Frequent toggling between alternative amino acids is driven by selection in HIV-1

Abstract

Host immune responses against infectious pathogens exert strong selective pressures favouring the emergence of escape mutations that prevent immune recognition. Escape mutations within or flanking functionally conserved epitopes can occur at a significant cost to the pathogen in terms of its ability to replicate effectively. Such mutations come under selective pressure to revert to the wild type in hosts that do not mount an immune response against the epitope. Amino acid positions exhibiting this pattern of escape and reversion are of interest because they tend to coincide with immune responses that control pathogen replication effectively. We have used a probabilistic model of protein coding sequence evolution to detect sites in HIV-1 exhibiting a pattern of rapid escape and reversion. Our model is designed to detect sites that toggle between a wild type amino acid, which is susceptible to a specific immune response, and amino acids with lower replicative fitness that evade immune recognition. Through simulation, we show that this model has significantly greater power to detect selection involving immune escape and reversion than standard models of diversifying selection, which are sensitive to an overall increased rate of non-synonymous substitution. Applied to alignments of HIV-1 protein coding sequences, the model of immune escape and reversion detects a significantly greater number of adaptively evolving sites in env and nef. In all genes tested, the model provides a significantly better description of adaptively evolving sites than standard models of diversifying selection. Several of the sites detected are corroborated by association between Human Leukocyte Antigen (HLA) and viral sequence polymorphisms. Overall, there is evidence for a large number of sites in HIV-1 evolving under strong selective pressure, but exhibiting low sequence diversity. A phylogenetic model designed to detect rapid toggling between wild type and escape amino acids identifies a larger number of adaptively evolving sites in HIV-1, and can in some cases correctly identify the amino acid that is susceptible to the immune response.

Figure 1. Codon model of amino acid toggling.

The immune escape and reversion model has three classes of codons: codons encoding the wild type amino acid (class x); codons separated from the wild type by a single nucleotide substitution (y); and codons separated from the wild type by more than one substitution (z). In the example shown, phenylalanine (F) is the wild type amino acid. Rates of substitution between F and each of the six amino acids within one nucleotide substitution of F or from these amino acids back to F are affected by the parameter ρ, the amino acid toggling rate. All other non-synonymous substitutions have a multiplier ω instead of ρ. Rates of all substitutions depend on the frequency parameters , where c represents the codon class. The parameters take account of the codon bias estimated across the entire alignment and free parameters t_c which describe the proportion of time spent by the site in each of the three codon classes.

Figure 2. Simulation along HIV-1 nef phylogeny.

Data was simulated under purifying selection, neutrality, diversifying selection, or toggling to evaluate power and false positives rates. (A) Amino acid sequence logos of ten randomly drawn codon sites for each category of simulated site. (B) Simulated toggling site mapped to HIV-1 phylogeny showing the occurrence of escape (red arrows) and reversion (green arrows) mutations. (C) Simulated diversifying selection site mapped to HIV-1 phylogeny.

Figure 3. Evaluation of power and false positives.

ROC curves indicating power and false positive rates for the detection of diversifying selection (left panel), diversifying selection and toggling (centre panel), and toggling (right panel) for each of the five parameter sets (Table 1) simulated.

Figure 4. Evidence of toggling and diversifying selection in real data.

Tree branches are coloured according to the codon category of the node at the right end of the branch for sites detected to be (A) diversifying selection or (B) toggling, and (C) a previously identified HLA-associated polymorphism in gag (TW10). Potential escape and reversion mutations are mapped as red and green arrows, respectively. P, likelihood ratio test statistic p-value; D, diversifying selection model; T, toggling model. Both env and gag trees are rooted on HIV-1 subtype B.

Figure 5. Amino acid diversity at positively selected sites.

Amino acid sequence logos of positively selected sites, diversifying selection (D), toggling (T), or both (D&T), indexed by HXB2 position in nef are shown.