The core 2 beta-1,6-N-acetylglucosaminyltransferase-mucin encoded by bovine herpesvirus 4 was acquired from an ancestor of the African buffalo

Abstract

The Bo17 gene of bovine herpesvirus 4 (BoHV-4) is the only viral gene known to date that encodes a homologue of the cellular core 2 beta-1,6-N-acetylglucosaminyltransferase-mucin type (C2GnT-M). To investigate the origin and evolution of the Bo17 gene, we analyzed its distribution among BoHV-4 strains and determined the sequences of Bo17 from nine representative strains and of the C2GnT-M gene from six species of ruminants expected to encompass the group within which the gene acquisition occurred. Of 34 strains of BoHV-4, isolated from four different continents, all were found to contain the Bo17 gene. Phylogenetic analyses indicated that Bo17 was acquired from a recent ancestor of the African buffalo, implying that cattle subsequently acquired BoHV-4 by cross-species transmission. The rate of synonymous nucleotide substitution in Bo17 was estimated at 5 x 10(-8) to 6 x 10(-8) substitutions/site/year, consistent with previous estimates made under the assumption that herpesviruses have cospeciated with their hosts. The Bo17 gene acquisition was dated to around 1.5 million years ago. Bo17 sequences from BoHV-4 strains from African buffalo and from cattle formed two separate clades, estimated to have split about 700,000 years ago. Analysis of the ratio of nonsynonymous to synonymous nucleotide substitutions revealed a burst of amino acid replacements subsequent to the transfer of the cellular gene to the viral genome, followed by a return to a strong constraint on nonsynonymous changes during the divergence of contemporary BoHV-4 strains. The Bo17 gene represents the most recent of the known herpesvirus gene acquisitions and provides the best opportunity for learning more about this important process of viral evolution.

FIG. 1.

Phylogenetic relationships among C2GnT-M sequences from ruminants, with the human gene as the out-group, as estimated by NJ (A and B) and ML (C and D) analysis of DNA (A and C) and protein (B and D) sequences. Values on internal branches refer to the percentage of bootstrap replicates in which the branch was found; only values greater than 60% are shown. Horizontal branch lengths are drawn to scale, with the bars indicating 0.02 nucleotide (A and C) or amino acid (B and D) replacements per site.

FIG. 2.

Phylogenetic relationship of the BoHV-4 Bo17 gene to ruminant C2GnT-M DNA sequences as estimated by NJ (A and B) and ML (C and D) analysis with (A and C) or without (B and D) the human sequence as the out-group. BoHV-4 strains are presented in italics. Values on internal branches refer to the percentage of bootstrap replicates in which the branch was found; only values greater than 60% are shown. The relationship shown in panel A is inferred to be incorrect (see text). Horizontal branch lengths are drawn to scale, with the bars indicating 0.02 (A and C) or 0.01 (B and D) nucleotide replacements per site.

FIG. 3.

Distribution of the Bo17 gene among BoHV-4 strains. The DNA of 34 BoHV-4 strains isolated throughout the world (see Table 1) was analyzed by EcoRI restriction (A) and submitted to Southern blot analysis for the detection of the Bo17 gene (B). Marker sizes (in kilobase pairs) are indicated on the left.