Polypyrimidine Tract-Binding Protein Enhances Zika Virus Translation by Binding to the 5'UTR of Internal Ribosomal Entry Site

Objectives To identify the 5'untranslated region of Zika virus(ZIKV 5'UTR)RNA-binding proteins and to investigate the impact of the binding protein on the activity of internal ribosomal entry site(IRES)located in ZIKV 5'UTR and virus production.Methods Interacting proteins in U251 cells were captured using tRSA-tagged ZIKV 5'UTR RNA and tRSA-ZIKV 5'UTR RNA-binding proteins were visualized by SDS-PAGE silver staining,Subsequently,liquid chromatographytandem mass spectrometry(LC-MS/MS),bioinformatics analysis,and Western blot were used to identify the candidate proteins binding to ZIKV 5'UTR.Dicistronic expression assay and plaque forming assay were performed to analyze the effect of the binding protein on ZIKV IRES activity and ZIKV production,respecitvely.Results tRSA RNA pull-down assay,LC-MS/MS,and Western blot analysis showed that polypyrimidine tractbinding protein(PTB)bound to the ZIKV 5'UTR.Furthermore,dual luciferase reporter assay revealed that overexpression of PTB significantly enhanced the IRES activity of ZIKV(t=10.220,P<0.001),while PTB knockdown had the opposite effect(t=4.897,P<0.01).Additionally,virus plaque forming assay demonstrated that up-regulation of PTB expression significantly enhanced viral titer(t=6.400,P<0.01),whereas reducing PTB expression level weakened virus infectivity(t=5.055,P<0.01).Conclusion PTB positively interacts with the ZIKV 5'UTR and enhances IRES activity and virus production.

超声波均质化对仙台病毒ELISA测试中抗原稳定性的影响

目的研究超声波均质化(homogenization)处理对于仙台病毒在ELISA测试中抗原稳定性的影响。方法对BHK-21细胞内扩增培养的仙台病毒通过差速离心,富集后使用超声波做均质化处理,和未处理的病毒分别包被酶标板,使用标准免疫小鼠血清和SPF小鼠血清对包被平板进行测试,比较样品测试孔间测量数据的变异率。结果对免疫血清做梯度稀释,各梯度样品在未经超声处理仙台病毒抗原包被ELISA检测中测量值的变异率在1.97%～6.02%之间;在经超声处理仙台病毒抗原包被ELISA检测中测量值的变异率在0.53%～2.26%之间;SPF血清测量值的变异率均高于免疫血清;所有样品在经超声处理的病毒抗原包被ELISA检测中测量值的变异率均较小。结论超声波处理有效的提高了仙台病毒抗原的均质性,在ELISA测试中提高了抗原的稳定性。

从中国恒河猴感染猴免疫缺陷病毒后的不同进展情况看艾滋病的发病机制

目的中国恒河猴感染猴免疫缺陷病毒（SIV）后，根据其病毒载量水平、疾病进程速度等，可分为普通进展型（NP）、快速进展型（RP）以及长期不进展型（LTNP）／精英控制者（Ec）等3类。对属于不同进展类型的动物的各项参数进行比较，有助于进一步理解AIDS的发病机制。方法使用SIVmae239毒株静脉感染中国恒河猴后，定期采血进行血液学、免疫学、病毒学及病理学检查；并对各参数进行比较。结果16只感染动物中，有1例动物（RM449猴）快速进展并死亡于感染后4．5个月（RP型）；2例（RM450和RM453）的血浆病毒载量被控制至低于检测水平（Ec型）；其余13例属于NP型。与13只NP型的猴相比较，RM449（RP型）病毒载量高，SIV特异IgG低，效应记忆型CD4’T亚群细胞数低，且流式图出现类似“分化阻滞”的现象；外周血B细胞数的降幅大，以组织样B细胞和活化的记忆B细胞为主；淋巴组织耗竭，胸腺消失；具更高的抗淋巴组织的自身抗体的水平。而RM450和RM453猴（EC型）大致与之相反。结论AIDS的形成可能与T、B淋巴细胞亚群的分化和功能不足有关；而胸腺、淋巴组织等的结构破坏，可能是其病理学基础。