GPCR Kinase-2 in TNFalpha Signaling in Macrophages

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Macrophages are important immune cells involved in the pathogenesis of many inflammatory diseases, including atherosclerosis and are one of the major immune cells in the atherosclerotic lesions. In addition to becoming foam cells upon uptake of modified lipoproteins in the vessel walls, macrophages also mediate the inflammatory responses that are associated with atherosclerotic plaque formation by producing cytokines, chemokines and growth factors. These agents further influence chemoattraction of more monocytes and other immune cells as well as profoundly affect the functions of smooth muscle cells and endothelial cells that are in the vicinity. Therefore, macrophages play an essential role in all stages of atherogenesis including fatty streaks to advanced plaques and eventually to plaque rupture. Thus, understanding the biochemical mechanisms by which macrophage biology is regulated is highly critical. Preliminary data demonstrates that macrophage biology is regulated by G-protein coupled receptor kinase-2 (GRK2). GRK2 is a serine/threonine kinase that was discovered for its role in the phosphorylation of 􀀂-adrenergic receptors. Evidence indicates that tumor necrosis factor-􀀁(TNF􀀁)-induced NF􀀃B signaling, and the consequent macrophage biology are critically regulated by GRK2, via interaction with, and potentially phosphorylation of the inhibitor of NF􀀃B signaling namely I􀀃B􀀁. These results suggest a novel and significant role for this kinase in macrophage biology and therefore in the pathogenesis of atherosclerosis. The objective of this proposal is to further expand on our preliminary findings and examine the mechanisms by which GRK2 regulates macrophage biology and importantly, also test if, loss of GRK2 affects the pathophysiology of atherosclerosis. The overall hypothesis is that GRK2 interaction with and phosphorylation of I􀀃B􀀁regulates TNF􀀁-induced NF􀀃B signaling, inflammatory mediator production, and macrophage survival/apoptosis and therefore, GRK2 plays a crucial role in the pathogenesis of atherosclerosis. To test this hypothesis we will examine the following specific aims: 1. Describe the mechanisms by which GRK-2 regulates TNF􀀁-induced NF􀀃B pathway in macrophages. 2. Examine the functional relevance of GRK2 in TNF􀀁signaling in macrophages. 3. Determine whether myeloid cell-specific loss of GRK2 affects the pathogenesis of atherosclerosis in LDLR knockout mice. Taken together, our studies should provide important mechanistic insight into TNF􀀁signaling in macrophages as well as identify potential therapeutic targets in the treatment of chronic inflammatory diseases