The extent of virus transmission among individuals and species is generally determined by the presence of specific membrane-embedded virus receptors required for virus entry. Interaction of the viral envelope glycoprotein (Env) with a specific cellular receptor is the first and crucial step in determining host specificity. Using a well-established retroviral model-avian Rous sarcoma virus (RSV)-we analyzed changes in an RSV variant that had repeatedly been able to infect rodents. By envelope gene (env) sequencing, we identified eight mutations that do not match the already described mutations influencing the host range. Two of these mutations-one at the beginning (D32G) of the surface Env subunit (SU) and the other at the end of the fusion peptide region (L378S)-were found to be of critical importance, ensuring transmission to rodent, human, and chicken cells lacking the appropriate receptor. Furthermore, we carried out assays to examine the virus entry mechanism and concluded that these two mutations cause conformational changes in the Env variant and that these changes lead to an activated, or primed, state of Env (normally induced after Env interaction with the receptor). In summary, our results indicate that retroviral host range extension is caused by spontaneous Env activation, which circumvents the need for original cell receptor. This activation is, in turn, caused by mutations in various env regions.

• Klíčová slova
• Rous sarcoma virus, envelope glycoprotein, receptor-independent entry, retrovirus, virus entry,
• MeSH
• genetické vektory * genetika   metabolismus   MeSH
• genové produkty env * genetika   metabolismus   MeSH
• krysa rodu Rattus MeSH
• kur domácí MeSH
• lidé MeSH
• missense mutace * MeSH
• nádorové buněčné linie MeSH
• substituce aminokyselin MeSH
• transdukce genetická * MeSH
• virus Rousova sarkomu * genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• genové produkty env * MeSH

UNLABELLED: Transformation of rodent cells with avian Rous sarcoma virus (RSV) opened new ways to studying virus integration and expression in nonpermissive cells. We were interested in (i) the molecular changes accompanying fusion of RSV-transformed mammalian cells with avian cells leading to virus rescue and (ii) enhancement of this process by retroviral gene products. The RSV-transformed hamster RSCh cell line was characterized as producing only a marginal amount of env mRNA, no envelope glycoprotein, and a small amount of unprocessed Gag protein. Egress of viral unspliced genomic RNA from the nucleus was hampered, and its stability decreased. Cell fusion of the chicken DF-1 cell line with RSCh cells led to production of env mRNA, envelope glycoprotein, and processed Gag and virus-like particle formation. Proteosynthesis inhibition in DF-1 cells suppressed steps leading to virus rescue. Furthermore, new aberrantly spliced env mRNA species were found in the RSCh cells. Finally, we demonstrated that virus rescue efficiency can be significantly increased by complementation with the env gene and the highly expressed gag gene and can be increased the most by a helper virus infection. In summary, Env and Gag synthesis is increased after RSV-transformed hamster cell fusion with chicken fibroblasts, and both proteins provided in trans enhance RSV rescue. We conclude that the chicken fibroblast yields some factor(s) needed for RSV replication, particularly Env and Gag synthesis, in nonpermissive rodent cells. IMPORTANCE: One of the important issues in retrovirus heterotransmission is related to cellular factors that prevent virus replication. Rous sarcoma virus (RSV), a member of the avian sarcoma and leukosis family of retroviruses, is able to infect and transform mammalian cells; however, such transformed cells do not produce infectious virus particles. Using the well-defined model of RSV-transformed rodent cells, we established that the lack of virus replication is due to the absence of chicken factor(s), which can be supplemented by cell fusion. Cell fusion with permissive chicken cells led to an increase in RNA splicing and nuclear export of specific viral mRNAs, as well as synthesis of respective viral proteins and production of virus-like particles. RSV rescue by cell fusion can be potentiated by in trans expression of viral genes in chicken cells. We conclude that rodent cells lack some chicken factor(s) required for proper viral RNA processing and viral protein synthesis.

• MeSH
• fúze buněk MeSH
• genové produkty env genetika   metabolismus   MeSH
• genové produkty gag genetika   metabolismus   MeSH
• křečci praví MeSH
• kur domácí MeSH
• nemoci drůbeže virologie   MeSH
• ptačí sarkom virologie   MeSH
• testy genetické komplementace MeSH
• transformované buněčné linie MeSH
• virová transformace buněk MeSH
• virus Rousova sarkomu genetika   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• křečci praví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• genové produkty env MeSH
• genové produkty gag MeSH