Tetherin/BST-2 is an antiviral protein that blocks the release of enveloped viral particles by linking them to the membrane of producing cells. At first, BST-2 genes were described only in humans and other mammals. Recent work identified BST-2 orthologs in nonmammalian vertebrates, including birds. Here, we identify the BST-2 sequence in domestic chicken (Gallus gallus) for the first time and demonstrate its activity against avian sarcoma and leukosis virus (ASLV). We generated a BST-2 knockout in chicken cells and showed that BST-2 is a major determinant of an interferon-induced block of ASLV release. Ectopic expression of chicken BST-2 blocks the release of ASLV in chicken cells and of human immunodeficiency virus type 1 (HIV-1) in human cells. Using metabolic labeling and pulse-chase analysis of HIV-1 Gag proteins, we verified that chicken BST-2 blocks the virus at the release stage. Furthermore, we describe BST-2 orthologs in multiple avian species from 12 avian orders. Previously, some of these species were reported to lack BST-2, highlighting the difficulty of identifying sequences of this extremely variable gene. We analyzed BST-2 genes in the avian orders Galliformes and Passeriformes and showed that they evolve under positive selection. This indicates that avian BST-2 is involved in host-virus evolutionary arms races and suggests that BST-2 antagonists exist in some avian viruses. In summary, we show that chicken BST-2 has the potential to act as a restriction factor against ASLV. Characterizing the interaction of avian BST-2 with avian viruses is important in understanding innate antiviral defenses in birds.IMPORTANCE Birds are important hosts of viruses that have the potential to cause zoonotic infections in humans. However, only a few antiviral genes (called viral restriction factors) have been described in birds, mostly because birds lack counterparts of highly studied mammalian restriction factors. Tetherin/BST-2 is a restriction factor, originally described in humans, that blocks the release of newly formed virus particles from infected cells. Recent work identified BST-2 in nonmammalian vertebrate species, including birds. Here, we report the BST-2 sequence in domestic chicken and describe its antiviral activity against a prototypical avian retrovirus, avian sarcoma and leukosis virus (ASLV). We also identify BST-2 genes in multiple avian species and show that they evolve rapidly in birds, which is an important indication of their relevance for antiviral defense. Analysis of avian BST-2 genes will shed light on defense mechanisms against avian viral pathogens.
- MeSH
- antigen stromálních buněk kostní dřeně genetika imunologie MeSH
- buněčné linie MeSH
- fibroblasty imunologie virologie MeSH
- Galliformes genetika imunologie virologie MeSH
- genové produkty gag - virus lidské imunodeficience genetika imunologie MeSH
- HEK293 buňky MeSH
- HIV-1 genetika imunologie MeSH
- interakce hostitele a patogenu genetika imunologie MeSH
- lidé MeSH
- molekulární evoluce * MeSH
- Passeriformes genetika imunologie virologie MeSH
- ptačí proteiny genetika imunologie MeSH
- ptačí sarkom genetika imunologie virologie MeSH
- regulace genové exprese MeSH
- replikace viru MeSH
- sekvence aminokyselin MeSH
- sekvenční homologie aminokyselin MeSH
- sekvenční seřazení MeSH
- selekce (genetika) MeSH
- signální transdukce MeSH
- uvolnění viru z buňky MeSH
- viry ptačího sarkomu genetika imunologie patogenita MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
- Research Support, N.I.H., Intramural MeSH
- MeSH
- dějiny 20. století MeSH
- experimentální nádory MeSH
- infekce onkogenními viry genetika přenos MeSH
- kur domácí MeSH
- leukemie * veterinární virologie MeSH
- nádorová transformace buněk MeSH
- nádory dějiny etiologie klasifikace virologie MeSH
- onkogenní viry klasifikace patogenita MeSH
- onkogeny * MeSH
- proviry * enzymologie genetika MeSH
- pseudogeny MeSH
- ptačí sarkom * etiologie virologie MeSH
- ptáci MeSH
- replikace viru MeSH
- retroelementy MeSH
- Retroviridae účinky léků MeSH
- reverzní transkripce MeSH
- reverzní transkriptasa dějiny MeSH
- savci MeSH
- zvířata MeSH
- Check Tag
- dějiny 20. století MeSH
- zvířata MeSH
In my article I tried to present the results of early experiments suggesting a significant role for cell association in Rous sarcoma virus transformation of non-permissive cells and revealing that infectious virus can be efficiently rescued from such cells by their fusion with permissive chicken fibroblasts.
- MeSH
- krysa rodu rattus MeSH
- kur domácí virologie MeSH
- proviry patogenita fyziologie MeSH
- ptačí sarkom virologie MeSH
- replikace viru MeSH
- virová transformace buněk MeSH
- virus Rousova sarkomu patogenita fyziologie MeSH
- zvířata MeSH
- Check Tag
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy 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
- MeSH
- dějiny 20. století MeSH
- dějiny 21. století MeSH
- geny src * MeSH
- heterografty MeSH
- integrace viru MeSH
- krysa rodu rattus MeSH
- kur domácí virologie MeSH
- nádorové buněčné linie MeSH
- nemoci drůbeže dějiny virologie MeSH
- periodika jako téma dějiny MeSH
- proviry genetika fyziologie MeSH
- ptačí sarkom dějiny virologie MeSH
- transplantace nádorů MeSH
- virová transformace buněk MeSH
- virus Rousova sarkomu genetika izolace a purifikace fyziologie MeSH
- zvířata MeSH
- Check Tag
- dějiny 20. století MeSH
- dějiny 21. století MeSH
- krysa rodu rattus MeSH
- zvířata MeSH
- Publikační typ
- autobiografie MeSH
- biografie MeSH
- historické články MeSH
- úvodníky MeSH
- Geografické názvy
- Československo MeSH
- Maryland MeSH
- O autorovi
- Rous, Francis Payton, 1879-1970 Autorita
The group of closely related avian sarcoma and leukosis viruses (ASLVs) evolved from a common ancestor into multiple subgroups, A to J, with differential host range among galliform species and chicken lines. These subgroups differ in variable parts of their envelope glycoproteins, the major determinants of virus interaction with specific receptor molecules. Three genetic loci, tva, tvb, and tvc, code for single membrane-spanning receptors from diverse protein families that confer susceptibility to the ASLV subgroups. The host range expansion of the ancestral virus might have been driven by gradual evolution of resistance in host cells, and the resistance alleles in all three receptor loci have been identified. Here, we characterized two alleles of the tva receptor gene with similar intronic deletions comprising the deduced branch-point signal within the first intron and leading to inefficient splicing of tva mRNA. As a result, we observed decreased susceptibility to subgroup A ASLV in vitro and in vivo. These alleles were independently found in a close-bred line of domestic chicken and Indian red jungle fowl (Gallus gallus murghi), suggesting that their prevalence might be much wider in outbred chicken breeds. We identified defective splicing to be a mechanism of resistance to ASLV and conclude that such a type of mutation could play an important role in virus-host coevolution.
- MeSH
- Alpharetrovirus genetika fyziologie MeSH
- genetická predispozice k nemoci MeSH
- introny MeSH
- kur domácí genetika metabolismus virologie MeSH
- molekulární sekvence - údaje MeSH
- nemoci drůbeže genetika metabolismus virologie MeSH
- ptačí proteiny genetika metabolismus MeSH
- ptačí sarkom genetika metabolismus virologie MeSH
- sekvence aminokyselin MeSH
- sekvence nukleotidů MeSH
- sekvenční delece MeSH
- sestřih RNA MeSH
- virové receptory genetika metabolismus MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Retroviruses and retrovirus-derived vectors integrate nonrandomly into the genomes of host cells with specific preferences for transcribed genes, gene-rich regions, and CpG islands. However, the genomic features that influence the transcriptional activities of integrated retroviruses or retroviral vectors are poorly understood. We report here the cloning and characterization of avian sarcoma virus integration sites from chicken tumors. Growing progressively, dependent on high and stable expression of the transduced v-src oncogene, these tumors represent clonal expansions of cells bearing transcriptionally active replication-defective proviruses. Therefore, integration sites in our study distinguished genomic loci favorable for the expression of integrated retroviruses and gene transfer vectors. Analysis of integration sites from avian sarcoma virus-induced tumors showed strikingly nonrandom distribution, with proviruses found prevalently within or close to transcription units, particularly in genes broadly expressed in multiple tissues but not in tissue-specifically expressed genes. We infer that proviruses integrated in these genomic areas efficiently avoid transcriptional silencing and remain active for a long time during the growth of tumors. Defining the differences between unselected retroviral integration sites and sites selected for long-terminal-repeat-driven gene expression is relevant for retrovirus-mediated gene transfer and has ramifications for gene therapy.
- MeSH
- chromozomy virologie MeSH
- exprese genu MeSH
- genetická terapie metody MeSH
- genetické vektory MeSH
- integrace viru MeSH
- kur domácí MeSH
- proviry genetika fyziologie MeSH
- ptačí sarkom virologie MeSH
- viry ptačího sarkomu genetika fyziologie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Unmethylated CpG islands are known to keep adjacent promoters transcriptionally active. In the CpG island adjacent to the adenosine phosphoribosyltransferase gene, the protection against transcriptional silencing can be attributed to the short CpG-rich core element containing Sp1 binding sites. We report here the insertion of this CpG island core element, IE, into the long terminal repeat of a retroviral vector derived from Rous sarcoma virus, which normally suffers from progressive transcriptional silencing in mammalian cells. IE insertion into a specific position between enhancer and promoter sequences led to efficient protection of the integrated vector from silencing and gradual CpG methylation in rodent and human cells. Individual cell clones with IE-modified reporter vectors display high levels of reporter expression for a sustained period and without substantial variegation in the cell culture. The presence of Sp1 binding sites is important for the protective effect of IE, but at least some part of the entire antisilencing capacity is maintained in IE with mutated Sp1 sites. We suggest that this strategy of antisilencing protection by the CpG island core element may prove generally useful in retroviral vectors.
- MeSH
- biologické modely MeSH
- CpG ostrůvky MeSH
- financování organizované MeSH
- genetická transkripce MeSH
- koncové repetice MeSH
- lidé MeSH
- mutace MeSH
- průtoková cytometrie MeSH
- ptačí sarkom genetika virologie MeSH
- reportérové geny MeSH
- transkripční faktor Sp1 metabolismus MeSH
- umlčování genů MeSH
- vazebná místa MeSH
- virus ptačí leukózy metabolismus MeSH
- virus Rousova sarkomu metabolismus MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- MeSH
- Alpharetrovirus imunologie MeSH
- autoimunitní nemoci genetika veterinární vrozené MeSH
- hlavní histokompatibilní komplex imunologie účinky léků MeSH
- kur domácí genetika imunologie virologie MeSH
- kuřecí embryo MeSH
- onkogeny genetika imunologie MeSH
- ptačí sarkom genetika imunologie virologie MeSH
- techniky in vitro MeSH
- významné osobnosti dějiny MeSH
- zvířata MeSH
- Check Tag
- kuřecí embryo MeSH
- zvířata MeSH
