戊型肝炎病毒是一种单链病毒,可导致免疫功能低下患者慢性感染的正义性RNA病毒。已经在此类患者中描述了病毒宿主重组变体(VHRV)。这些变体将部分人类基因整合到富含聚脯氨酸的区域,可以引入新的翻译后修饰(PTM)。比如泛素化。本研究的目的是表征不同VHRV的复制能力,即,RNF19A,ZNF787,KIF1B,EEF1A1、RNA18、RPS17和RPL6。我们用了一个编码Kernow菌株的质粒,其中编码S17插入的片段被删除(Kernowp6delS17)或被编码不同插入的片段替换。通过RT-qPCR测定上清液和HepG2/C3A细胞裂解物中的HEVRNA浓度。对衣壳蛋白ORF2进行免疫染色。还评估了利巴韦林的作用。对于携带RNF19A的变体,上清液和细胞裂解物中的HEVRNA浓度较高,ZNF787,KIF1B,RPS17和EEF1A1插入比Kernowp6delS17,而不是RNA18或RPL6片段。RNF19A的ORF2病灶数量较高,ZNF787,KIF1B,和RPS17比Kernowp6delS17。具有复制优势的VHRV对利巴韦林的抗病毒作用不太敏感。在具有复制优势的VHRV和没有复制优势的VHRV之间没有发现PTM的差异。总之,我们的研究表明,插入并没有系统地赋予体外复制优势。需要进一步的研究来确定复制能力差异的潜在机制。
目的:戊型肝炎病毒(HEV)是病毒性肝炎的主要病因。HEV可导致免疫功能低下患者的慢性感染。利巴韦林治疗目前用于治疗此类慢性感染。最近,在免疫功能低下患者中对7种病毒宿主重组病毒进行了表征.这些病毒已将人基因片段的一部分整合到其基因组中。我们研究了这些插入对复制能力的影响。我们发现这些插入的片段可以增强七个重组变体中的五个的病毒复制。我们还表明,具有复制优势的重组变体在体外对利巴韦林的敏感性较低。最后,我们发现,导致这种复制优势的机制似乎并不依赖于可能改变病毒蛋白功能的人类基因片段引入的翻译后修饰。涉及改善此类重组病毒复制的机制仍有待探索。
Hepatitis E virus is a single-strand, positive-sense RNA virus that can lead to chronic infection in immunocompromised patients. Virus-host recombinant variants (VHRVs) have been described in such patients. These variants integrate part of human genes into the polyproline-rich region that could introduce new post-translational modifications (PTMs), such as ubiquitination. The aim of this study was to characterize the replication capacity of different VHRVs, namely, RNF19A, ZNF787, KIF1B, EEF1A1, RNA18, RPS17, and RPL6. We used a plasmid encoding the Kernow strain, in which the fragment encoding the S17 insertion was deleted (Kernow p6 delS17) or replaced by fragments encoding the different insertions. The HEV RNA concentrations in the supernatants and the HepG2/C3A cell lysates were determined via RT-qPCR. The capsid protein ORF2 was immunostained. The effect of ribavirin was also assessed. The HEV RNA concentrations in the supernatants and the cell lysates were higher for the variants harboring the RNF19A, ZNF787, KIF1B, RPS17, and EEF1A1 insertions than for the Kernow p6 del S17, while it was not with RNA18 or RPL6 fragments. The number of ORF2 foci was higher for RNF19A, ZNF787, KIF1B, and RPS17 than for Kernow p6 del S17. VHRVs with replicative advantages were less sensitive to the antiviral effect of ribavirin. No difference in PTMs was found between VHRVs with a replicative advantage and those without. In conclusion, our study showed that insertions did not systematically confer a replicative advantage in vitro. Further studies are needed to determine the mechanisms underlying the differences in replicative capacity.
OBJECTIVE: Hepatitis E virus (HEV) is a major cause of viral hepatitis. HEV can lead to chronic infection in immunocompromised patients. Ribavirin treatment is currently used to treat such chronic infections. Recently, seven virus-host recombinant viruses were characterized in immunocompromised patients. These viruses have incorporated a portion of a human gene fragment into their genome. We studied the consequences of these insertions on the replication capacity. We found that these inserted fragments could enhance virus replication for five of the seven recombinant variants. We also showed that the recombinant variants with replicative advantages were less sensitive to ribavirin in vitro. Finally, we found that the mechanisms leading to such a replicative advantage do not seem to rely on the post-translational modifications introduced by the human gene fragment that could have modified the function of the viral protein. The mechanisms involved in improving the replication of such recombinant viruses remain to be explored.