Human miR-27a* serves as a regulator of Prf1 and GzmB protein expression. (A) Basal suppression of GzmB and Prf1 expression by miR-27a* under resting conditions. (Top) IB analysis. (Bottom) Cr release assay at ET5. (B-C) miR-27a* regulates the level of Prf1 and GzmB protein during activation. mNK cells were freshly incubated in the presence of IL-15 (60 ng/mL) after 24-hour deprivation of IL-15. The cells were harvested at the times (0, 6, 12, and 30 hours) indicated relative to treatment of 60 ng/mL IL-15 and analyzed for kinetics of Prf1/GzmB mRNA and miR-27a* (B top). mNK cells were left untreated or treated for 6 hours with various concentrations of IL-15 (0, 30, and 60 ng/mL) after 24-hour deprivation of IL-15. miR-27a* (B middle) and Prf1/GzmB mRNA (B bottom) were analyzed by real-time qPCR. Concurrently, the mNK cells were subjected to IB analysis (C top) and Cr release assay (C bottom). (D) High expression of Prf1 and GzmB protein in NK-92 is because of low level of endogenous miR-27a*. (Top) IB analysis. (Middle) Semi-qPCR analysis. (Bottom) Real-time qPCR. Data are representative of 3 independent experiments (mean ± SEM of triplicates; Student t test, ***P < .001 vs Ctrl_miR.