质蛋白中的晚期结构域对于介导有效的病毒出芽至关重要，但对于病毒核衣壳蛋白中晚期结构域的作用知之甚少。在这里，我们表征了与埃博拉病毒核衣壳蛋白VP24中潜在的后域功能的YxxL母题的功能相关性。YxxL基序中的突变对VP24的功能有两个相反的影响。一方面，这种突变影响了VP24在病毒RNA转录和复制中的调控功能，这与将微型基因组掺入释放的具有转录和复制能力的病毒样颗粒（trVLP）中有关。因此，被那些trVLPs感染的细胞显示出更高水平的病毒转录。另一方面，YxxL基序的突变极大地损害了由病毒蛋白NP，VP35和VP24组成的核衣壳样结构（NCLSs）的细胞内运输以及释放的trVLP的长度。试图挽救表达YxxL缺失VP24的重组埃博拉病毒的尝试失败，突显了该基序在病毒生命周期中的重要性。重要事项埃博拉病毒（EBOV）引起严重发烧，病死率很高，到目前为止，尚无可用的特异性疗法。了解病毒蛋白和宿主蛋白之间的相互作用对于确定新的治疗方法很重要。VP24是必不可少的核衣壳成分之一，在调节病毒RNA的合成和将病毒核衣壳转运到质膜之前必须进行浓缩。我们对VP24中YxxL基序的功能分析表明，它可作为核衣壳样结构（NCLSs）与细胞蛋白之间的接口，以Alix无关的方式促进NCLSs的细胞内转运。而且，YxxL基序对于VP24在病毒RNA合成中的抑制功能是必需的。无法挽救具有突变的YxxL基序的EBOV编码VP24的信号表明YxxL基序的完整性对于EBOV生长至关重要。因此，该基序可能代表抗病毒干扰的潜在目标。无法挽救具有突变的YxxL基序的EBOV编码VP24的信号表明YxxL基序的完整性对于EBOV生长至关重要。因此，该基序可能代表抗病毒干扰的潜在目标。无法挽救具有突变的YxxL基序的EBOV编码VP24的信号表明YxxL基序的完整性对于EBOV生长至关重要。因此，该基序可能代表抗病毒干扰的潜在目标。