This application requests five years of additional support for a Program Project that is currently in year 30. The theme for the program is cellular and molecular biology of the lung surfactant system. The hypothesis is that the surfactant cell cycle is regulated through control of lipid and protein synthesis, packaging in secretory organelles, regulated and constitutive secretion, cellular reuptake, and intracellular processing of endocytosed lipid and protein components. This hypothesis will be studied through 5 continuing projects. Project 1 will evaluate the interaction of SP-A with alveolar type II cells with emphasis on receptor-mediated uptake of SP-A and DPPC and the pathways for routing of SP-A to lamellar bodies. This project provides novel information on P63 as a biologically-relevant SP-A receptor. Project 2 will focus on molecular and functional studies of ABCA3, a lamellar body membrane protein that was originally described by this program and that is now known to be essential for surfactant processing. Project 3 has been discontinued as the project leader has relocated to another university. Project 4 has as its focus the regulation of gene expression for surfactant-related genes with emphasis on genes regulated by TTF. This project has developed an in vitro hormonally responsive model system for study of type II cell differentiation. The project will focus on regulation of genes for the lamellar body relevant proteins, ABCA3 and LAMPS. Project 5 has as its focus the intracellular processing of SP-C using a variety of unique immunological reagents to identify subcellular sites for conversion of the primary translation product to the mature protein. This project has identified a novel pathway to aggresome formation due to misprocessing of SP-C and will evaluate the subsequent mechanisms for cell injury that result in a lung fibrosis syndrome. Project 6 will continue study of perioxiredoxin 6, a novel lysosomal type phospholipase A2 that was identified by this program. This PLA2 is responsible for DPPC degradation following internalization by type II cells and is also required for PC synthesis by the reacylation pathway. The 5 projects are supported by a molecular core that will provide adenovector constructs and recombinant proteins, an imaging and morphology core which provides support for confocal fluorescence imaging, electron microscopy including immunogold localization, and morphometric analysis, a cell culture core that provides primary cultures of human, rat and mouse granular pneumocytes, and an administrative core. These projects and cores form a cohesive unit with extensive interactions between investigators and projects, all focused on the common aim to understand the cellular and molecular biology of the lung surfactant system with the goal of enhancing the treatment for surfactant-related diseases.
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