The activating immune receptor NKG2D binds to multiple stress-induced ligands that are structurally different: MHC-class I-related Chain (MIC) A/B molecules have a transmembrane domain whereas most UL16 binding proteins (ULBPs) are glycosylphosphatidyl-inositol (GPI)-linked molecules. The significance of this variability in membrane anchor is unclear. Here, we demonstrate that ULBP2, but not ULBP1 or 3, can reach the cell surface without the GPI modification. Several proteins are expressed at the cell surface as both transmembrane and GPI-linked molecules, either via alternative splicing or by the expression of linked genes. However, to our knowledge, ULBP2 is the first single mammalian cDNA that can be expressed as either a transmembrane or a GPI-anchored protein. The rate of maturation and the levels of cell surface expression of the non-GPI linked form were lower than those of the GPI-linked ULBP2. Nonetheless, non-GPI ULBP2 was recognised by NKG2D and triggered NK cell cytotoxicity. These data show that differences in membrane attachment by NKG2D-ligands are more important for regulation of their surface expression than for cytotoxic recognition by NKG2D and emphasise that detailed characterization of the cell biology of individual NKG2Dligands will be necessary to allow targeted modulation of this system

The authors would like to thank Drs Medoff, Stevens and Vainauskas for GPIdeficient cell lines; C. Gross and F. Colucci for critically reading the manuscript. This work was supported by the MRC (New Investigator Grant to MVG) and FIS (PS09/00181 and PI08/1701). OA was supported by the Leukaemia Research Fund and The Newton Trust. SA was supported by Fundación Caja Madrid and Ibercaja.

Lola Fernández-Messina ...et al.

Peer reviewed