Diefenbach and colleagues identified two splice variants of mouse NKG2D — a long form (NKG2D-L) that has a 13-amino-acid extension at the amino terminus, and a short form (NKG2D-S). Resting NK cells express messenger RNA encoding NKG2D-L, and after activation, mRNA encoding NKG2D-S is upregulated. Neither isoform can be detected in resting macrophages or CD8+ T cells. After T-cell receptor stimulation, the expression of both isoforms is upregulated on CD8+ T cells, whereas stimulation of macrophages with bacterial lipopolysaccharide upregulates the expression of NKG2D-S preferentially. Immunoprecipitation studies showed that the two isoforms associate differentially with the adaptor molecules DAP10 and DAP12 (also known as KARAP) — NKG2D-L associates only with DAP10, whereas NKG2D-S can associate with both DAP10 and DAP12. Because CD8+ T cells do not express DAP12, the two NKG2D isoforms that are expressed by activated T cells can interact only with DAP10, whereas activated NK cells can transmit signals through DAP10 and DAP12. Could the failure of T cells to respond directly to signals through NKG2D in the absence of co-stimulation be due to the absence of DAP12 in these cells? Ectopic expression of DAP12 in CD8+ T cells allowed a direct stimulatory signal to be transmitted through NKG2D-S. NK cells from Dap12−/− mice have defective signalling through NKG2D, which would not have been predicted from previous results.
Gilfillan and colleagues generated Dap10−/− mice to investigate the function of this adaptor. Although DAP10 was known previously to associate only with NKG2D, expression studies showed that the expression of NKG2D is low, but detectable, on NK cells, but absent on T cells. Activation of NK cells resulted in the upregulation of expression of NKG2D, but this did not occur when T cells were activated. How can NKG2D be expressed by NK cells in the absence of DAP10? The obvious explanation is that NKG2D can associate with another adaptor molecule, possibly DAP12. Immunoprecipitation studies showed that this is indeed the case.
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