We have found that the level of airway epithelial-leukocyte adhesion is controlled at least in part by the amount of intercellular adhesion molecule-1 (CAM-1) on the epithelial cell surface. Thus, determining the mechanisms that regulate ICAM-1 expression on airway epithelial cells provides an avenue for understanding the development of leukocyte influx and trafficking into airway tissues and for identifying pharmacologic targets in airway disease. Our results indicate that there is only little ICAM-1 (or leukocyte adhesion) in cultured airway epithelial cells under basal conditions and that the level of ICAM-1 (and leukocyte adhesion) is especially sensitive to stimulation by interferon-gamma (IFN-gamma). We propose that the selective IFN-gamma-responsiveness of ICAM-1 levels on epithelial cells may allow for selective localization of leukocytes in the epithelium during inflammatory conditions with augmented IFN-gamma production and that an overexuberant TH1-type inflammatory response may even lead to excessive local activation of leukocytes and consequent epithelial damage. In studies aimed at determining the biochemical basis for the IFN-gamma effect on ICAM-1 expression, we have found that the effect is regulated at a transcriptional level, and an analysis of the ICAM-1 gene promoter region has led to the identification of a cis-acting interferon-response element (IRE) and a trans-acting IRE-binding complex (IRE-BC) that mediate IFN-gamma control of ICAM-1 gene transcription. The proposed studies aim at more precisely identifying and characterizing the DNA elements and then the corresponding DNA-binding proteins that mediate the selective response of the ICAM-1 gene to IFN-gamma in epithelial cells. The principal cell system to be employed for this project is primary culture human airway mucosal epithelial cells, but pilot experiments may also be performed using transformed lung epithelial cell lines. Distinct features of ICAM-1 gene IRE behavior in airway epithelial cells will be defined by comparison to: (i) the behavior of the same element in other cell types (especially vascular endothelial cells) in which the element is no longer IFN-responsive; and Iii) the behavior of the two other closely- matched IFN-gamma-responsive elements found in two other immune-response genes. These comparisons are aimed at identifying determinants for cell- specific and gene-specific regulation of ICAM-1.
Specific aims are to: I. Precisely identify cis-acting regulatory elements)s) in the ICAM-1 gene IRE that are critical for IFN-gamma-responsiveness in airway epithelial cells.
This aim will be accomplished by analyzing the activity of ICAM-1 gene/reporter gene constructs in DNA-transfection experiments and by DNase I protection mapping (footprinting). II. Identify and characterize specific trans-acting protein(s) that bind to the ICAM-1 gene IRE in airway epithelial cells.
This aim will be accomplished by analyzing the characteristics of nuclear proteins (and cytosolic precursors) from IFN-gamma stimulated versus unstimulated cells in DNA-protein binding experiments.
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| Nakajima, S; Roswit, W T; Look, D C et al. (1995) A hierarchy for integrin expression and adhesiveness among T cell subsets that is linked to TCR gene usage and emphasizes V delta 1+ gamma delta T cell adherence and tissue retention. J Immunol 155:1117-31 |
