Veletanlic Vanesa, Sartalamacchia Kylie, Diller Julia R, Ogden Kristen M
Department of Pediatrics, Vanderbilt University Medical Center, Nashville, Tennessee, USA.
Department of Pathology, Microbiology, and Immunology, Vanderbilt University Medical Center, Nashville, Tennessee, USA.
bioRxiv. 2023 Apr 8:2023.04.07.536061. doi: 10.1101/2023.04.07.536061.
Fusion-associated small transmembrane (FAST) proteins are viral nonstructural proteins that mediate cell-cell fusion to form multinucleated syncytia. We previously reported that human species B rotavirus NSP1-1 is a FAST protein that induces syncytia in primate epithelial cells but not rodent fibroblasts. We hypothesized that the NSP1-1 proteins of other rotavirus species could also mediate cell-cell fusion and that fusion activity might be limited to cell types derived from homologous hosts. To test this hypothesis, we predicted the structure and domain organization of NSP1-1 proteins of species B rotavirus from a human, goat, and pig, species G rotavirus from a pigeon and turkey, and species I rotavirus from a dog and cat. We cloned these sequences into plasmids and transiently expressed the NSP1-1 proteins in avian, canine, hamster, human, porcine, and simian cells. Regardless of host origin of the virus, each NSP1-1 protein induced syncytia in primate cells, while few induced syncytia in other cell types. To identify the domains that determined cell-specific fusion activity for human species B rotavirus NSP1-1, we engineered chimeric proteins containing domain exchanges with the p10 FAST protein from Nelson Bay orthoreovirus. Using the chimeric proteins, we found that the N-terminal and transmembrane domains determined the cell type specificity of fusion activity. Although the species and cell type criteria for fusion activity remain unclear, these findings suggest that rotavirus species B, G, and I NSP1-1 are functional FAST proteins whose N termini play a role in specifying the cells in which they mediate syncytia formation.
融合相关小跨膜（FAST）蛋白是介导细胞间融合以形成多核巨细胞的病毒非结构蛋白。我们之前报道过，人B组轮状病毒NSP1-1是一种能在灵长类上皮细胞中诱导形成巨细胞但不能在啮齿类成纤维细胞中诱导形成巨细胞的FAST蛋白。我们推测，其他轮状病毒种类的NSP1-1蛋白也可能介导细胞间融合，并且融合活性可能仅限于源自同源宿主的细胞类型。为了验证这一假设，我们预测了来自人、山羊和猪的B组轮状病毒、来自鸽子和火鸡的G组轮状病毒以及来自狗和猫的I组轮状病毒的NSP1-1蛋白的结构和结构域组织。我们将这些序列克隆到质粒中，并在禽、犬、仓鼠、人、猪和猿猴细胞中瞬时表达NSP1-1蛋白。无论病毒的宿主来源如何，每种NSP1-1蛋白都能在灵长类细胞中诱导形成巨细胞，而很少能在其他细胞类型中诱导形成巨细胞。为了确定决定人B组轮状病毒NSP1-1细胞特异性融合活性的结构域，我们构建了与尼尔森湾正呼肠孤病毒的p10 FAST蛋白进行结构域交换的嵌合蛋白。使用这些嵌合蛋白，我们发现N端和跨膜结构域决定了融合活性的细胞类型特异性。尽管融合活性的物种和细胞类型标准仍不清楚，但这些发现表明，B组、G组和I组轮状病毒的NSP1-1是功能性FAST蛋白，其N端在指定它们介导巨细胞形成的细胞中发挥作用。
