c-Myc overexpression leads to an increase in KIR expression during NK-cell development. CD34⁺ cells were isolated from umbilical cord blood and transduced with MSCV retroviral constructs containing egfp or c-myc. These cells were cultured on the EL08.1D2 cell line in the presence of exogenous cytokines. After 21 days, cultured cells were harvested for (A) quantitative RT-PCR to determine c-myc transcript levels. All values are normalized to GAPDH (n = 5). Error bars represent the SEM for each group of samples. *P < .05. (B) These cells were also immunophenotyped with APC-conjugated NCAM16.2 and PE-conjugated DX9, EB6, GL183, and FES172 monoclonal antibodies. The FACS plots in panel B are representative examples of cells harvested from day 21 cultures. (C) The percentage of KIR⁺ NK cells in eGFP- and c-Myc–transduced cultures was determined by FACS analysis (n = 28). (D) RNA was harvested from day 21 cultures and used for quantitative RT-PCR using primers designed to amplify coding KIR transcripts and to detect transcripts originating from the distal promoter element for KIR2DL3, KIR2DL1/2DL2/2DL3/2DS1/2DS2 (2D distal), and KIR3DL1 (n = 8). Expression levels were normalized to an IL-2–activated peripheral blood NK population known to express all KIR genes. Error bars represent the SEM for each group of samples. *P < .05.