Defining the caprine γδ T cell WC1 multigenic array and evaluation of its expressed sequences and gene structure conservation among goat breeds and relative to cattle.

Workshop cluster 1 (WC1) molecules are part of the scavenger receptor cysteine-rich (SRCR) superfamily and act as hybrid co-receptors for the γδ T cell receptor and as pattern recognition receptors for binding pathogens. These members of the CD163 gene family are expressed on γδ T cells in the blood of ruminants. While the presence of WC1+ γδ T cells in the blood of goats has been demonstrated using monoclonal antibodies, there was no information available about the goat WC1 gene family. The caprine WC1 multigenic array was characterized here for number, structure and expression of genes, and similarity to WC1 genes of cattle and among goat breeds. We found sequence for 17 complete WC1 genes and evidence for up to 30 SRCR a1 or d1 domains which represent distinct signature domains for individual genes. This suggests substantially more WC1 genes than in cattle. Moreover, goats had seven different WC1 gene structures of which 4 are unique to goats. Caprine WC1 genes also had multiple transcript splice variants of their intracytoplasmic domains that eliminated tyrosines shown previously to be important for signal transduction. The most distal WC1 SRCR a1 domains were highly conserved among goat breeds, but fewer were conserved between goats and cattle. Since goats have a greater number of WC1 genes and unique WC1 gene structures relative to cattle, goat WC1 molecules may have expanded functions. This finding may impact research on next-generation vaccines designed to stimulate γδ T cells.

Goat γδ T cell subpopulations defined by WC1 expression, responses to pathogens and cytokine production.

The major functions of γδ T cells in mammals overlap with those of αß T cells but differ in that γδ T cells are rapid responders and see different types of antigens. While γδ T cells have been shown to be a major population of circulating lymphocytes in artiodactyl species such as cattle, sheep, and pigs, less is known about these cells in goats, an important agricultural species. We have recently shown that WC1, a γδ T cell-specific family of hybrid pattern recognition receptors/co-receptors, is a multigenic family in goats expanded beyond what occurs in cattle. This study was conducted to address some of the limitations of previous studies in determining the proportions of γδ T cells, WC1+ γδ T cells as well as the WC1.1+ and WC1.2+ subpopulations in blood and to evaluate their responses to various pathogens. Previously, the proportion of caprine γδ T cells was determined using a monoclonal antibody (mAb) 86D that we show here does not react with all γδ T cells thereby underestimating their contribution to the lymphocyte population. Using a mAb reactive with the TCRδ constant region we found the proportion of γδ T cells in blood was not significantly less than that of either CD4 or CD8 T cells and did not decrease with age after 6 months. γδ T cells that expressed WC1 ranged from ~20 to 85% of the total γδ T cells. Less than half of those were classified as WC1.1+ or WC1.2+ by mAb staining thus indicating a third major WC1+ population. We found that naïve γδ T cells proliferated in cultures of PBMC stimulated with antigens of Leptospira or Mycobacterium avium paratuberculosis (MAP) more than they did in control medium cultures or in those stimulated with M. bovis BCG antigens and that the responding γδ T cells included both WC1+ and WC1- cells. In ex vivo PMA/ionomycin-stimulated cultures of WC1- γδ T cells but not WC1+ cells produced both IL-17 and IFNγ. In longterm cultures with Leptospira or MAP both WC1- and WC1+ cells proliferated but only WC1- γδ T cells produced IL-17. In conclusion, goats have a substantial number of WC1- and WC1+ γδ T cells in PBMC that do not decrease with animal age after 6 months; both populations respond to bacterial antigens as naïve cells but in these cultures only the WC1- γδ cells produc IL-17 and IFNγ .