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IL-12 exerts several regulatory effects on natural killer (NK) cells by activating IL-12 signaling. IL-12 signaling is tightly auto-regulated to control its onset and termination, with prolonged IL-12 treatment resulting in IL-12 hyporesponsiveness. However, the mechanisms underlying IL-12 auto-regulation are still unclear. In this study we report that prolonged IL-12 treatment significantly up-regulates microRNAs (miRNAs), including miR-132, -212, and -200a in primary human NK cells. This up-regulation correlates temporally with gradually decreasing STAT4 levels and decreasing IFN-γ expression, after an initial increase within the first 16 hours of IL-12 treatment. The IL-12 hyporesponsiveness is dependent on IL-12 concentration, and associated up-regulation of miR-132, -212, and -200a. Furthermore, IL-12-hyporesponsive cells regain responsiveness of IFN-γ production 24 hours after IL-12 removal, which correlates with decreases in miR-132, -212, and -200a levels. Overexpression of miR-132, -212, and -200a by transfection into NK cells mimics IL-12 priming, inducing IL-12 hyporesponsiveness, whereas transfection of miR-132, -212, and -200a inhibitors largely abolishes IL-12 induction of IL-12 hyporesponsiveness. These data suggest that miR-132, -212, and -200a up-regulation during prolonged IL-12 treatment, negatively regulates the IL-12 signaling pathway by reducing STAT4 expression in primary human NK cells.

The effector domain of the Rev protein is a nuclear export signal (NES) that is responsible for transporting Rev and its bound congeners out of the nucleus and into the cytoplasm. Previous work has identified several critical residues in the NES and has led to the belief that NESs of the Rev type are necessarily leucine rich. Here we present the sequences of a large number of functional Rev molecules with NES mutations. The data indicate a previously unreported diversity in allowable residues at a number of positions, including each of the leucine residues previously considered essential.