One project is focused on the structure of the receptor for Interferon- gamma (IFN-gamma). We have characterized 2 subunits (Mr of 38 Kd and 200 Kd) by SDS-PAGE analysis of biosynthetically labelled human monocytes. These subunits are in addition to the previously cloned IFN-gamma binding protein (Mr, 80-90kD). We are now beginning to preparatively purify these subunits so that eventually antibodies against them can be raised. We are also investigating whether or not somatic cell hybrids expressing combinations of the transfected IFN-gamma binding protein and chromosome 21 exhibit the ability to co-immunoprecipitate the 38 and 200 Kd subunits. Another project is directed toward an understanding of the role of phosphorylation of the IFN-gamma receptor during signal transduction. When exposed to the phorbol ester, PMA, THP1 cells, a monocyte-like cell line, now become capable of responding to IFN-gamma as measured by the induction of the gene for IP-10, an IL-8/PF4 family member. The mechanisms underlying the development of competency in these cells for IFN-gamma responsiveness is under investigation. Whether or not phosphorylation by various kinases or dephosphorylation by phosphatases are involved in this mechanism is under study. In parallel to this, we are also studying the ability of IFN-gamma to induce phosphorylation of the IFN-gamma receptor on human monocytes. Using a phosphatase inhibitor, such as okadaic acid, we have been able to measure phosphorylation of the receptor. We are determining if this is necessary for signal transduction as measured by the expression of mRNA for the IP-10 and the FcgammaRI genes. We are also involved in examining the mechanisms underlying the ability of IL-4 to inhibit many of the effects of IFN-gamma and IFN-alpha. DNA sequences in the promotor regions of IFN-inducible genes will be identified and tested for their role in the inhibitory effect of IL-4 on IFN-induced gene expression.
