Scientific Report 2005

Immunology

Initiation of Inflammation by the Innate Immune System

P.S. Tobias, H.-K. Lee, L.K. Curtiss,* P. Dawson,** T. Kirkland,*** D. Liebler****

* Department of Immunology, Scripps Research
** Department of Cell Biology, Scripps Research
*** University of California, San Diego, California
**** Vanderbilt University, Nashville, Tennessee

We focus on understanding the mechanisms by which cells use the innate immune system to initiate defensive inflammatory responses. First, we seek to understand the structural features of the Toll-like receptors (TLRs) and their allied proteins lipopolysaccharide-binding protein, CD14, MD-2, and CD36, which enable the receptors to bind their ligands. Second, we seek to understand the structural changes by which binding of a microbial ligand to the extracellular domain of the receptor leads to signal transduction across the cell membrane and initiation of intracellular signaling cascades. Third, we seek to understand the involvement of endogenous and exogenous inflammatory stimuli in atherosclerosis.

Ten TLRs are known. For most of these, ligands derived from microorganisms are known; binding to the ligands initiates signaling, leading to expression of inflammatory mediators and other defensive responses. In addition, some of the TLRs that may be involved in sterile inflammatory conditions such as arthritis or atherosclerosis may have endogenous ligands. However, these ligands are not yet clearly identified.

To understand the structural features of ligand-receptor binding, we use 2 approaches. In the traditional mutation approach, amino acid residues in the proteins are mutated, and the proteins are then studied for functional changes. In the second approach, we use cross-linking agents to create covalent attachments of the ligands to the proteins. The proteins are then degraded chemically to determine the site of attachment.

Binding of ligands to TLRs starts an intracellular signaling cascade that results in activation of a number of cellular responses. Prominent hypothesized mechanisms by which ligand binding to TLRs incurs transmembrane signaling are (1) the ligand induces dimerization of receptors and (2) binding of the ligand induces conformational changes in the receptor. Our studies indicate that pairs of TLRs are associated even in the absence of ligand and that the pairs undergo a conformational change upon ligand binding. We are using a variety of approaches to understand the structural basis for associations among the TLRs and their associated intracellular signaling partners.Atherosclerosis is an inflammatory disease of the large arteries. Evidence suggests that inflammatory components derived from microbes can induce progression of atherosclerosis. However, most of the development of atherosclerotic lesions is due to endogenous inflammatory factors. Because the TLR system is so intimately involved with inflammation, we are determining whether the TLRs are involved in atherosclerosis. Our initial data clearly indicate that TLR2, whether activated by endogenous ligands or by exogenous ligands, drives progression of atherosclerosis. For these experiments, we are using mouse models of the disease and mice deficient in individual TLRs.

Publications

Dunzendorfer, S., Lee, H.-K., Soldau, K., Tobias, P.S. TLR4 is the signaling but not the lipopolysaccharide uptake receptor. J. Immunol. 173:1166, 2004.

Dunzendorfer, S., Lee, H.-K., Tobias, P.S. Flow-dependent regulation of endothelial Toll-like receptor 2 expression through inhibition of SP1 activity. Circ. Res. 95:684, 2004.

Tobias, P., Curtiss, L.K. Paying the price for pathogen protection: Toll receptors in atherogenesis. J. Lipid Res. 46:404, 2005.