Department of BioSciences, Rice University , Houston, Texas, USA.
Departamento de Genética del Desarrollo y Fisiología Molecular, Universidad Nacional Autónoma de México , Cuernavaca, Morelos, Mexico.
J Virol. 2023 Aug 31;97(8):e0080223. doi: 10.1128/jvi.00802-23. Epub 2023 Jul 28.
The human astrovirus (HAstV) is a non-enveloped, single-stranded RNA virus that is a common cause of gastroenteritis. Most non-enveloped viruses use membrane disruption to deliver the viral genome into a host cell after virus uptake. The virus-host factors that allow for HAstV cell entry are currently unknown but thought to be associated with the host-protease-mediated viral maturation. Using liposome disruption analysis, we identified a trypsin-dependent lipid disruption activity in the capsid protein of HAstV serotype 8. This function was further localized to the P1 domain of the viral capsid core, which was both necessary and sufficient for membrane disruption. Site-directed mutagenesis identified a cluster of four trypsin cleavage sites necessary to retain the lipid disruption activity, which is likely attributed to a short stretch of sequence ending at arginine 313 based on mass spectrometry of liposome-associated peptides. The membrane disruption activity was conserved across several other HAstVs, including the emerging VA2 strain, and effective against a wide range of lipid identities. This work provides key functional insight into the protease maturation process essential to HAstV infectivity and presents a method to investigate membrane penetration by non-enveloped viruses . IMPORTANCE Human astroviruses (HAstVs) are an understudied family of viruses that cause mild gastroenteritis but have recent cases associated with a more severe neural pathogenesis. Many important elements of the HAstV life cycle are not well understood, and further elucidating them can help understand the various forms of HAstV pathogenesis. In this study, we utilized an liposome-based assay to describe and characterize a previously unreported lipid disruption activity. This activity is dependent on the protease cleavage of key sites in HAstV capsid core and can be controlled by site-directed mutagenesis. Our group observed this activity in multiple strains of HAstV and in multiple lipid conditions, indicating this may be a conserved activity across the AstV family. The discovery of this function provides insight into HAstV cellular entry, pathogenesis, and a possible target for future therapeutics.
人星状病毒（HAstV）是一种无包膜的单链 RNA 病毒，是引起肠胃炎的常见原因。大多数无包膜病毒在病毒摄取后，通过破坏膜来将病毒基因组递送到宿主细胞中。允许 HAstV 细胞进入的病毒-宿主因子目前尚不清楚，但被认为与宿主蛋白酶介导的病毒成熟有关。使用脂质体破坏分析，我们在 HAstV 血清型 8 的衣壳蛋白中鉴定出一种依赖于胰蛋白酶的脂质破坏活性。该功能进一步定位于病毒衣壳核心的 P1 结构域，该结构域对于膜破坏既必要又充分。定点突变鉴定出一组四个胰蛋白酶切割位点对于保留脂质破坏活性是必需的，这可能归因于基于脂质体相关肽的质谱分析的精氨酸 313 末端的短序列。该膜破坏活性在包括新兴的 VA2 株在内的几种其他 HAstV 中是保守的，并且对广泛的脂质种类有效。这项工作为 HAstV 感染性所必需的蛋白酶成熟过程提供了关键的功能见解，并提出了一种研究无包膜病毒膜穿透的方法。
人星状病毒（HAstV）是一组研究不足的病毒，它们引起轻度肠胃炎，但最近有病例与更严重的神经发病机制有关。HAstV 生命周期的许多重要元素还不是很清楚，进一步阐明这些元素可以帮助我们了解各种形式的 HAstV 发病机制。在这项研究中，我们利用脂质体测定法来描述和表征以前未报道的脂质破坏活性。这种活性依赖于 HAstV 衣壳核心关键位点的蛋白酶切割，并且可以通过定点突变来控制。我们的研究小组在多种 HAstV 株和多种脂质条件下观察到这种活性，表明这可能是 AstV 家族的一种保守活性。这种功能的发现为 HAstV 细胞进入、发病机制以及未来治疗的可能靶点提供了新的见解。
