Inhibition of CD4 Translation Mediated by Human Immunodeficiency Virus Type 1 Envelope Protein in a Cell-free System

Stephan Bour,Romas Geleziunas,Mark A Wainberg

doi:10.1074/jbc.272.46.29005

Stephan Bour, Romas Geleziunas + Show 1 more

Open Access

https://doi.org/10.1074/jbc.272.46.29005

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Abstract

The human immunodeficiency virus type 1 (HIV-1) employs a number of complex strategies to interfere with the synthesis, stability, and subcellular localization of its specific cellular receptor CD4. To define better the mechanisms of inhibition of CD4 expression, we used a rabbit reticulocyte lysate in vitro system, in which cDNAs derived from HIV-1-infected cells were used to generate mRNA for the Tat, Vpu, and gp160 envelope proteins that were translated together with CD4-encoding mRNA. In the presence of microsomal membranes, we observed that cotranslation of Env mRNA resulted in a dose-dependent inhibition of CD4 translation. This effect was enhanced further when an mRNA-encoding Vpu in addition to Env mRNA was utilized. However, the activity of Vpu was mostly post-translational, since translation of Vpu alone, but not Env, was able to destabilize CD4 molecules presynthesized into microsomes. The Env-mediated inhibitory effect was specifically targeted at CD4 and did not affect the synthesis or stability of the CD8 molecule. Interestingly, mutated CD4 species, with a 20-fold lower affinity for HIV-1 Env than wild-type, were less sensitive to cotranslational inhibition. Our report identifies the envelope as the HIV-1 protein responsible for down-regulation of CD4 translation. We further propose a mechanism whereby direct interactions between gp160 and nascent CD4 molecules can cause interference with and premature termination of CD4 protein elongation.

Highlights

Infection of CD4ϩ cells by the human immunodeficiency virus type 1 (HIV-1)1 generally leads to cell surface depletion of the CD4 receptor
Vpu induces CD4 degradation in the endoplasmic reticulum and preferably targets CD4 molecules trapped in complexes with the gp160 envelope precursor [14, 19], whereas Nef has been shown to induce CD4 internalization from the cell surface followed by lysosomal degradation
In Vitro Expression and Identification of env, tat, and vpu—To study inhibition of CD4 translation in the presence of the HIV-1 envelope protein, mRNAs similar to those expressed in HIV-infected cells were used [37]

Summary

Introduction

Infection of CD4ϩ cells by the human immunodeficiency virus type 1 (HIV-1) generally leads to cell surface depletion of the CD4 receptor (for review, see Ref. 1) This down-regulation of CD4 is largely dependent on the formation of intracellular complexes between CD4 and the gp160 viral envelope precursor protein [2,3,4,5]. The effect of HIV-1 infection on CD4 translation could not be clearly addressed because of the use of in vivo models in which both transcriptional and post-translational levels of CD4 down-regulation contributed to the overall decrease in CD4 abundance Both the mechanisms and viral proteins involved in a presumed inhibition of CD4 translation remained unknown.

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