MOLECULAR PATHOLOGY OF PNEUMOCYSTIS PNEUMONIA

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DESCRIPTION: Alveolar macrophages from Pneumocystis carinii-infected hosts are defective in phagocytosis, and the expression of the transcription factor GATA-2 in these cells is severely down-regulated. Introduction of a GATA-2-specific antisense oligonuceotide into alveolar macrophages from normal uninfected animals also resulted in a decrease in the phagocytic activity of these cells. In this proposed study, experiments will be performed to further investigate the role of GATA-2 in alveolar macrophage phagocytosis. A GATA-2 expression vector will be introduced into alveolar macrophages from P. carinii-infected hosts to investigate whether GATA-2 over-expression could correct the defect. To further understand the involvement of GATA-2 in P. carinii pathogenesis, genes that are regulated by GATA-2 in monocytes will be identified. DNA microarrays will be probed with labeled cDNA from wild type and GATA-2 knockout monocytes. The hybridized microarrays will then be analyzed to determine which genes are regulated by GATA-2. Experiments will also be performed to determine whether genes such as those encoding MMR, MIP-1alpha, MCP-1, IL6 and TNF-alpha that are associated with functions of alveolar macrophages are regulated by GATA-2. The mechanisms by which P. carinii causes down regulation of GATA-2 in alveolar macrophages will be explored. Normal alveolar macrophages will be incubated directly or indirectly with live or dead P. carinii organisms to determine whether a P. carinii protein is responsible for down regulation of GATA-2 transcription leading to a reduction in the phagocytic activity of alveolar macrophages. Proteins such as vitronectin, fibronectin, surfactant proteins A and D, and P. carinii major surface glycoprotein, which are known to interact with alveolar macrophages during P. carinii infection, will also be examined. These proteins will be incubated either alone or in combination with normal macrophages to determine whether they have any effect on GATA-2 down regulation. Different fractions of bronchoalveolar lavage fluids from P. carinii-infected lung will also be tested. Any P. carinii or host protein that is found to have the ability to down regulate GATA-2 expression will be identified by sequencing a portion of the protein.