Diseases of the circulatory system constitute the leading cause of death in humans. Atherosclerosis involved in such diseases is initiated by attachment of monocytes to the arterial endothelial cells followed by migration through the arterial endothelial layer to the subendothelial space. this process is thought to involve monocyte chemotactic protein-1 (MCP-1) generated by endothelial and other cells in the vascular wall. Monocyte migration into injury sites is also involved in inflammation and wound healing. Understanding of the structure, function and regulation of production of MCP-1 could lead to new ways to deal with major diseases of man. To achieve such long-term goals, we propose the following specific objectives: 1. Determine structural features that mediate cell-type specificity of MCP-1 and the related neutrophil attractant protein (NAP-1/IL-8) using site-directed mutagenesis to produce structurally altered forms, bioassays to determine the activity of the mutants towards monocytes and neutrophils, and 2-D NMR to elucidate structural alterations in the mutants. 2. Clone a cDNA encoding the MCP-1 receptor, either using the likely similarity of this receptor to other chemotactic peptide receptors, or using labeled MCP-1 binding to COS cells expressing the cDNA clone, and attempts to engineer the receptor to elucidate the binding regions that confer specificity towards MCP-1 or IL-8. Knowledge of the structures of MCP-1 receptor, the regions of MCP that bind to the receptor, cloning of receptor and its structure and function will allow us to characterize the modulation of monocyte chemotaxis. This information would be important in designing treatments for atherosclerosis and other diseases that involve monocyte migration.
