Mattion N M, Mitchell D B, Both G W, Estes M K
Baylor College of Medicine, Division of Molecular Virology, Houston, Texas 77030.
Virology. 1991 Mar;181(1):295-304. doi: 10.1016/0042-6822(91)90495-w.
The nucleotide sequence of rotavirus genome segment 11 shows that this gene contains three potential open reading frames. We used several approaches to determine whether any polypeptides other than NS26, the primary protein product, are expressed. In particular, we sought to determine whether the strong out-of-phase start codon present at nucleotides 80-82, which would encode a protein of 92 amino acids, is used in vivo or in cell-free systems. Several modifications of gene 11 were made and found to produce proteins from the different initiation codons in cell-free transcription-translation systems. The protein from the out-of-phase open reading frame was shown to be expressed in rotavirus-infected MA104 cells; this was demonstrated using monospecific sera prepared to this protein expressed in Spodoptera frugiperda insect cells infected with a baculovirus recombinant containing only the out-of-phase open reading frame. The origin of some of the lower-molecular-weight bands serologically related to the primary product of gene 11, NS26, was also studied by selective immunoprecipitation using two different sera made from recombinant baculovirus lysates. All of these polypeptides are present in infected cells in a complex which is still incompletely defined.
轮状病毒基因组第11节段的核苷酸序列表明，该基因包含三个潜在的开放阅读框。我们采用了几种方法来确定除主要蛋白产物NS26之外是否还表达其他多肽。具体而言，我们试图确定位于核苷酸80 - 82处的强异相起始密码子（该密码子可编码一个92个氨基酸的蛋白质）在体内或无细胞系统中是否被使用。我们对基因11进行了几处修饰，并发现其在无细胞转录 - 翻译系统中可从不同的起始密码子产生蛋白质。结果表明，来自异相开放阅读框的蛋白质在轮状病毒感染的MA104细胞中表达；这是通过使用针对在仅含有异相开放阅读框的杆状病毒重组体感染的草地贪夜蛾昆虫细胞中表达的该蛋白质制备的单特异性血清来证明的。我们还通过使用由重组杆状病毒裂解物制备的两种不同血清进行选择性免疫沉淀，研究了一些与基因11的主要产物NS26血清学相关的低分子量条带的来源。所有这些多肽都存在于感染细胞中的一个复合物中，该复合物的具体情况仍未完全明确。
