Quantifying Selection against Synonymous Mutations in HIV-1 env Evolution

Abstract

Intrapatient evolution of human immunodeficiency virus type 1 (HIV-1) is driven by the adaptive immune system resulting in rapid change of HIV-1 proteins. When cytotoxic CD8؉ T cells or neutralizing antibodies target a new epitope, the virus often escapes via nonsynonymous mutations that impair recognition. Synonymous mutations do not affect this interplay and are often assumed to be neutral. We test this assumption by tracking synonymous mutations in longitudinal intrapatient data from the C2-V5 part of the env gene. We find that most synonymous variants are lost even though they often reach high frequencies in the viral population, suggesting a cost to the virus. Using published data from SHAPE (selective 2=-hydroxyl acylation analyzed by primer extension) assays, we find that synonymous mutations that disrupt base pairs in RNA stems flanking the variable loops of gp120 are more likely to be lost than other synonymous changes: these RNA hairpins might be important for HIV-1. Computational modeling indicates that, to be consistent with the data, a large fraction of synonymous mutations in this genomic region need to be deleterious with a cost on the order of 0.002 per day. This weak selection against synonymous substitutions does not result in a strong pattern of conservation in cross-sectional data but slows down the rate of evolution considerably. Our findings are consistent with the notion that large-scale patterns of RNA structure are functionally relevant, whereas the precise base pairing pattern is not. Human immunodeficiency virus type 1 (HIV-1) evolves rapidly within a single host during the course of the infection. This evolution is driven by strong selection imposed by the host immune system via cytotoxic CD8 ϩ T cells (CTLs) and neutralizing antibodies (nAbs) (1) and is facilitated by HIV-1's high mutation rate (2, 3). Escape mutations in epitopes targeted by CTLs are typically observed during early infection and spread rapidly through the population (4). During chronic infection, the most rapidly evolving parts of the HIV-1 genome are the variable loops (V1 to V5) in the envelope protein gp120 (V loops), which change to avoid recognition by nAbs. Escape mutations in env, the gene encoding gp120, spread through the viral population within a few months. Consistent with this time scale, it is found that serum from a particular time typically neutralizes autologous virus extracted more than 3 to 6 months earlier but not contemporary virus (5).Escape mutations are selected because they change the amino acid sequences of viral proteins in a way that reduces antibody binding or epitope presentation. Conversely, synonymous mutations do not modify the viral proteins and are commonly used as approximately neutral markers in studies of viral evolution, i.e., as a negative control for detecting selected sites (6-8). In addition to maintaining protein function and avoiding the adaptive immune recognition, however, the HIV-1 genome has to ensure efficient processing and translation, nuclear export, and packaging ...

Experimental Estimation of the Effects of All Amino-Acid Mutations to HIV’s Envelope Protein on Viral Replication in Cell Culture

“…This overall trend is consistent with the fact that synonymous mutations usually have smaller functional effects than nonsynonymous mutations. However, synonymous mutations can sometimes have substantial effects [21, 80–82], particularly in viruses like HIV that are under strong selection for RNA secondary structure and codon usage [83, 84]. To assess selection on synonymous mutations on a more site-specific level, we examined the change in frequency of multi-nucleotide codon mutations across env ’s primary sequence (Fig 4).…”

Uneven Genetic Robustness of HIV-1 Integrase

“…While it has previously been suggested that HIV-1 RNA structure might constrain variability and evolution (72)(73)(74), silent mutations introduced into some of the most highly conserved RNA hairpins in HIV-1 result in no significant virus replication defects (75). Similarly, introduction of synonymous mutations into other regions of the HIV-1 genome with apparently strong purifying selection against synonymous substi- tutions does not reduce viral fitness in vitro (76).…”

Identifying Selection in the Within-Host Evolution of Influenza Using Viral Sequence Data

“…Weak selection acting upon synonymous mutations has been identified for codon usage in influenza [45] and against mutations that disrupt RNA structure in HIV [46], although the magnitude of selection inferred here is significantly higher than in either case. While inferring the presence of selection, our method cannot match occurrences of selection to specific biological mechanisms; further data would generally be required to do this.…”