N蛋白磷酸化位点突变影响猪繁殖与呼吸综合征病毒的复制及亚基因组的转录

doi:10.11843/j.issn.0366-6964.2019.12.011

摘要/Abstract

摘要： 为探究猪繁殖与呼吸综合征病毒（porcine reproductive and respiratory syndrome virus，PRRSV）的N蛋白磷酸化修饰是否对病毒的复制及亚基因组的转录产生影响，将XH-GD PRRSV N蛋白的已知磷酸化位点Ser-105和Ser-120分别或同时突变为Ala，拯救突变病毒，命名为A105、A120、A105-120，测定其生长曲线和亚基因组的转录水平。结果显示：突变病毒在MARC-145的病毒复制速率与滴度显著低于亲本毒株（P<0.01或P<0.05），而在原代PAMs复制速率降低（P<0.01），不影响病毒最终的滴度。突变毒株的基因组RNA（genomic RNA，gRNA）转录水平显著降低（P<0.01或P<0.05），磷酸化位点突变能显著降低长链亚基因组mRNA（subgenomic RNA，sgmRNA）2、3的转录（P<0.01或P<0.05），但短链sgmRNA4、5表达量随时间呈增加趋势。以gRNA表达量为基准对各亚基因组表达量进行整体分析，发现N蛋白磷酸化显著影响了sgmRNA4和sgmRNA5的表达（P<0.01或P<0.05）。结果提示，N蛋白磷酸化位点突变影响PRRSV的复制及亚基因组转录。

关键词: 猪繁殖与呼吸综合征病毒, N蛋白, 磷酸化, 亚基因组

Abstract: To explore whether the N-protein phosphorylation of porcine reproductive and respiratory syndrome virus (PRRSV) affects viral replication, the ser-105 and ser-120 of the PRRSV N-protein were mutated into alanine alone or simultaneously. The rescued viruses were named as A105, A120 and A105-120, and their growth curve and subgenomic transcription level were determined respectively. The viral replication rate and titer of the mutant virus in MARC-145 were significantly lower than that of parental strain PRRSV XH-GD (P<0.01 or P<0.05). While in the primary PAM cells, the replication rate of the mutant virus decreased (P<0.01), but the final virus titer was not affected. The genomic (g) RNA transcription level of the mutant virulence group was significantly decreased (P<0.01 or P<0.05), and the phosphorylation site mutation significantly reduced the transcription of the longer subgenomic mRNA (sgmRNA)2/3 (P<0.01 or P<0.05), but increased the expression levels of the shorter sgmRNA 4/5 with time. Based on the gRNA expression level, the expression of sgmRNAs expression was analyzed. Our current work showed that the N-protein phosphorylation significantly affects the expression of sgmRNA4/5(P<0.01 or P<0.05). Mutation of N protein phosphorylation site affects PRRSV replication and subgenomic transcription.

Key words: porcine reproductive and respiratory syndrome virus, N protein, phosphorylation, subgenome

中图分类号:

S852.659.6

易和友, 于之清, 陈耀, 李琪, 韦应芳, 张桂红. N蛋白磷酸化位点突变影响猪繁殖与呼吸综合征病毒的复制及亚基因组的转录[J]. 畜牧兽医学报, 2019, 50(12): 2470-2478.

YI Heyou, YU Zhiqing, CHEN Yao, LI Qi, WEI Yingfang, ZHANG Guihong. Mutation of N Protein Phosphorylation Site Affects Replication and Subgenomic Transcription of Porcine Reproductive and Respiratory Syndrome Virus[J]. Acta Veterinaria et Zootechnica Sinica, 2019, 50(12): 2470-2478.

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