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Optimal Expression of the Envelope Glycoprotein of Orthobornaviruses Determines the Production of Mature Virus Particles.

ABSTRACT: 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 genus Orthobornavirus, including that of BoDV-1, in the production of infectious particles. By using G-pseudotype assay in which the lack of G in G-deficient REVec (ΔG-REVec) was compensated for 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 vgRNA 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.IMPORTANCE Most viruses causing persistent infection produce few infectious particles from the infected cells. Borna disease virus 1, a member of the genus Orthobornavirus, 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.

SUBMITTER: Sakai M

PROVIDER: S-EPMC8092845 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:The mature envelope glycoprotein (Env) spike on the surfaces of human immunodeficiency virus type 1 (HIV-1)-infected cells and virions is derived from proteolytic cleavage of a trimeric gp160 glycoprotein precursor. In these studies, we compared the conformations of cleaved and uncleaved membrane Envs with truncated cytoplasmic tails to those of stabilized soluble gp140 SOSIP.664 Env trimers. Deletion of the gp41 cytoplasmic tail did not significantly affect the sensitivity of viruses with the HIV-1AD8 Env to inhibition by antibodies or a CD4-mimetic compound. After glutaraldehyde fixation and purification from membranes, a cleaved Env exhibited a hydrodynamic radius of ?10 nm and an antibody-binding profile largely consistent with that expected based on virus neutralization sensitivity. The purified cleaved Env trimers exhibited a hollow architecture with a central void near the trimer axis. Uncleaved Env, cross-linked and purified in parallel, exhibited a hydrodynamic radius similar to that of the cleaved Env. However, the uncleaved Env was recognized by poorly neutralizing antibodies and appeared by negative-stain electron microscopy to sample multiple conformations. Compared with membrane Envs, stabilized soluble gp140 SOSIP.664 Env trimers appear to be more compact, as reflected in their smaller hydrodynamic radii and negative-stain electron microscopy structures. The antigenic features of the soluble gp140 SOSIP.664 Env trimers differed from those of the cleaved membrane Env, particularly in gp120 V3 and some CD4-binding-site epitopes. Thus, proteolytic maturation allows the membrane-anchored Env to achieve a conformation that retains functional metastability but masks epitopes for poorly neutralizing antibodies.IMPORTANCE The entry of human immunodeficiency virus type 1 (HIV-1) into host cells is mediated by the envelope glycoprotein (Env) spike on the surface of the virus. Host antibodies elicited during natural HIV-1 infection or by vaccination can potentially recognize the Env spike and block HIV-1 infection. However, the changing shape of the HIV-1 Env spike protects the virus from antibody binding. Understanding the shapes of natural and man-made preparations of HIV-1 Envs will assist the development of effective vaccines against the virus. Here, we evaluate the effects of several Env modifications commonly used to produce Env preparations for vaccine studies and the determination of structure. We found that the cleavage of the HIV-1 Env precursor helps Env to assume its natural shape, which resists the binding of many commonly elicited antibodies. Stabilized soluble Envs exhibit more compact shapes but expose some Env elements differently than the natural Env.

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Optimal Expression of the Envelope Glycoprotein of Orthobornaviruses Determines the Production of Mature Virus Particles.

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