An investigation into specific therapeutic targeting of the HIV-1 translational frameshift element
HIV-1 is a pathogenic retrovirus, which often develops into acquired immune deficiency syndrome (AIDS) and inevitably leads to death. The virus has directly caused an estimated 36 million deaths worldwide, and is the second highest attributed contributor to deaths caused by communicable diseases. This high mortality rate is due to the virus’ ability to evade treatment regimes by having an incredibly high rate of genomic mutation. However a highly conserved mechanism has been discovered that exhibits a novel control of protein expression. The -1 programmed ribosomal frameshifting region (-1PRF) is located between the gag and pol coding regions of the HIV-1 genome. By developing a mechanism that allows shifting between frames, HIV-1 has been able to condense its genome and tightly control expression of the Gag-Pol protein products. The PRF element is made up of three sub-elements, known as the slippery sequence, intercodon and secondary structural element (SSE). Although previous studies have hypothesised the importance of each sub-element, the interaction and causative effect was unknown. Substitution of the SSEs of HIV-1 and Paternally Expressed Gene 10 (PEG10) showed that the much larger PEG10 pseudoknot increased the efficiency of frameshifting in HIV-1 by 60%, whereas the contrasting substitution dropped PEG10 efficiency down 95%. This indicates that PEG10 relies heavily on the SSE to produce its high native frameshift efficiency. Substitution of the intercodon revealed a degree of ribosomal sampling in HIV-1 as a small increase was observed, however no variation in the PEG10 construct implied that no such sampling occurs in this mechanism. Due to the mechanism’s highly conserved nature, it is the ideal candidate for RNA targeting molecules. The Tate laboratory previously discovered three small molecule modulatory compounds, which were shown to affect HIV-1 frameshifting efficiency. However as there are several known retrotransposon-derived elements within mammals, there was a possibility of interaction between the compound and human -1PRF mechanisms. When exposed to a range of concentrations up to 20μM of compound A3, CCR5 exhibited minimal increases, similar to those seen in PEG10 and far below the interaction seen in HIV-1. Binding of the compounds were shown to be in the SSE of HIV-1, which due to the differing nature of -1PRF SSEs may provide insight into the specificity observed. Thus, small molecule compounds developed for the targeting of the HIV-1 -1PRF region are unlikely to negatively affect the efficacy of known human orthologues, due to the differences in secondary structure and the underlying mechanism of action.
Advisor: Tate, Warren
Degree Name: Master of Science
Degree Discipline: Genetics
Publisher: University of Otago
Keywords: HIV-1; PEG10; CCR5; Translational; Ribosome; Frameshift; gag-pol
Research Type: Thesis