Different rates of (non-)synonymous mutations in astrovirus genes; correlation with gene function

Abstract

Background: Complete genome sequences of the Astroviridae include human, non-human mammalian and avian species. A consensus topology of astroviruses has been derived from nucleotide substitutions in the full-length genomes and from non-synonymous nucleotide substitutions in each of the three ORFs. Analyses of synonymous substitutions displayed a loss of tree structure, suggesting either saturation of the substitution model or a deviant pattern of synonymous substitutions in certain virus species.

Results: We analyzed the complete Astroviridae family for the inference of adaptive molecular evolution at sites and in branches. High rates of synonymous mutations are observed among the non-human virus species. Deviant patterns of synonymous substitutions are found in the capsid structural genes. Purifying selection is a dominant force among all astrovirus genes and only few codon sites showed values for the dN/dS ratio that may indicate site-specific molecular adaptation during virus evolution. One of these sites is the glycine residue of a RGD motif in ORF2 of human astrovirus serotype 1. RGD or similar integrin recognition motifs are present in nearly all astrovirus species.

Conclusion: Phylogenetic analysis directed by maximum likelihood approximation allows the inclusion of significantly more evolutionary history and thereby, improves the estimation of dN and dS. Sites with enhanced values for dN/dS are prominent at domains in charge of environmental communication (f.i. VP27 and domain 4 in ORF1a) more than at domains dedicated to intrinsic virus functions (f.i. VP34 and ORF1b (the virus polymerase)). Integrin recognition may play a key role in astrovirus to target cell attachment.

Figure 1

PAML trees of non-synonymous (A) and synonymous (B) substitutions in ORF1a (left panel), ORF1b (middle panel) and ORF2 (right panel) of astrovirus. PAML trees have been constructed by decorating the topology supplied by the input treefile with the branch lengths estimated by means of PAML approximation. The scale bars indicate the relative extent of mutational rates as (non-) synonymous substitutions per (non-) synonymous site corrected for multiple hits at the same site.

Figure 2

PAML trees of synonymous substitutions in the astrovirus capsid proteins VP34 (left panel) and VP27 (right panel). See text and legend to Fig 1.

Figure 3

Values for dN/dS of amino acid sites in relation to domain functions in astrovirus ORF1a (A), ORF1b (B) and ORF2 (C). For each site, the BEB-derived value of dN/dS (Y-axis) is plotted against the position of the residue in the polyprotein encoded by the ORF (X-axis). Arrows and scale bars indicate sites proposed for proteolytic digestion and predicted for phosphorylation (T, Y and S in ORF1b) as well as domain functions described in publications or predicted by servers. Numbering is according to human astrovirus serotype 1 Z25771. 3A: CC: coiled coil, TMH: transmembrane helix Nsp1a: non-structural protein 1a. 1: Region putatively involved in viral RNA helicase activity (1–80). 2: Protease domain (447–589). 3: Reported death domain and nuclear localization signal (620–714). 4: Hypervariable region and cell-growth induced deletion (760–838). 5: Ribosomal frameshifting signal (918–935). 6: Variable region prone to O-glycosylation and phosphorylation (608–632). 7: Putative VPg region associated with viral RNA replication (664–757). 8: KKXX-like ER retention (931–934). 3B: 1: Ribosomal frameshifting signal (1–28). 2: Predicted furin-type cleavage site (32–38). 3: Conserved polymerase domain (...-...). 3C: Rxxx: Arg-residues reported to mark tryptic digestion. T227: Thr-residue reported to bind viral RNA. 1a & 1b: NGR, RGD-like cell attachment motif (16–52). 2: LDV, RGD-like cell attachment motif (182–184). 3: Reported common epitope (340–376). 4: RGD: integrin recognition motif for cell attachment (572–574). 5: Predicted serotypic epitope (580–606). 6: Region of intracellular caspase digestion (701–787). 7: Region of extracellular tryptic digestion (300–423). 8, 9, 10: VP34, VP27, VP25, viral capsid proteins.

Figure 4

Routing of astrovirus sequence data from GenBank to PAML input data files. Main features of astrovirus genomes are given at the top. Numbers in parentheses indicate the length of an ORF in nucleotides. RFS points to the ribosomal frame shift signal between ORF1a and ORF1b. In GenBank, the number of complete ORF2 sequences surpasses those of ORF1a or ORF1b. At the ultimate right side, resources are boxed to indicate the actions undertaken and results obtained.