Genomic organization and evolution of the ULBP genes in cattle

The cattle UL16-binding protein 1 (ULBP1) and ULBP2 genes encode members of the MHC Class I superfamily that have homology to the human ULBP genes. Human ULBP1 and ULBP2 interact with the NKG2D receptor to activate effector cells in the immune system. The human cytomegalovirus UL16 protein is known to disrupt the ULBP-NKG2D interaction, thereby subverting natural killer cell-mediated responses. Previous Southern blotting experiments identified evidence of increased ULBP copy number within the genomes of ruminant artiodactyls. On the basis of these observations we hypothesized that the cattle ULBPs evolved by duplication and sequence divergence to produce a sufficient number and diversity of ULBP molecules to deliver an immune activation signal in the presence of immunogenic peptides. Given the importance of the ULBPs in antiviral immunity in other species, our goal was to determine the copy number and genomic organization of the ULBP genes in the cattle genome.

Sequencing of cattle bacterial artificial chromosome genomic inserts resulted in the identification of 30 cattle ULBP loci existing in two gene clusters. Evidence of extensive segmental duplication and approximately 14 Kbp of novel repetitive sequences were identified within the major cluster. Ten ULBPs are predicted to be expressed at the cell surface. Substitution analysis revealed 11 outwardly directed residues in the predicted extracellular domains that show evidence of positive Darwinian selection. These positively selected residues have only one residue that overlaps with those proposed to interact with NKG2D, thus suggesting the interaction with molecules other than NKG2D.

The ULBP loci in the cattle genome apparently arose by gene duplication and subsequent sequence divergence. Substitution analysis of the ULBP proteins provided convincing evidence for positive selection on extracellular residues that may interact with peptide ligands. These results support our hypothesis that the cattle ULBPs evolved under adaptive diversifying selection to avoid interaction with a UL16-like molecule whilst preserving the NKG2D binding site. The large number of ULBPs in cattle, their extensive diversification, and the high prevalence of bovine herpesvirus infections make this gene family a compelling target for studies of antiviral immunity.