Interferon-gamma priming effects in the activation and deactivation of ISGF3 in K562 cells

ISGF3 is a major protein factor which mediates the transcriptional activation of interferon-inducible genes. ISGF3 is composed of two subunits, ISGF3 gamma and ISGF3 alpha, which are stimulated by interferon-gamma (IFN-gamma) and interferon-alpha (IFN-alpha), respectively. In this paper, the effect of pretreatment of IFN-gamma on the response of K562 cells to IFN-alpha, termed "gamma-priming," is examined. Using techniques of gene transfection and mobility shift assay, we studied the gamma-priming effects on the kinetics of appearance and disappearance of (i) the protein product of a luciferase reporter gene driven by an IFN-inducible promoter and (ii) the binding of ISGF3 to the interferon-stimulated response element. We found that exposure of cells to IFN-gamma prior to IFN-alpha greatly increased both the levels and the rate of ISGF3 binding activity during the early phase of IFN-alpha treatment and caused the amount of ISGF3 bound to the interferon-stimulated response element to decrease more rapidly with time during the later phase. Such effects were reflected also on the kinetics of expression of the interferon-inducible luciferase gene induced by IFN-alpha. In order to understand the basis of these gamma-priming effects, we use an in vitro reconstitution method to examine the kinetics of the subcomponents of ISGF3 in response to different IFN treatments in K562 cells. It was found that gamma-priming not only increased the levels of ISGF3 gamma, but it also stimulated both the rates of rise and fall of the activated alpha-component of ISGF3. A molecular model is proposed to explain these findings.