| Human immunodeficiency virus type 1 (HIV-1) and Hepatitis C virus (HCV) are responsible for two major viral pandemics and around 40 and 200 million people, respectively, are infected with these viruses worldwide. During the course of my Ph.D. studies, I investigated unconventional translation mechanisms used by these two viruses.; One of these unconventional translation mechanisms is a programmed -1 ribosomal frameshift that is used by HIV-1. In HIV-1, Gag, the precursor of the viral structural proteins, and Pol, the precursor of the viral enzymes, are synthesized from the same RNA, i.e. the full-length viral RNA, but the coding sequence of Pol is in a -1 reading frame relative to the coding sequence of Gag. The majority of ribosomes translate the viral RNA according to conventional rules and synthesize Gag, but a minority of ribosomes that initiated translation at the initiation codon of Gag shift the reading frame at a specific sequence before they encounter the stop codon of Gag and extend the translation to the pol gene, synthesizing Gag-Pol. The expression of Pol is thus regulated by a -1 frameshift and the efficiency of this -1 frameshift event controls the Gag-Pol to Gag ratio, which is critical for particle assembly, replication and viral infectivity.; We also investigated the translation mechanism leading to the synthesis of a HCV viral protein, the F protein, which was recently discovered and could be implicated in the development of the pathogenesis in patients infected with HCV. Contradictory results that describe the mechanism accounting for the synthesis of the F protein have been published. Among them, a major study showed that the F protein was synthesized via a +1 frameshift by a minority of ribosomes that initiated translation at the AUG codon for the viral polyprotein on a stretch of 10A encompassing codons 9 to 11 of the polyprotein. However, these results were obtained with an in vitro translation system and repetitive sequences, like this stretch of 10A, are known to promote unspecific -1 and +1 frameshifts in vitro . With translation assays in mammalian cultured cells, we demonstrated that the F protein is not synthesized by a +1 frameshift, but results from direct initiation of translation in the +1 reading frame relative to the polyprotein of HCV. (Abstract shortened by UMI.)... |
