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Extracellular matrix proteoglycans regulate toll-like receptors 4 and 9

细胞外基质蛋白聚糖调节 Toll 样受体 4 和 9

基本信息

批准号：
10563132
负责人：
Shukti Chakravarti
金额：
$ 41万
依托单位：
NEW YORK UNIVERSITY SCHOOL OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-01-01 至 2024-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10563132
关键词：
Address Anti-Bacterial Agents Antibiotic Resistance Autoimmunity Bacterial DNA Bacterial Infections Binding Binding Sites C-terminal CAV1 gene CD14 gene Cell Wall Cell membrane Cell surface Cells Chronic Collagen Fibril Cornea DNA Data Dendritic Cells Early Endosome Endocytosis Endosomes Extracellular Matrix Extracellular Matrix Proteins Eye Future Homeostasis Host Defense Infection Inflammation Inflammatory Injections Interferon Type I Keratitis Knockout Mice Leucine-Rich Repeat Ligands Lipopolysaccharides Location Macrophage Mediating Membrane Microdomains Molecular Mus Mutate N-terminal Nuclear Pathway interactions Peritoneal Macrophages Phenylalanine Play Production Property Proteins Proteoglycan Pseudomonas Infections Pseudomonas aeruginosa Recombinants Regulation Role Signal Pathway Signal Transduction Site Sterility Structure TLR4 gene TLR9 gene Testing Therapeutic Intervention Tissues Toll-like receptors Tyrosine Viral Vision Wild Type Mouse Work caveolin 1 conjunctiva corneal regeneration cytokine healing in vivo knowledge base lumican monocyte novel pathogen permissiveness prototype receptor response therapeutic target therapy development trafficking translational impact translational potential treatment strategy viral DNA

项目摘要

Pseudomonas (P.) aeruginosa bacterial infections cause sight-threatening inflammation of the cornea. With antibiotic resistance on the rise, additional treatment strategies are needed. To address this gap, the current proposal focuses on novel interactions of pathogen recognition toll-like receptors (TLRs) with lumican (Lum), an extracellular matrix protein of the cornea. The TLR-4 and TLR-9 receptors recognize P. aeruginosa cell wall-derived lipopolysaccharides (LPS) and nuclear DNA, respectively, to induce pro-inflammatory cytokines and type I interferons. LPS recognition by TLR4 at the cell surface of macrophages and dendritic cells (DCs) leads to MyD88-dependent pro-inflammatory cytokine production, while endosomal TLR4 stimulates an alternative pathway that additionally produces type I interferons. Endolysosomal TLR9, on the other hand, stimulated by bacterial DNA or the synthetic TLR9 ligand CpGDNA, leads to the production of both pro- inflammatory cytokines and type I interferons. Interestingly, Lum has opposing effects on TLR4 and TLR9; it promotes TLR4 but suppresses TLR9 responses. We demonstrated that a tyrosine at the N-terminal end (Y20) of Lum binds with the TLR-adaptor CD14 to promote cell surface TLR4 signals. Our preliminary data suggest a potential caveolin-1 binding site at the C-terminal end (F228) of Lum that may be involved in caveolar trafficking of TLRs. Our central hypothesis is that through binding CD14 and CAV1 Lum regulates TLR4 and TLR9 locations within cells to promote TLR4 but restrict TLR9 signals, and this will impact pro-inflammatory cytokine and type I interferon inductions to modulate inflammation and regain of tissue homeostasis in keratitis. In the following aims we will test our hypothesis using wild type mice, Lum-null mice, and macrophages and dendritic cells from these mice. Aim 1) determine if Lum regulates TLR4 location and degradation, with CD14-binding preferentially promoting the cell surface while CAV1 the endosomal TLR4 pathway, Aim 2) test if Lum increases the inactive TLR9 pool in the cell surface and endosomal compartments to suppress its signals, and Aim 3) determine the course of sterile keratitis mediated by LPS or CpGDNA and P. aeruginosa mediated infectious keratitis in Lum- deficient and wild type mice and the effects of subconjunctival injections of mutated recombinant Lum with loss of CD14 or Cav1 binding activity. Type I interferons have broad antibacterial and tissue protective roles in addition to their well-known antiviral properties. Thus, a provocative translational potential is that by manipulating the CD14 and the CAV1 binding properties of Lum, it will be possible to regulate the pro- inflammatory cytokine and type I interferon induction pathways separately to resolve inflammation and expedite corneal healing keratitis. Our study will develop a strong knowledge base on the role played by the ECM in infection and inflammation for developing novel molecular therapeutic interventions in the future.

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