摘要
猪繁殖与呼吸综合征病毒(PRRSV)侵入宿主细胞后能够强烈抑制宿主天然免疫反应。早期研究表明,PRRSV 4个非结构蛋白NSP1、NSP2、NSP4和NSP11均可以抑制I型干扰素(IFN-I)的产生,但NSP4抑制IFN-I的机理尚无相关报道。本研究显示PRRSV感染猪肺泡巨噬细胞(PAMs)导致宿主蛋白NEMO(NF-κB essential modulator)被切割,产生一条约40 ku的蛋白条带。通过检测PRRSV编码蛋白酶的切割功能,进一步表明其NSP4是切割NEMO的关键蛋白酶。免疫共沉淀试验表明NSP4与NEMO具有特异性相互作用;激光共聚焦试验结果表明两者在细胞浆中共定位。通过双荧光素酶报告基因试验显示,NSP4过表达可以显著抑制仙台病毒(SeV)和NEMO诱导的IFNβ启动子的激活,导致IFNβ表达量显著下降(p<0.01)。本研究结果表明,PRRSV NSP4可以切割宿主蛋白NEMO,从而显著抑制IFNβ启动子的激活,从分子水平解释了PRRSV NSP4抑制IFNβ产生的机理。
To investigate the molecular mechanism that porcine reproductive and respiratory syndrome virus （PRRSV） inhibits the production of type I interferon （IFN-I）, we detected the expressions of several immunity molecules through western blot and found that host protein of NF-κB essential modulator （NEMO） in porcine alveolar macrophages （PAMs） was cleaved and resulted in decreased expression of IFNβ when infected with PRRSV. In addition, we identified that the NEMO was cleaved by PRRSV NSP4 which had the 3C-like protease activity. While, the interaction and co-localization of NSP4 and NEMO protein was confirmed by the Co-IP assay and confocal examination in co-transfected HEK293 cells with the recombinant expression plasmids.Furthermore, the overexpression of wide-type NSP4 significantly inhibited the activation of IFNβ reporter induced by Sendai vires and NEMO using dual-luciferase reporter gene assay. In conclusion, we found that PRRSV NSP4 cleaved the host protein NEMO and inhibited the production of IFNβ in PAMs.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2014年第3期169-173,共5页
Chinese Journal of Preventive Veterinary Medicine
基金
国家自然科学基金(31172333)
中国农业科学院增量项目(ZGKJ201106)
