Lyssavirus P Gene Characterisation Provides Insights into the Phylogeny of the Genus and Identifies Structural Similarities and Diversity within the Encoded Phosphoprotein

Abstract

A comprehensive phylogenetic analysis of the Lyssavirus genus, employing P gene sequences from 128 isolates recovered globally, is presented. While confirming prior suggestions of the genetic distinctness of the Australian bat lyssaviruses, these data also revealed a clear division within the rabies virus clade (Genotype 1) between globally distributed viruses circulating predominantly in canid species (subgroup 1-1), and American strains harbored by both chiropteran and terrestrial hosts (subgroup 1-2). Nucleotide substitution patterns within the P locus suggested differential selection operating along the length of the open reading frame (ORF) between rabies viruses of these two subgroups. Comparison of the deduced primary sequences of the encoded phosphoproteins of all isolates provided insights into the product's structural organization. Two conserved (CD1,2) and two variable (VD1,2) domains were evident; high variation in the VD2 region was reflected by differences in hydropathic profiles. Only two of five serine residues reported to function as phosphate acceptors in the P protein of the rabies challenge virus standard (CVS) strain were absolutely conserved; similarly, out of four internal methionines reported to direct internal translation initiation of the CVS strain to produce N-terminally truncated P proteins, only Met(20) was universally retained. In contrast, two sequence motifs, one believed to confer binding to the cytoplasmic dynein light chain LC8, and a lysine-rich sequence probably contributing to N protein binding, were conserved throughout the genus. Most rabies viruses of the carnivora (1-1) contain a potential C ORF in alternate frame to that of P, a feature limited or absent in most other isolates of the genus, an observation interpreted with consideration to the predicted course of lyssavirus evolution.

Genetic Analysis of Phosphoprotein and Matrix Protein of Rabies Viruses Isolated in Brazil

“…Taken together, these findings suggest that phylogenetic analyses using these two genes are capable of identifying epidemiological characteristics of Brazilian RVs. Conservation of the P protein in Brazilian RV isolates was low, with two conserved (CD1 and 2) and two variable domains (VD1 and 2), which corroborates the findings of Nadin-Davis et al [30,32]. Variable amino acid substitutions characteristic of Brazilian RV variants occurred at high frequencies in the variable domains, and this was reflected in the hydrophilic regions of the observed hydropathic profiles.…”

“…With the exception of several RV isolates, the Ser and Ala differences were observed in the consensus sequences of isolates from either the carnivora-or chiroptera-related RV isolates, as reported by Nadin-Davis et al [32]. The variable domains thought to be located on the surface structure of the P protein have been suggested to be involved in host/viral interactions and adaptation to the host environment [31,32]. Our results support this hypothesis and studies of the variable domains are expected to be useful in elucidating the adaptive evolution of the virus.…”

“…In addition, Brazilian carnivorarelated RVs formed a monophyletic cluster and were divided into dog-and fox-related RV lineages. Based on multiple alignments of P protein amino acid sequence data, two conserved domains (CD1 and 2) comprising positions 1-50 and 201-245, respectively, and two variable domains (VD1 and 2) comprising positions 61-80 and 134-180, respectively, have been identified [30,32]. These conserved and variable domains were observed in the Brazilian RV isolates of this study (Fig.…”

“…Additionally, the LC8-binding motif, involved in viral nucleocapsid axoplasmic transport [21,39], formed consensus sequence K(S/A)TQT in the VD2 region. With the exception of several RV isolates, the Ser and Ala differences were observed in the consensus sequences of isolates from either the carnivora-or chiroptera-related RV isolates, as reported by Nadin-Davis et al [32]. The variable domains thought to be located on the surface structure of the P protein have been suggested to be involved in host/viral interactions and adaptation to the host environment [31,32].…”

“…In addition, few studies on P and M proteins in wild-type RV strains have been reported to date [30,31,32,33], and no reports have genetically characterized the P and M genes in Brazilian RV isolates. In this study, we conducted genetic analyses of P and M genes of several RV field isolates prevalent in Brazil.…”