Arginine residue in the transmembrane domain of KIR2DL1 plays an important role in its inhibitory function. Arginine in the transmembrane domain of KIR2DL1*010 was replaced with alanine. A stable YT-Indy cell line expressing the KIR2DL1*010 mutant was generated. YT-Indy transfected with vector was used as control. (A) Specific killing of YT-Indy expressing wild-type KIR2DL1*010 (*010), KIR2DL1*010 mutants (*010 R245A), and KIR2DL1*00401 (*00401) was assessed against 721.221-Cw6 by BADTA release assay. *P = .62, **P < .05. (B) Relative expression of CD107 at the surface of YT-Indy cells expressing wild-type (*010) and mutated (*010 R245A) KIR2DL1*010 was assessed by CD107 mobilization assay. **P < .05. (C) Production of IFN-γ was assessed after YT-Indy cells expressing wild-type (top) and mutated (bottom) KIR2DL1*010 were stimulated with target 721.221-Cw6 cells. Results are representative of 3 independent experiments. (D) YT-Indy cells expressing wild-type and mutated KIR2DL1*010 alleles were mixed with target 721.221-Cw6 cells at a 10:1 (E/T) ratio and injected into mice subcutaneously. A total of 10 mice, 5 for each group, were given injections. After tumors reached 20% of the body mass in some mice, the mice were humanely killed, dissected, and photographed. The image of a representative tumor from each group is shown. The tumor shown at left is from a mouse that was given YT-Indy expressing wild-type KIR2DL1*010 (*010), and the tumor shown at right is from a mouse that was given YT-Indy cells expressing the mutated KIR2DL1*010 (*010 R245A) allele. Mass and volume of these tumors are shown at bottom. **P < .05, ***P < .01. The experiments were repeated 3 times. Error bars represent SD.