NKG2D splice variants: a reexamination of adaptor molecule associations

NKG2D is a homodimeric C-type lectin-related receptor expressed on natural killer (NK) cells and T cells. In mice, alternative deoxyribonucleic acid (DNA) splicing generates two isoforms of NKG2D that differ in the length of their cytoplasmic domains. Their ability to induce cellular activation is mediated via association with two membrane-bound, signaling adaptor molecules, DAP10 and DAP12. It has been reported that the long form of NKG2D associates exclusively with DAP10, whereas the short variant can interact with either adaptor. The short isoform was reported to be almost undetectable in naïve NK cells. Using two distinct cell types, we demonstrate that like the short isoform, the long variant of NKG2D also associates not only with DAP10 but also with DAP12. Using reporter cells (70Z/3), we demonstrate that DAP12 can compete equally with DAP10 for association with both variants of NKG2D when DAP10 and DAP12 are coexpressed. Cross-linking either isoform of NKG2D induces a calcium flux when associated exclusively with DAP10 or DAP12. Moreover, using quantitative polymerase chain reaction (PCR), we also show that the short isoform of NKG2D is expressed in naïve NK cells. Our data suggest that signaling via mouse NKG2D isoforms is more complex than originally presented.