狂犬病病毒HEP-Flury M基因重排后在小鼠神经母细胞瘤细胞中的表型分析

Phenotypic consequences of M gene rearrangement of rabies virus HEP-Flury in mouse neuroblastoma cells

【目的】探究狂犬病病毒HEP-Flury M基因重排对基因转录和蛋白表达的影响,揭示病毒在小鼠神经母细胞瘤(NA)细胞中的表型变化与M基因重排的相关性。【方法】通过荧光定量PCR、Western blot以及病毒在NA细胞中的生长和扩散试验,对亲本毒株rHEP-Flury和M基因重排毒株M2、M4在NA细胞中的基因转录、表达、生长和扩散进行比较。【结果】狂犬病病毒结构基因的转录和表达主要受病毒基因组RNA合成的影响,但是在一个完整转录过程中单个结构基因的转录比例与其所在位置相关,M基因重排病毒的Leader RNA(LeRNA)和L mRNA的转录比例显著高于亲本毒株rHEP-Flury。M基因重排病毒在NA细胞中的生长和扩散都劣于亲本毒株rHEPFlury。【结论】狂犬病病毒亲本毒株rHEP-Flury具有狂犬病病毒原始的基因组顺序,在NA细胞中的生长和扩散都明显优于M基因重排病毒。结构基因在基因组中的位置主要决定其在一次转录过程中的转录比例,进而影响病毒在NA细胞中的生长和扩散。

【Objective】To explore the effects of M gene rearrangement of rabies virus HEP-Flury on gene transcription and protein expression,reveal the correlation between phenotypic consequences of virus in mouse neuroblastoma(NA)cells and M gene rearrangement.【Method】The gene transcription,expression,growth and spread of parent strain rHEP-Flury and M gene rearranged virus stains(M2 and M4)in infected NA cells were compared by fluorescence quantitative PCR,Western blot,growth and spread experiments of virus in NA cells.【Result】The transcription and expression of rabies virus structural gene were mainly affected by RNA synthesis of virus genome.The gene position mainly affected the ratio of transcription in a complete transcription process.The transcription ratios of leader RNA(LeRNA)and L mRNA of M gene rearranged viruses were significanthy higher than those of rHEP-Flury.The growth and spread of M gene rearranged viruses in NA cells were inferiorer than rHEP-Flury.【Conclusion】rHEP-Flury has wild-type gene order of rabies virus.Its growth and spread in NA cells are superiorer to M gene rearranged viruses.The position of structural gene in genome determines its transcription ratio in a complete transcription process and affects the growth and spread of virus in NA cells.