gag, vif, and nef genes contribute to the homologous vital interference induced by a nonproducer human immunodeficiency virus type 1 (HIV-1) variant: Identification of novel HIV-1-inhibiting viral protein mutants

Paola D'Aloja, Eleonora Olivetta, Roberta Bona, Filomena Nappi, Daniela Pedacchia, Katherina Pugliese, Giuliana Ferrari, Paola Verani, Maurizio Federico

Research output: Contribution to journalArticle

Abstract

We previously demonstrated that expression of the nonproducer F12- human immunodeficiency virus type 1 (HIV-1) variant induces a block in the replication of superinfecting HIV that does not depend on the down- regulation of CD4 HIV receptors. In order to individuate the gene(s) involved in F12-HIV-induced interference, vectors expressing each of the nine F12-HIV proteins were transfected in HIV-susceptible HeLa CD4 cells. Pools of cell clones stably producing each viral protein were infected with HIV-1, and virus release was measured in terms of reverse transcriptase activity in supernatants. We hereby demonstrate that HeLa CD4 cells expressing the F12-HIV gag, vif, or nef gene were resistant, to different degrees, to infection with T-cell-line-adapted HIV-1 strains. Conversely, expression of either the tat, rev, or vpu F12-HIV gene increased the rate of HIV release, and no apparent effects on HIV replication were observed in cells expressing either the F12-HIV vpr, pol, or env gene. No variation of CD4 exposure was detected in any of the uninfected HeLa CD4 pools. These data indicate that F12-HIV homologous viral interference is the consequence of the synergistic anti-HIV effects of Gag, Vif, and Nef proteins. Retrovirus vectors expressing F12-HIV vif or nef allowed us to further establish that the expression of each mutated protein (i) inhibits the replication of clinical HIV-1 isolates as well, (ii) impairs the infectivity of the virus released by cells chronically infected with HIV-1, and (iii) limitedly to F12-HIV Vif protein, induces HIV resistance in both vif- permissive and vif-nonpermissive cells. The levels of action of F12-HIV vif and nef anti-HIV effects were also determined. We observed that HIV virions emerging from the first viral cycle on F12-HIV vif-expressing cells, although released in unaltered amounts, had a strongly reduced ability to initiate the retrotranscription process when they reinfected parental HeLa CD4 cells. Differently, we observed that expression of F12-HIV Nef protein affects the HIV life cycle at the level of viral assembling and/or release. For the first time, an inhibitory effect on the HIV life cycle in both acutely and chronically infected cells induced by mutated Vif and Nef HIV-1 proteins is described. These genes could thus be proposed as new useful reagents for anti-HIV gene therapy.

Original languageEnglish
Pages (from-to)4308-4319
Number of pages12
JournalJournal of Virology
Volume72
Issue number5
Publication statusPublished - May 1998

Fingerprint

nef Genes
vif Genes
gag Genes
viral proteins
Viral Proteins
Human immunodeficiency virus 1
crossover interference
HIV-1
HIV
mutants
genes
cells
proteins
life cycle (organisms)
Retroviridae
vif Gene Products
HeLa Cells
Human Immunodeficiency Virus Proteins
viruses
gene therapy

ASJC Scopus subject areas

  • Immunology

Cite this

gag, vif, and nef genes contribute to the homologous vital interference induced by a nonproducer human immunodeficiency virus type 1 (HIV-1) variant : Identification of novel HIV-1-inhibiting viral protein mutants. / D'Aloja, Paola; Olivetta, Eleonora; Bona, Roberta; Nappi, Filomena; Pedacchia, Daniela; Pugliese, Katherina; Ferrari, Giuliana; Verani, Paola; Federico, Maurizio.

In: Journal of Virology, Vol. 72, No. 5, 05.1998, p. 4308-4319.

Research output: Contribution to journalArticle

D'Aloja, Paola ; Olivetta, Eleonora ; Bona, Roberta ; Nappi, Filomena ; Pedacchia, Daniela ; Pugliese, Katherina ; Ferrari, Giuliana ; Verani, Paola ; Federico, Maurizio. / gag, vif, and nef genes contribute to the homologous vital interference induced by a nonproducer human immunodeficiency virus type 1 (HIV-1) variant : Identification of novel HIV-1-inhibiting viral protein mutants. In: Journal of Virology. 1998 ; Vol. 72, No. 5. pp. 4308-4319.
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AU - Olivetta, Eleonora

AU - Bona, Roberta

AU - Nappi, Filomena

AU - Pedacchia, Daniela

AU - Pugliese, Katherina

AU - Ferrari, Giuliana

AU - Verani, Paola

AU - Federico, Maurizio

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N2 - We previously demonstrated that expression of the nonproducer F12- human immunodeficiency virus type 1 (HIV-1) variant induces a block in the replication of superinfecting HIV that does not depend on the down- regulation of CD4 HIV receptors. In order to individuate the gene(s) involved in F12-HIV-induced interference, vectors expressing each of the nine F12-HIV proteins were transfected in HIV-susceptible HeLa CD4 cells. Pools of cell clones stably producing each viral protein were infected with HIV-1, and virus release was measured in terms of reverse transcriptase activity in supernatants. We hereby demonstrate that HeLa CD4 cells expressing the F12-HIV gag, vif, or nef gene were resistant, to different degrees, to infection with T-cell-line-adapted HIV-1 strains. Conversely, expression of either the tat, rev, or vpu F12-HIV gene increased the rate of HIV release, and no apparent effects on HIV replication were observed in cells expressing either the F12-HIV vpr, pol, or env gene. No variation of CD4 exposure was detected in any of the uninfected HeLa CD4 pools. These data indicate that F12-HIV homologous viral interference is the consequence of the synergistic anti-HIV effects of Gag, Vif, and Nef proteins. Retrovirus vectors expressing F12-HIV vif or nef allowed us to further establish that the expression of each mutated protein (i) inhibits the replication of clinical HIV-1 isolates as well, (ii) impairs the infectivity of the virus released by cells chronically infected with HIV-1, and (iii) limitedly to F12-HIV Vif protein, induces HIV resistance in both vif- permissive and vif-nonpermissive cells. The levels of action of F12-HIV vif and nef anti-HIV effects were also determined. We observed that HIV virions emerging from the first viral cycle on F12-HIV vif-expressing cells, although released in unaltered amounts, had a strongly reduced ability to initiate the retrotranscription process when they reinfected parental HeLa CD4 cells. Differently, we observed that expression of F12-HIV Nef protein affects the HIV life cycle at the level of viral assembling and/or release. For the first time, an inhibitory effect on the HIV life cycle in both acutely and chronically infected cells induced by mutated Vif and Nef HIV-1 proteins is described. These genes could thus be proposed as new useful reagents for anti-HIV gene therapy.

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