Abstract
Macrophages also possess a calmodulin (CaM)-dependent myosin light chain kinase. The accumulation of calcium by phagocytic vesicles isolated from pulmonary alveolar macrophages is stimulated by CaM, and this stimulation is partially inhibited by trifluoperazine. Studies on the murine macrophage-like cell line J774 have demonstrated that trifluoperazine and W7 inhibit cell growth, migration, Fc-mediated phagocytosis, and phagocytosis of latex particles. A band of calmodulin-binding activity in extracts of J774 was detected by exposure to 125I-labeled CaM ([125I]CaM) followed by polyacrylamide gel electrophoresis in calcium-containing buffers. A two-stage immunologic approach was then taken to purify and study this protein: First, a rabbit antibody to the partially purified protein was obtained and a radioimmunoassay established. Utilizing the experience and information derived therefrom, monoclonal antibodies to the binding protein were produced. These were then used to immunopurify the binding protein, and to study its biosynthesis and cellular distribution. The results indicate that the J774 CaM-BP is a macrophage-specific, membrane-associated glycoprotein. The major CaM-BP in the macrophage-like cell line J774 is widely applicable not only to other calmodulin-binding proteins, but to any protein that binds ligands that can be specifically radiolabeled.
Published Version
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