Optimal Expression of the Envelope Glycoprotein of Orthobornaviruses Determines the Production of Mature Virus Particles.
Optimal Expression of the Envelope Glycoprotein of Orthobornaviruses Determines the Production of Mature Virus Particles.
复制标题
正博尔纳病毒包膜糖蛋白的最佳表达决定了成熟病毒颗粒的产生。
DOI：
10.1128/jvi.02221-20
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Makino A.
中科院分区：
文献类型：
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作者：
Sakai M;Fujita Y;Komorizono R;Kanda T;Komatsu Y;Noda T;Tomonaga K;Makino A.
基于RNA病毒的附加型载体（REVec），其骨架是博尔纳病病毒1（BoDV-1），可以在转导的细胞中提供长期基因表达。为了提高REVec的转导效率，我们评估了包括BoDV-1在内的正核糖核酸病毒属病毒包膜糖蛋白（G）在感染性颗粒产生中的作用。通过使用G-假型测定，其中G-缺陷型REVec（ΔG-REVec）中G的缺乏被G的表达补偿，我们发现BoDV-1-G的过量表达不影响颗粒本身的产生，但导致细胞中未切割的和异常的成熟G表达，导致产生具有低转导滴度的REVec颗粒。我们发现，未切割的G在细胞中的表达抑制了成熟G和病毒基因组RNA掺入到颗粒中。G的这一特征在哺乳动物和禽类正核糖核酸病毒中是保守的;然而，金丝雀核糖核酸病毒1型（CnBV-1）-G的切割效力例外地不受其过量表达的影响，这导致产生具有最高滴度的假型ΔG-REVec。CnBV-1和CnBV-2之间的嵌合G蛋白揭示了CnBV-1-G的信号肽通过与细胞内furin的相互作用负责切割效率。我们表明，CnBV-1-G导致具有高转导效率和高滴度重组REVec的假型REVec的发展。我们的研究表明，正核糖核酸病毒G的限制性表达有助于调节感染性颗粒的产生，其机制可以提高REVec的转导效率。重要的是，大多数引起持续感染的病毒从感染的细胞中产生很少的感染性颗粒。博尔纳病病毒1型属于正核糖核酸病毒属，是一种持续感染细胞核的RNA病毒，已被应用于长期基因表达载体。在这项研究中，我们发现，在正核糖核酸病毒中常见的是，过量的G表达不会影响颗粒本身的产生，但会减少具有成熟G和基因组RNA的感染性颗粒的产生。该结果表明，有限的G表达有助于抑制异常病毒颗粒的产生。另一方面，我们发现金丝雀博尔纳病毒1具有特殊的G成熟机制，并产生高滴度病毒。本研究不仅有助于了解正核糖核酸病毒感染性颗粒的产生机制，而且有助于改进正核糖核酸病毒的载体系统。
An RNA virus-based episomal vector (REVec) whose backbone is Borna disease virus 1 (BoDV-1) can provide long-term gene expression in transduced cells. To improve the transduction efficiency of REVec, we evaluated the role of the viral envelope glycoprotein (G) of the genusOrthobornavirus, including that of BoDV-1, in the production of infectious particles. By using a G-pseudotype assay in which the lack of G in G-deficient REVec (ΔG-REVec) was compensated for the expression of G, we found that excess expression of BoDV-1-G does not affect particle production itself but results in uncleaved and aberrant mature G expression in the cells, leading to the production of REVec particles with low transduction titers. We revealed that the expression of uncleaved G in the cells inhibits the incorporation of mature G and viral genomic RNA into the particles. This feature of G was conserved among mammalian and avian orthobornaviruses; however, the cleavage efficacy of canary bornavirus 1 (CnBV-1)-G was exceptionally not impaired by its excess expression, which led to the production of the pseudotype ΔG-REVec with the highest titer. Chimeric G proteins between CnBV-1 and -2 revealed that the signal peptide of CnBV-1-G was responsible for the cleavage efficacy through the interaction with intracellular furin. We showed that CnBV-1-G leads to the development of pseudotyped REVec with high transduction efficiency and a high-titer recombinant REVec. Our study demonstrated that the restricted expression of orthobornavirus G contributes to the regulation of infectious particle production, the mechanism of which can improve the transduction efficiency of REVec.IMPORTANCEMost viruses causing persistent infection produce few infectious particles from the infected cells. Borna disease virus 1, a member of the genusOrthobornavirus, is an RNA virus that persistently infects the nucleus and has been applied to vectors for long-term gene expression. In this study, we showed that, common among orthobornaviruses, excessive G expression does not affect particle production itself but reduces the production of infectious particles with mature G and genomic RNA. This result suggested that limited G expression contributes to suppressing abnormal viral particle production. On the other hand, we found that canary bornavirus 1 has an exceptional G maturation mechanism and produces a high-titer virus. Our study will contribute to not only understanding the mechanism of infectious particle production but also improving the vector system of orthobornaviruses.