The research in this application will accomplish three specific objectives regarding the biochemistry and molecular biology of the Type I interferon (IFN) receptor. The first objective is to develop procedures for the preparation of highly purified interferon receptor from human and bovine tissues. There are, to our knowledge, no receptor studies deomonstrating the binding of radiolabeled IFN to receptors derived from the homogenates of human and animal tissues. The procedures in this proposal include: the preparation of membranes by differential centrifugation of tissue homogenates; extraction of the interferon receptor from the membranes with non-denaturing detergents and the subsequent purification of the receptor by ion-exchange and gel-permeation chromatography, preparative electrophoresis, isoelectric focusing, high performance liquid chromatography (HPLC), lectin-specific and affinity chromatography. The second objective is to provide a rigorous biochemical characterization of the Type I interferon receptor. The characterization will include: the use of density gradient centrifugation, gel-filtration, isoelectric focusing, covalent and photoaffinity labeling and determination of putative carbohydrate moieties of the receptor. Information about the molecular weight, possible subunit composition and carbodydrate composition of the receptor will be obtained. The third objective is to examine the role of the Type I IFN receptor in the interferon response in cultured cells. Through the use of photoaffinity labeling the determination of Type I interferon receptor structures in a series of cell lines which exhibit differential responses to interferon will be compared to the effect(s) of antireceptor monoclonal antibodies on the biological activities of interferon. It is intended that possible differences in the structure of the receptors will be related to the biological activities of interferon and that this correlation will serve to establish that there are separate and distinct effector systems which can interact with the Type I interferon receptor and elicit the well-established variety of biological responses of interferons. The existence of different effector systems for the Type I interferon receptor would provide for an explanation of the observed deficiencies of certain cell types with respect to the antiviral and, in particular, the antiproliferative activities of interferon.
