The ability of ribosomes to maintain the correct translational reading frame is fundamental to the integrity of protein synthesis and to cell growth and viability. However, there are now a number of examples utilized by viruses in which elongating ribosomes are programmed to shift their translational reading frame one base in the 5' direction. This process is called programmed -1 ribosomal frameshifting. Programmed -1 ribosomal frameshifting is utilized uniquely by eucaryotic viruses, making it a compelling target for developing antiviral agents. The human immunodeficiency virus type 1 (HIV-1) utilizes a programmed -1 ribosomal frameshift to synthesize both the gag and gag-pol proteins from a single transcript. We have been investigating the cis-acting elements and trans-acting factors that determine programmed -1 ribosomal frameshifting efficiencies in the yeast Saccharomyces cerevisiae. Using the double-stranded L-A virus system, we have shown that small changes in the ratio of the gag to gag-pol synthesized by altering frameshifting efficiencies leads to a loss of the killer virus. These findings have led us to develop the concept that antiviral agents can be identified that alter the efficiency of programmed frameshifting without dramatically affecting global protein synthesis. We have successfully demonstrated this principle using the yeast killer virus system as the model, and more recently, with HIV in mammalian cells. Based on these investigations, we propose to characterize programmed -1 frameshifting in HIV-1 with the goal of developing this virus specific mechanism as a target for antiviral intervention.
The aims of the experiments proposed in this grant proposal will be to characterize the sequences that promote efficient frameshifting in HIV-1 and to determine the affects of altering frameshifting efficiency on HIV production. We will also determine if putative compounds that alter programmed -1 frameshifting and promote loss of the killer virus will also reduce or eliminate HIV production. Additional compounds that affect programmed frameshifting will also be identified. Finally, we will investigate at the molecular level how compounds that affect programmed frameshifting function. Our long term goal will be to develop compounds that affect frameshifting to the point that proof of principle has been established and antiviral agents targeting this process will be subsequently developed for clinical use.
