Dual Mechanisms of Translation Initiation of the Full-Length HIV-1 mRNA Contribute to Gag Synthesis

Anne Monette,Fernando Valiente-Echeverría,Éric A Cohen,Marcelo Lopez-Lastra,Matias Rivero,Andrew J Mouland,David Harrich

doi:10.1371/journal.pone.0068108

Abstract

The precursor group-specific antigen (pr55Gag) is central to HIV-1 assembly. Its expression alone is sufficient to assemble into virus-like particles. It also selects the genomic RNA for encapsidation and is involved in several important virus-host interactions for viral assembly and restriction, making its synthesis essential for aspects of viral replication. Here, we show that the initiation of translation of the HIV-1 genomic RNA is mediated through both a cap-dependent and an internal ribosome entry site (IRES)-mediated mechanisms. In support of this notion, pr55Gag synthesis was maintained at 70% when cap-dependent translation initiation was blocked by the expression of eIF4G- and PABP targeting viral proteases in two in vitro systems and in HIV-1-expressing cells directly infected with poliovirus. While our data reveal that IRES-dependent translation of the viral genomic RNA ensures pr55Gag expression, the synthesis of other HIV-1 proteins, including that of pr160Gag/Pol, Vpr and Tat is suppressed early during progressive poliovirus infection. The data presented herein implies that the unspliced HIV-1 genomic RNA utilizes both cap-dependent and IRES-dependent translation initiation to supply pr55Gag for virus assembly and production.

Highlights

Translation initiation of most eukaryotic mRNAs occurs by a scanning mechanism whereby the 40S ribosomal subunit is recruited to the vicinity of the 59-cap-structure, a 7-methylguanylic acid residue located at the 59 terminus of eukaryotic mRNAs, and scans in the 5’ to 3’ direction until an initiation codon is encountered [1,2,3]
Controversy exists regarding the mechanism used by the human immunodeficiency virus type 1 (HIV-1) genomic mRNA to initiate protein synthesis during an infection cycle [15,16,17,18]
The most important feature of this system is that the translation product from the artificially generated upstream open reading frame does not encode a functional firefly luciferase reporter gene (FLuc) reporter protein

Summary

Introduction

Translation initiation of most eukaryotic mRNAs occurs by a scanning mechanism whereby the 40S ribosomal subunit is recruited to the vicinity of the 59-cap-structure, a 7-methylguanylic acid residue located at the 59 terminus of eukaryotic mRNAs, and scans in the 5’ to 3’ direction until an initiation codon is encountered [1,2,3]. We first programmed protein synthesis with in vitro synthesized RNA to show that the complete cleavage of eIF4G, resulting from the in vitro expression of the foot and mouth disease virus (FMDV) L protease, only moderately impacted the translation of a monocistronic mRNA harboring the HIV-1 59UTR.

Results

Conclusion