摘要
目的探讨基于维甲酸诱导基因Ⅰ样受体(RLR)信号通路的去泛素化酶对肠道病毒71型(EV71)感染的调控机制。方法将人横纹肌肉瘤细胞A-204分为空白对照组、空载质粒组、去泛素化酶组,空白对照组不作任何处理,空载质粒组转染10 nmol/L空载质粒48 h后加入EV71(MOI=0.5),去泛素化酶组转染10 nmol/L去泛素化酶USP4过表达质粒48 h后加入EV71(MOI=0.5)。采用RT-PCR检测各组细胞中EV71 mRNA表达,Western blot检测各组细胞USP4、维甲酸诱导基因Ⅰ(RIG-Ⅰ)、黑色素瘤分化相关基因5(MDA5)、LGP2蛋白表达。结果空载质粒组细胞USP4蛋白表达明显低于空白对照组(P<0.05),去泛素化酶组细胞USP4蛋白表达明显高于空载质粒组和空白对照组(P<0.05)。空载质粒组和去泛素化酶组细胞EV71 mRNA表达明显高于空白对照组(P<0.05),去泛素化酶组细胞EV71 mRNA表达明显低于空载质粒组(P<0.05)。空载质粒组细胞RIG-Ⅰ、MDA5、LGP-2蛋白表达明显低于空白对照组(P<0.05),去泛素化酶组细胞RIG-Ⅰ、MDA5、LGP-2蛋白表达明显高于空载质粒组和空白对照组(P<0.05)。结论去泛素化酶USP4可能通过激活RLR信号通路而发挥抗EV71感染作用。
Objective To investigate the regulatory mechanism of deubiquitinase enzyme on enterovirus 71(EV71)infection based on retinoic-acid-inducible geneⅠ-like receptor(RLR)signaling pathway.Methods Human rhabdomyosarcoma cells A-204 were divided into the blank control group,empty load plasmid group and deubiquitinating enzyme group.The blank control group had no treatment;the empty plasmid group was transfected with 10 nmol/L empty plasmid for 48 h before adding EV71(MOI=0.5);the deubiquitinating enzyme group was transfected with 10 nmol/L deubiquitinating enzyme USP4 overexpression plasmid for 48 h before adding EV71(MOI=0.5);real-time quantitative reverse transcription polymerase chain reaction(RT-PCR)was used to detect the expression of EV71 mRNA in A-204 cells.The expression levels of USP4,retinoic acid-inducible gene-Ⅰ(RIG-Ⅰ),melanoma differentiation-associated gene 5(MDA5)and LGP2 protein in A-204 cells were detected by Western blot.Results The expression of cellular USP4 protein in the empty load plasmid group was significantly lower than that in the blank control group(P<0.05),the expression of cellular USP4 protein in the deubiquitinating enzyme group was significantly higher than that in the empty load plasmid group and blank control group(P<0.05).The expression of EV71 mRNA in the empty load plasmid group and deubiquitinating enzyme group was significantly higher than that in the blank control group,and the expression of EV71 mRNA in the deubiquitinating enzyme group was significantly lower than that in the empty load plasmid group(P<0.05);the expression levels of cellular RIG-Ⅰ,MDA5 and LGP-2 in the emptyload plasmid group were significantly lower than those in the blank control group,and the levels of cellular RIG-Ⅰ,MDA5 and LGP-2 in the deubiquitinating enzyme group were significantly higher than those in the empty load plasmid group and blank control group(P<0.05).Conclusion Deubiquitinating enzyme USP4 plays the anti-EV71 infection role possibly by activating the RLR signaling pathway.
作者
梁赣锋
沈扬
俞逊婕
吴海燕
LIANG Ganfeng;SHEN Yang;YU Sunjie;WU Haiyan(Department of Infection, Taizhou Municipal First People′s Hospital,Taizhou,Zhejiang 318020,China)
出处
《重庆医学》
CAS
2021年第7期1100-1103,共4页
Chongqing medicine
