Purpose: Natural killer (NK) cells represent a specialized lymphoid population of the innate immune response against tumor or virally infected cells. NK cell cytotoxicity is closely regulated by numerous inhibitory and activating receptors. Major Histocompatibility Complex (MHC) I acts as an inhibitory receptor by presenting normal self antigen on nucleated cells, thus sparing the healthy cell from NK cell mediated lysis. On cells lacking MHC I other activating receptors, such as 2B4, NKG2D, and a group termed the Natural Cytotoxicity Receptors (NCRs), induce cytotoxicity and subsequent target cell death. The NCRs play a key role in recognition and killing of MHC I deficient cells and include the receptors NKp30, NKp46, and NKp44. Binding of one or more of these receptors to specific, pathologically induced ligands induces strong NK mediated cytotoxicity. NKp44 is of particular interest because it is only expressed on activated NK cells, implicated in dramatic increases in cytotoxicity, and involved in HIV infection. Ligands for NKp44 have been reported but not yet characterized. Previous work in our lab utilizing a NKp44-IgG fusion protein and FACS analysis has shown that a B cell lymphoma cell line (DB) expresses a ligand for NKp44. The purpose of this study is to clone and characterize this ligand to further understand the interactions between NKp44 and target cells.
Methods: Messenger RNA was isolated from DB cells and constructed into a directional complimentary DNA (cDNA) library for transformation into ultra competent E. coli bacteria. The cDNA library was amplified and then transiently transfected into Cos cells.
Results: The cDNA encoding the NKp44 ligand was isolated utilizing mammalian expression cloning and sequenced.
Conclusions: Characterization of this ligand and further understand of ligand/receptor interactions of NKp44 will help to develop future strategies in NK cell modulation in immunotherapy.