T103A变异MxA蛋白抑制水泡性口膜炎病毒复制的体外研究

In vitro Inhibition of VSV Replication by the T103A Mutant MxA Protein

目的研究T103A变异MxA蛋白抑制水疱性口膜炎病毒(VSV)复制活性。方法将野生型、T103A变异MxA蛋白表达载体和对照质粒分别瞬时转染Wish细胞,24h后VSV感染细胞,48h后用MTT法检测各组细胞增殖;另取Wish细胞转染上述3种质粒,转染24h加入VSV感染细胞,24h后收集细胞采用RT-PCR检测VSV mRNA水平;Western blot检测各组MxA蛋白表达。结果野生型、T103A变异MxA蛋白均在Wish细胞有较好表达;MTT检测结果提示T103A变异组细胞增殖数显著低于野生型组(P<0.01);RT-PCR结果显示T103A变异组VSVmRNA水平显著高于野生型组(P<0.01),但与对照组比较差异无统计学意义(P>0.05)。结论T103A变异MxA蛋白失去了抑制VSV复制活性。

Objective To investigate the antiviral activity of T103A mutant MxA protein in vesicular stomatitis virus （VSV） infected cells. Methods Wish cells were transfected with control vector or pcDNA3.1-MxA carrying the wild-type or T103A mutant MxA gene. Twenty-four hours after transfection, the cells were infected with VSV and the viability of the infected cells was determined By the MTT reduction method at 48 h after infction. Level of VSV mRNA was determined by RT-PCR at 24 h after VSV infection. Results Both wild-type MxA and T103A mutant proteins were expressed in infected cells. The viability of cells expressing T103A mutant proteins was markedly lower than the cells expressing the wild-type proteins （P〈0.01）. Similar to the control group, the level of VSV mRNA in T103A mutant protein expressing cells was markedly higher than the cells expressing the wild-type proteins （P〈0.01）. Conclusion T103A mutation is responsible for the lost of antiviral activity of the MxA protein.