Regulation of cellular growth is mediated by a vast array of factors. Several proteins that regulate growth have been identified by their association with the aberrantly controlled growth of tumor cells. Macrophages play a critical role in immune recognition, and their uncontrolled growth results in myeloid leukemias. In atherosclerosis, macrophage adherence to arterial endothelium is one of the earliest events in the disease. Proliferation of macrophages is controlled at many levels. A specific regulator of macrophage growth is the intercellular cytokine, macrophage colony stimulating factor (M-CSF or CSF-1), that binds to the extracellular portion of a macrophage-specific cell surface receptor, the integral membrane protein Fms. Our recent results have shown that another growth regulator, the macrophage-specific transcription factor PU.1, enhances the CSF-1 stimulated proliferation of macrophages. Transcription factors directly control gene expression by binding to DNA and regulating RNA expression. The product of the PU.1 or spi-1 gene is a sequence-specific DNA binding phosphoprotein expressed in the nuclei of macrophages and B-cells. The PU.1 gene is a member of the ets oncogene family, and aberrant expression of the PU.l/spi-1 gene is associated with erythroid tumors in mice. PU.1 transactivates genes whose promoter-- enhancer regions contain the PU.1 DNA binding sequences. Phosphorylation of PU.1 influences its ability to transactivate responsive genes without affecting its binding to its DNA target sequence. In addition, phosphorylation affects the ability of PU.1 to stimulate macrophage growth. The long-term goal of this study is to understand growth regulation at the level of gene expression.
The specific aims of the proposal will focus on 1) delineating the sites of phosphorylation in the PU.l/spi-1 protein; 2) the effect of regulators of macrophage proliferation on phosphorylation of PU.1; 3) the role of the CSF-1 receptor in regulating activity of the PU.1 protein; and 4) the role of the PU.1 protein in regulating CSF-1-dependent macrophage proliferation.
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