The goal of this Program Project Grant is to advance cellular, biochemical and molecular understanding of phagocyte biology, microbicidal systems and clinically relevant disorders of antimicrobial function. The target cell is the human neutrophil and the major theme is the fundamental basis for oxygen-dependent systems in the inflammatory response and host defenses. Three projects, each directed by an established investigator, constitute an integrated approach to the scientific goals. Project 1 will determine the structure and function of cytosolic proteins that play essential roles in activation of the superoxide-generating NADPH oxidase system. The studies include characterization of the inherited deficiency state of two cytosolic proteins in two different autosomal forms of chronic granulomatous disease. In Project 2 electron spin resonance spectroscopy will be the key analytical tool for defining physical chemical and cellular factors that modulate the formation of hydroxyl radical by activated neutrophils. Project 3 deals with molecular aspects of the expression and intracellular processing of myeloperoxidase, an important enzymatic constituent of oxygen-dependent microbicidal function. The Xenopus oocyte expression system and sequence analysis of abnormal restriction enzyme fragments from patients with hereditary myeloperoxidase deficiency will be used to characterize normal and aberrant proenzyme processing. The interplay among the individual projects is extensive at both conceptual and technical levels. Specific receptor-ligand-cytoskeletal interactions lead to oxidase activation events (Project 1 which in turn result in the formation of activated species of oxygen that require in part the secretion of myeloperoxidase for the expression of microbicidal activity. The investigators' laboratories are in favorable proximity to facilitate programmatic approaches and core resource development. Two proposed scientific cores will provide cell preparation and culture support and analytical facilities whereas overall program management will be coordinated through the administrative core. Institutional environment and support are also very favorable including such core facilities as protein structure, molecular biology, ESR spectroscopy, hybridoma, electron microscopy, image analysis, and fluorescence- activated cell sorting. Thus, the Program Project Grant will take maximum advantage of the complementary scientific expertise of the investigators, the proposed core facilities and existing institutional features to attack important problems in neutrophil biology.
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