Asymmetric evolution in viral overlapping genes is a source of selective protein adaptation

Abstract

Overlapping genes represent an intriguing puzzle, as they encode two proteins whose ability to evolve is constrained by each other. Overlapping genes can undergo "symmetric evolution" (similar selection pressures on the two proteins) or "asymmetric evolution" (significantly different selection pressures on the two proteins). By sequence analysis of 75 pairs of homologous viral overlapping genes, I evaluated their accordance with one or the other model. Analysis of nucleotide and amino acid sequences revealed that half of overlaps undergo asymmetric evolution, as the protein from one frame shows a number of substitutions significantly higher than that of the protein from the other frame. Interestingly, the most variable protein (often known to interact with the host proteins) appeared to be encoded by the de novo frame in all cases examined. These findings suggest that overlapping genes, besides to increase the coding ability of viruses, are also a source of selective protein adaptation.

Keywords: Ancestral frame; De novo frame; Homologs; Non-synonymous nucleotide substitution; Selection pressure; Synonymous nucleotide substitution; Virus adaptation.

Fig. 1

Orientation of overlapping genes, with the downstream frame having a shift of one nucleotide 3′ with respect to the upstream frame. There are 3 types of codon position (cp): cp13 (bold character), in which the first position of the upstream frame overlaps the third position of the downstream frame; cp21 (underlined character), in which the second position of the upstream frame overlaps the first position of the downstream frame; cp32 (italic character), in which the third position of the upstream frame overlaps the second position of the downstream frame. Based on the genetic code, a nucleotide substitution at first codon position causes an amino acid change in 95.4% of cases, at second codon position in 100% of cases, and at third codon position in 28.4% of cases. Thus, nucleotide substitutions at the codon positions “13” and “32” are usually non-synonymous in one frame and synonymous in the other. Nucleotide substitutions at the codon position “21” are almost all non-synonymous in both frames.

Fig. 2

Analysis of the amino acid diversity in the 75 pairs of homologous overlapping genes. Each pair of columns shows: i) the percent amino acid identity between the protein encoded by the upstream frame of the overlap and that encoded by the homolog (dark column); ii) the percent amino acid identity between the protein encoded by the downstream frame of the overlap (shifted of one nucleotide 3′ with respect to the upstream frame) and that encoded by the homolog (gray column). The horizontal line separates well-conserved homologous pairs (aa identity >50%) from not well-conserved homologous pairs (aa identity <50%). (A) Subset of the 37 overlapping genes under symmetric evolution. (B) Subset of the 38 overlapping genes under asymmetric evolution. The numbering of overlapping genes is in accordance with that given in Supplementary Table S1. The underlined numbers indicate the overlaps in which the pattern of symmetric evolution (4 cases out of 37) or that of asymmetric evolution (6 cases out of 38) was not confirmed by chi-square analysis of the nucleotide diversity.

Fig. 3

Correlation between the normalized chi-square value (from analysis of amino acid substitutions) and the absolute value (Abs) of the difference between the percent frequency (%F) of nucleotide substitutions at the codon position “32” (%F.cp32) and that at the codon position “13” (%F.cp13). Empty circles indicate the 33 overlapping genes under symmetric evolution. Black circles indicate the 32 overlapping genes under asymmetric evolution.