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Mutations in the principal neutralization determinant of human immunodeficiency virus type 1 affect syncytium
R J Grimaila1, B A Fuller, P D Rennert
1Repligen Corporation, Cambridge, Massachusetts 02139.
Abstract:
The principal neutralization determinant (PND) of human immunodeficiency virus type 1 envelope glycoprotein gp120 contains a conserved GPG sequence. The effects of a 29-amino-acid deletion of most of the PND, a 3-amino-acid deletion in the GPG sequence, and 16 single-amino-acid substitutions in the GPG sequence were determined in a transient expression assay. All mutant envelope glycoproteins were expressed at levels comparable to that of the wild-type envelope, and mutations in the GPG sequence did not affect processing to gp120 or, except for the 29-amino-acid deletion, binding to CD4. Of all of the mutants, only the GHG and GFG mutants induced formation of syncytia similar in size and number to those induced by the wild-type envelope. When the envelope expression level was increased 10-fold or more, several additional mutants (APG, GAG, GSG, GQG, GVG, and GPF) also induced syncytium formation. Transfection with infectious proviral molecular clones containing the GHG, GFG, APG, GAG, GSG, or GPF mutations induced production of viral particles; however, only the GPG, GHG, and GFG viruses produced active infections in CD4-bearing cells. Furthermore, whereas the wild-type virus was efficiently neutralized by PND polyclonal and monoclonal antibodies, the GHG- and GFG-containing viruses were not. These results show that mutations in the GPG sequence found within the PND do not affect envelope expression and do not significantly affect CD4 binding or production of viral particles but that they do affect the ability of the envelope to induce syncytia and those of the viral particles to infect CD4 cells and be neutralized by PND antibodies.
Insights
Mutations in the GPG sequence of the human immunodeficiency virus type 1 envelope glycoprotein gp120 affect viral infectivity and neutralization. These changes impact syncytia formation and antibody recognition, but not CD4 binding or viral particle production.
Area of Science:
- Virology
- Immunology
- Molecular Biology
Background:
- The principal neutralization determinant (PND) of human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein gp120 is crucial for viral infectivity and antibody recognition.
- A conserved GPG sequence within the PND is a key target for neutralizing antibodies.
Purpose of the Study:
- To investigate the functional impact of mutations within the GPG sequence of the HIV-1 PND on envelope glycoprotein expression, CD4 binding, syncytia formation, viral infectivity, and neutralization.
- To determine the role of the GPG sequence in HIV-1 pathogenesis and antibody evasion.
Main Methods:
- Transient expression assays were used to assess the effects of deletions and single-amino-acid substitutions in the PND and GPG sequence on envelope glycoprotein function.
- Mutant envelope glycoproteins were analyzed for expression levels, gp120 processing, and CD4 binding.
- Syncytia formation assays were performed to evaluate the cell-to-cell fusion capacity of mutant viruses.
- Infectious molecular clones were generated to assess viral particle production, infectivity in CD4-bearing cells, and neutralization by PND antibodies.
Main Results:
- Mutations in the GPG sequence did not significantly affect envelope expression, gp120 processing, or CD4 binding (except for a large deletion).
- Specific mutations (GHG, GFG) induced syncytia formation comparable to wild-type, and others (APG, GAG, GSG, GQG, GVG, GPF) induced syncytia at higher envelope expression levels.
- While several mutants produced viral particles, only wild-type, GHG, and GFG viruses were infectious in CD4+ cells.
- GHG and GFG viruses exhibited reduced neutralization by PND antibodies compared to wild-type virus.
Conclusions:
- Mutations within the GPG sequence of the HIV-1 PND can alter viral infectivity and antibody neutralization without significantly impacting envelope expression, CD4 binding, or viral particle production.
- The GPG sequence plays a critical role in mediating viral entry, syncytia formation, and susceptibility to antibody-mediated neutralization.
- These findings highlight the potential for HIV-1 to evade immune responses through mutations in key neutralization epitopes.