Visualization and localization of message, protein, or structural change is essential to the central theme of this proposal, the regulation of CFTR function. The function of the imaging core varies from low resolution studies of whole tissues, defining protein expression and cellular pathology, to high resolution light and electron microscopic observations of subcellular passaging and presentation of protein. The Center for Biologic Imaging, in which this core service will be performed, is designed with this function in mind. It is reequipped to perform a continuum of optical methods including all types of light and electron microscopy essential to this program project. Within the scope of this project the principal goals of the core will be various. For example, we will quantify the effects of membrane trafficking proteins (e.g., syntaxin) on the subcellular trafficking of CFTR in time and space. We will examine the relationship of mucin expression to CFTR expression in normal and CF airway cells. We will determine whether AKAPs and their subcellular localizing protein phosphatase beta subunits re targeted to the apical plasma membrane and trafficked in clathrin coated vesicles. To perform these functions we will use a diverse array of microscopy technologies. At the light microscopic level these include: histological, immuno-histological, in situ hybridization, laser confocal, and live cell technologies. At the electron microscopic level we will provide fine structural, and immuno-electron microscopic evaluations of specimens as a natural extension of the light microscopic analyses when needed. Furthermore, our considerable experience in computerized image processing and morphometry will allow quantitative analysis of observed phenomena to corroborate earlier, possibly quite subtle qualitative changes. This core will be used extensively by all projects, through the imaging tools used will vary from project to project.
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