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Analysis of the vif gene of feline immunodeficiency virus
1Department of Medical Pathology, University of California, Davis 95616.
Abstract:
We have examined FIV vif function in the context of molecular clone pF34 (GenBank Accession No. M25729). Rabbit antisera directed against the translation product of the vif gene identified a 29-kDa protein in tissue culture cells infected with FIV-pF34; this protein was not present in cultures of uninfected cells. Thus, the vif gene of this virus strain is expressed in infected cells. Three mutations were made in distinct regions of the vif gene of molecular clone FIV-pF34: (i) deletion of 223 bp from the central portion of the gene, (ii) site-directed mutation of a conserved N-terminal basic region, and (iii) site-directed mutation of a conserved C-terminal motif. FIV proviruses containing each of these mutations were assessed for replication following transfection into two feline adherent cell types, CrFK and G355-5. Reverse transcriptase and p24gag antigen assays of supernatants from transfected cultures revealed that all three vif mutants produced very little cell-free virus or viral protein in both cell types. Results of immunocytochemical staining of these cultures indicated that all three mutants expressed low levels of cell-associated FIV p24gag. These findings suggest that each of the three regions mutated in vif is critical for function. Our observations are consistent with studies showing marked attenuation of HIV-1 vif mutants in certain cell types. We conclude that the vif gene is a critical determinant of FIV-pF34 replication and infectivity in CrFK and G355-5 cells.
Insights
The feline immunodeficiency virus (FIV) vif gene is crucial for viral replication and infectivity. Mutations in key vif regions significantly impair FIV production in feline cells, highlighting its essential function.
Area of Science:
- Virology
- Molecular Biology
- Immunodeficiency Viruses
Background:
- The feline immunodeficiency virus (FIV) vif gene's role in viral replication is not fully understood.
- Previous studies on human immunodeficiency virus type 1 (HIV-1) suggest vif is important for infectivity.
- Understanding FIV vif function can provide insights into lentiviral replication mechanisms.
Purpose of the Study:
- To investigate the function of the FIV vif gene using molecular clone pF34.
- To determine the impact of mutations in critical vif regions on FIV replication.
- To assess the necessity of vif for FIV infectivity in feline cell lines.
Main Methods:
- Generated rabbit antisera against the vif gene's translation product to detect the FIV Vif protein.
- Created three distinct mutations within the FIV vif gene: a central deletion, an N-terminal mutation, and a C-terminal mutation.
- Transfected FIV proviruses with wild-type and mutant vif genes into CrFK and G355-5 feline cells.
- Assessed viral replication using reverse transcriptase assays, p24gag antigen detection, and immunocytochemical staining.
Main Results:
- A 29-kDa FIV Vif protein was detected in infected cells, confirming vif gene expression.
- All three vif mutants exhibited significantly reduced production of cell-free virus and viral proteins.
- Mutant viruses showed low levels of cell-associated FIV p24gag, indicating impaired replication and assembly.
- The mutations in distinct vif regions critically impaired viral function in both cell types.
Conclusions:
- The FIV vif gene is essential for efficient viral replication and infectivity in CrFK and G355-5 cells.
- Each of the three mutated regions within vif is critical for its function.
- FIV vif plays a vital role in FIV-pF34 replication, similar to the role of HIV-1 vif.
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