The sequence and phylogenetic analysis of avian reovirus genome segments M1, M2, and M3 encoding the minor core protein μA, the major outer capsid protein μB, and the nonstructural protein μNS

Abstract

The sequences and phylogenetic analyses of the M-class genome segments of 12 avian reovirus strains are described. The S1133 M1 genome segment is 2283 base pairs long, encoding a protein μA consisted of 732 amino acids. Each M2 or M3 genome segment of 12 avian reovirus strains is 2158 or 1996 base pairs long, respectively, encoding a protein μB or μNS consisted of 676 and 635 amino acids, respectively. The S1133 genome segment has the 5′ GCUUUU terminal motif, but each M2 and M3 genome segment displays the 5′ GCUUUUU terminal motif which is common to other known avian reovirus genome segments. The UCAUC 3′-terminal sequences of the M-class genome segments are shared by both avian and mammalian reoviruses. Noncoding regions of both 5′- and 3′-termini of the S1133 M1 genome segment consist of 12 and 72 nucleotides, respectively, those of each M2 genome segment consist of 29 and 98 nucleotides, respectively, and those of each M3 genome segment are 24 and 64 nucleotides, respectively. Analysis of the average degree of the M-class gene and the deduced μ-class protein sequence identities indicated that the M2 genes and the μB proteins have the greatest level of sequence divergence. Computer searches revealed that the μA possesses a sequence motif (NH 2-Leu-Ala-Leu-Asp-Pro-Pro-Phe-COOH) (residues 458–464) indicative of N-6 adenine-specific DNA methylase. Examination of the μB amino acid sequences indicated that the cleavage site of μB into μBN and μBC is between positions 42 and 43 near the N-terminus of the protein, and this site is conserved for each protein. During in vitro treatment of virions with trypsin to yield infectious subviral particles, both the N-terminal fragment δ and the C-terminal fragment φ were shown to be generated. The site of trypsin cleavage was identified in the deduced amino acid sequence of μB by determining the amino-terminal sequences of φ proteins: between arginine 582 and glycine 583. The predicted length of δ generated from μBC is very similar to that of δ generated from mammalian reovirus μ1C. Taken together, protein μB is structurally, and probably functionally, similar to its mammalian homolog, μ1. In addition, two regions near the C-terminal and with a propensity to form α-helical coiled-coil structures as previously indicated are observed for each protein μB. Phylogenetic analysis of the M-class genes revealed that the predicted phylograms delineated 3 M1, 5 M2, and 2 M3 lineages, no correlation with serotype or pathotype of the viruses. The results also showed that M2 lineages I–V consist of a mixture of viruses from the M1 and M3 genes of lineages I–III, reflecting frequent reassortment of these genes among virus strains.