The level of reverse transcriptase (RT) in human immunodeficiency virus type 1 particles affects susceptibility to nonnucleoside RT inhibitors but not to lamivudine

Abstract

We investigated the relationship between the level of reverse transcriptase (RT) in human immunodeficiency virus type 1 (HIV-1) particles and susceptibility to nonnucleoside reverse transcriptase inhibitors (NNRTIs). HIV-1 virions containing different active levels of RT were generated. Susceptibility to the NNRTIs efavirenz and nevirapine was inversely proportional to the level of enzymatically active RT. However, the sensitivity of HIV-1 to the nucleoside analog 3TC was not affected by the level of RT per particle. These data indicate that the susceptibility of HIV-1 to NNRTIs is influenced by RT activity.

FIG. 1.

The specific infectivity of HIV-1 correlates with the amount of polymerase-active RT per virion. Phenotypically mixed viruses were made by cotransfecting different ratios of plasmids encoding a WT HIV-1 vector and an HIV-1 vector containing the D110E and Y181I mutations in RT. Specific infectivity was measured as IU/p24, and HIV-1 containing 100% WT RT was normalized to 100%.

FIG. 2.

Sensitivity to NNRTIs depends on the RT activity in particles. Phenotypically mixed viruses (100%, 50%, or 25% WT RT) were inhibited in a single round of infection at similar multiplicities of infection by EFV (A) or NVP (B). Error bars represent the standard deviations of three or two independent experiments, respectively. Infection level was normalized to 100% for virus in the absence of drug.

FIG. 3.

Inhibition of HIV-1 by 3TC is not affected by the RT activity in particles. Phenotypically mixed viruses (100%, 50%, or 25% WT RT) were inhibited in a single round of infection assay at similar multiplicities of infection by AZT (A) or 3TC (B). Error bars represent the standard deviations of three or two independent experiments, respectively. Infection level was normalized to 100% for virus in the absence of drug.

FIG. 4.

Extra WT RT delivered to particles via Vpr-RT changes the susceptibility of HIV-1 to EFV but not 3TC. Immunobloting was performed on purified WT HIV-1, HIV with Vpr-RT, or HIV with Vpr-RT DM lysates, and blots were probed with an anti-Gag antibody (A) or anti-RT antibodies (B). HIV containing Vpr-RT or Vpr-RT DM was inhibited in a single round of infection at similar multiplicities of infection by EFV (C) or 3TC (D). Error bars represent the standard deviations of three independent experiments. Infection level was normalized to 100% for virus in the absence of drug.