异戊二烯半胱氨酸氧化酶1抗水疱性口炎病毒作用及机制的初步研究  

Preliminary study on antiviral effect and mechanism of prenylcysteine oxidase 1 against vesicular stomatitis virus

作　　者：李帅臣 孟祥博 许诺 李红坤 周孙欣 王恒心 张彤[1] LI Shuai-chen;MENG Xiang-bo;XU Nuo;LI Hong-kun;ZHOU Sun-xin;WANG Heng-xin;ZHANG Tong(Department of Stomatology,First Medical Center,PLA General Hospital,Beijing 100853,China;PLA Medical School,Beijing 100853,China)

机构地区：[1]解放军总医院第一医学中心口腔科,北京100853 [2]解放军医学院,北京100853

出　　处：《临床口腔医学杂志》2025年第1期3-8,共6页Journal of Clinical Stomatology

基　　金：北京市自然科学基金面上项目(编号:7232154);军队后勤科研重点项目(编号:BKJWS221C002)。

摘　　要：目的:探讨异戊二烯半胱氨酸氧化酶1(prenylcysteine oxidase 1,PCYOX1)对水疱性口炎病毒(vesicular stomatitis virus,VSV)感染复制及固有免疫I型干扰素应答的影响。方法:在人皮肤成纤维细胞(BJ-5ta)中转染小干扰RNA(small interfering RNA,siRNA)或过表达质粒以敲低或过表达PCYOX1基因。使用VSV-GFP感染细胞,通过实时荧光定量PCR(RT-qPCR)检测VSV mRNA的表达水平,免疫印记法(Western blotting)检测VSV-G、GFP蛋白的表达。在PCYOX1基因敲低细胞中转染poly(I∶C),通过RT-qPCR检测干扰素β(interferonβ,IFN-β)、C-X-C基序趋化因子配体10(C-X-C motif chemokine ligand 10,CXCL10)和干扰素刺激基因56(interferon-stimulated gene 56,ISG56)mRNA水平的变化,Western blotting检测干扰素调节因子3(interferon regulatory factor 3,IRF3)和TANK结合激酶1(TANK-binding kinase 1,TBK1)磷酸化水平的变化。结果:成功在BJ-5ta细胞中实现PCYOX1基因的高效敲低及过表达。使用VSV-GFP感染细胞,与对照组相比,PCYOX1敲低组的VSV拷贝数增加至2.50~3.69倍(P<0.001),病毒蛋白表达水平也显著升高,但PCYOX1过表达组病毒mRNA及蛋白表达水平未见明显差异。在PCYOX1基因敲低细胞中转染poly(I∶C)后,固有免疫I型干扰素应答中关键转录因子IRF3的磷酸化水平明显下降,IFN-β及干扰素刺激基因CXCL10和ISG56的mRNA水平显著降低。结论:PCYOX1缺失会增强VSV的感染与复制,抑制I型干扰素介导的固有免疫激活,其可能是机体抵抗RNA病毒的关键调控分子。Objective:To explore the effect of prenylcysteine oxidase 1(PCYOX1)on vesicular stomatitis virus(VSV)replication and innate type I interferon response.Methods:The small interfering RNA(siRNA)or overexpression plasmid was transfected to knock down or overexpress PCYOX1 gene in human skin fibroblast BJ-5ta cells.After infecting cells with VSV-GFP,the expression of VSV mRNA was detected by real-time quantitative PCR(RT-qPCR),and the expression of VSV-G and GFP protein was detected by Western blotting.After transfecting poly(I∶C)into PCYOX1 knockdown cells,the changes in mRNA levels of interferonβ(IFN-β),C-X-C motif chemokine ligand 10(CXCL10),and interferon-stimulated gene 56(ISG56)were detect by RT-qPCR,Western blotting was performed to detect the changes in phosphorylation levels of interferon regulatory factor 3(IRF3)and TANK-binding kinase 1(TBK1).Results:Efficient knockdown and overexpression of PCYOX1 in BJ-5ta cells were successfully achieved.Compared with the control group,the copy number of VSV in PCYOX1 knockdown cells increased by 2.50~3.69 times(P<0.001)when infected with VSV-GFP,and the expression level of viral protein also significantly increased.However,there were no significant differences in the expression levels of viral mRNA and protein between the PCYOX1 overexpression group and control group.After transfecting poly(I∶C)into PCYOX1 knockdown cells,the phosphorylation level of IRF3,which is key transcription factor in innate type I interferon response,was significantly reduced,and the mRNA levels of IFN-βand interferon-stimulated genes CXCL10 and ISG56 were also significantly reduced.Conclusion:PCYOX1 gene deletion enhances the infection and replication of VSV,and inhibits the activation of innate immunity mediated by type I interferon.It may serve as a key regulatory molecule in the resistance to RNA viruses.

关 键 词：异戊二烯半胱氨酸氧化酶1 异戊二烯化 水疱性口炎病毒 抗病毒固有免疫 

分 类 号：Q813[生物学—生物工程] Q782