鸡突变型和未突变型Mx蛋白体外抗病毒活性研究

Comparison of the In vitro Antiviral Activity Between the Chicken Mutant-Type and Wild-Type Mx Proteins

【目的】研究鸡Mx蛋白第631位氨基酸变异与抗病性的相关性。【方法】用已经构建成功的中国狼山鸡Mx蛋白基因突变型pcDNA3.0-MMx和未突变型pcDNA3.0-Mx真核表达载体分别转染鸡成纤维(CEF)细胞和鼠成纤维(NIH-3T3)细胞;利用RT-PCR检测突变型MMx基因和未突变型Mx基因的表达,微量细胞病变抑制法测定重组蛋白抗新城疫病毒(NDV)和水泡性口炎病毒(VSV)的效果。【结果】诱变鸡Mx基因的表达重组蛋白可保护CEF细胞在孵育48 h内免受NDV的感染;转染突变型pcDNA3.0-MMx真核表达载体的NIH-3T3细胞在孵育60 h内亦未受到VSV的浸染;而转染未突变型pcDNA3.0-Mx真核表达载体的CEF和NIH-3T3细胞在24 h内均发生了病变。【结论】体外重组突变的Mx蛋白在单细胞水平上具有延缓NDV和VSV感染的能力。

【Objective】This experiment was conducted to assess the relationship between the 631 amino acid variations of chicken Mx protein and the disease resistance.【Method】 Mutant pcDNA3.0-MMx and wild-type pcDNA3.0-Mx plasmids of Mx protein gene of Chinese Langshan chicken were constructed successfully,then the CEF and NIH-3T3 cells were transfected by mutant pcDNA3.0-MMx and wild-type pcDNA3.0-Mx plasmids,respectively.RT-PCR was used to identify the expression of mutant MMx and wild-type Mx genes and the mini-cytopathic effect inhibition assay was used to evaluate the antiviral activity for Newcastle disease virus(NDV) and vesicular stomatitis virus(VSV).【Result】 The recombinant Mx protein protected the CEF cells from NDV infection up to 48-h incubation while the NIH-3T3 cells transfected by pcDNA3.0-MMx were not infected by 60-h incubation with VSV,however,the CEF and NIH-3T3 cells transfected by pcDNA3.0-Mx developed pathological changes during 24-h incubation with the viruses.【Conclusion】 Recombinant Mx protein could delay the infection of NDV and VSV at single-cell level.