AbstractThe Ly-49 multigene receptor family regulates mouse NK cell functions. A number of Ly-49 genes exhibit allelic variation, but the functional significance of allelic differences in extracellular domains of Ly-49 receptors regarding ligand specificity is largely unknown. Amino acid differences exist in the extracellular domains of the B6 and BALB/c allele products of the inhibitory Ly-49G receptor. We constructed chimeric Ly-49 receptors consisting of common cytoplasmic and transmembrane regions of the activating Ly-49W receptor fused with the ectodomains of the B6 and BALB/c alleles of Ly-49G. Expression of these chimeras in the RNK-16 rat NK cell line allowed us to study the specificity of inhibitory receptor ectodomains as they stimulated NK lytic activity. We found that the ectodomain of the BALB/c allele of Ly-49G recognizes both H-2Dd and Dk class I MHC alleles, whereas the ectodomain of the B6 allele of Ly-49G recognizes Dd, and not Dk. The specificity for Dk as well as Dd of the wild-type Ly-49GBALB/c allele product was confirmed with RNK-16 transfectants of this inhibitory receptor. Furthermore, the ectodomain of the Ly-49GBALB/c allele recognizes a distinct repertoire of xenogeneic ligands that only partially overlaps with that recognized by Ly-49GB6. Our results indicate that allelic variation in Ly-49 extracellular domains can have functional significance by altering Ly-49 receptor specificity for mouse class I MHC and xenogeneic ligands.