Characterization and inhibition of higher-order assembly signalling in Toll-like receptor pathways (2016-2018)

Toll-like receptors (TLRs) recognize pathogens and endogenous danger signals and initiate innate immune responses that lead to the production of pro-inflammatory cytokines. TLR-mediated inflammation is associated with a number of pathological states, including infectious, autoimmune, inflammatory, cardiovascular and cancer-related disorders. The signaling depends on TIR (Toll/interleukin-1 receptor) domain-containing adaptor molecules interacting with TIR domains in the cytoplasmic segments of TLRs. We have found that TLR adaptor TIR domains form large assemblies, and hypothesize that they signal through a higher-order assembly signaling mechanism. We propose to apply a cross-disciplinary approach to characterize the structures and interactions of TLR adaptor proteins and their assemblies, test structure-based hypotheses for functional effects in cells, and design inhibitors of these interactions. The objective will be achieved through four specific aims: (1) define the molecular architecture of higher-order assemblies formed by TIR domains; (2) determine the role of cysteines in MAL in TLR signaling; (3) analyze structurally and functionally the SAM domains of SARM; and (4) design inhibitors targeting interactions by TLR adaptors. The outcomes of the proposed research will include an improved understanding of signaling in TLR pathways, identify signaling interfaces as new targets for therapeutic design, and provide inhibitory peptides and small molecules as leads for therapeutic development against chronic inflammatory diseases and related disorders.