Thi Kim Anh Nguyen, Ad P. Koets, Martin Vordermeier, Peter J. Jervis, Liam R. Cox, Simon P. Graham, Wiebren J. Santema, D. Branch Moody, Serge van Calenbergh, Dirk M. Zajonc, Gurdyal S. Besra, Ildiko Van Rhijn

Although CD1d and NKT cells have been proposed to have highly conserved functions in mammals, data on functions of CD1d and NKT cells in species other than humans and rodents are lacking. Upon stimulation with the CD1d-presented synthetic antigen α-galactosylceramide, human and rodent type I invariant NKT cells release large amounts of cytokines. The two bovine CD1D (boCD1D) genes have structural features that suggest that they cannot be translated into functional proteins expressed on the cell surface. Here we provide evidence that despite an intron-exon structure and signal peptide that are different from all other known CD1 genes, boCD1D can be translated into a protein that is expressed on the cell surface. However, in vivo treatment of cattle (Bos taurus) with 0.1, 1, or 10 µg kg⁻¹ of the most commonly used α-galactosylceramide, which has a C26 fatty acid, did not lead to an increase in body temperature and serum cytokine levels of the animals. This lack of reactivity is not due to a complete inability of boCD1d to present glycosphingolipids because α-galactosylceramide variants with shorter fatty acids could be presented by boCD1d to human NKT cells in vitro. This suggests that the natural ligands of boCD1d are smaller lipids.