title: MD-2 is required for disulfide HMGB1-dependent TLR4 signaling.
abstract: Innate immune receptors for pathogen- and damage-associated molecular patterns (PAMPs and DAMPs) orchestrate inflammatory responses to infection and injury. Secreted by activated immune cells or passively released by damaged cells, HMGB1 is subjected to redox modification that distinctly influences its extracellular functions. Previously, it was unknown how the TLR4 signalosome distinguished between HMGB1 isoforms. Here we demonstrate that the extracellular TLR4 adaptor, myeloid differentiation factor 2 (MD-2), binds specifically to the cytokine-inducing disulfide isoform of HMGB1, to the exclusion of other isoforms. Using MD-2-deficient mice, as well as MD-2 silencing in macrophages, we show a requirement for HMGB1-dependent TLR4 signaling. By screening HMGB1 peptide libraries, we identified a tetramer (FSSE, designated P5779) as a specific MD-2 antagonist preventing MD-2-HMGB1 interaction and TLR4 signaling. P5779 does not interfere with lipopolysaccharide-induced cytokine/chemokine production, thus preserving PAMP-mediated TLR4-MD-2 responses. Furthermore, P5779 can protect mice against hepatic ischemia/reperfusion injury, chemical toxicity, and sepsis. These findings reveal a novel mechanism by which innate systems selectively recognize specific HMGB1 isoforms. The results may direct toward strategies aimed at attenuating DAMP-mediated inflammation while preserving antimicrobial immune responsiveness.
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030 kjtracey@nshs.edu hyang@nshs.edu.
authors.forename: Huan
authors.initials: H
authors.lastname: Yang
authors.affiliation: Department of Emergency Medicine, North Shore University Hospital, Manhasset, NY 11030.
authors.forename: Haichao
authors.initials: H
authors.lastname: Wang
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030.
authors.forename: Zhongliang
authors.initials: Z
authors.lastname: Ju
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030.
authors.forename: Ahmed A
authors.initials: AA
authors.lastname: Ragab
authors.affiliation: Department of Medicine and Department of Women's and Children's Health, Karolinska Institutet, Karolinska University Hospital, SE-171 77 Stockholm, Sweden Department of Medicine and Department of Women's and Children's Health, Karolinska Institutet, Karolinska University Hospital, SE-171 77 Stockholm, Sweden.
authors.forename: Peter
authors.initials: P
authors.lastname: Lundbäck
authors.affiliation: Department of Emergency Medicine, North Shore University Hospital, Manhasset, NY 11030.
authors.forename: Wei
authors.initials: W
authors.lastname: Long
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030.
authors.forename: Sergio I
authors.initials: SI
authors.lastname: Valdes-Ferrer
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030.
authors.forename: Mingzhu
authors.initials: M
authors.lastname: He
authors.affiliation: Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213.
authors.forename: John P
authors.initials: JP
authors.lastname: Pribis
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030.
authors.forename: Jianhua
authors.initials: J
authors.lastname: Li
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030.
authors.forename: Ben
authors.initials: B
authors.lastname: Lu
authors.affiliation: Department of Anesthesiology, The University of Texas Medical Branch, Galveston, TX 77555.
authors.forename: Domokos
authors.initials: D
authors.lastname: Gero
authors.affiliation: Department of Anesthesiology, The University of Texas Medical Branch, Galveston, TX 77555.
authors.forename: Csaba
authors.initials: C
authors.lastname: Szabo
authors.affiliation: Medical Research Council Centre for Drug Safety Science, Department of Molecular and Clinical Pharmacology, University of Liverpool, Liverpool L69 3BX, England, UK.
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Antoine
authors.affiliation: Department of Medicine and Department of Women's and Children's Health, Karolinska Institutet, Karolinska University Hospital, SE-171 77 Stockholm, Sweden Department of Medicine and Department of Women's and Children's Health, Karolinska Institutet, Karolinska University Hospital, SE-171 77 Stockholm, Sweden.
authors.forename: Helena E
authors.initials: HE
authors.lastname: Harris
authors.affiliation: Division of Infectious Diseases and Immunology, University of Massachusetts Medical School, Worcester, MA 01655.
authors.forename: Doug T
authors.initials: DT
authors.lastname: Golenbock
authors.affiliation: Division of Infectious Diseases and Immunology, University of Massachusetts Medical School, Worcester, MA 01655.
authors.forename: Jianmin
authors.initials: J
authors.lastname: Meng
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030.
authors.forename: Jesse
authors.initials: J
authors.lastname: Roth
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030.
authors.forename: Sangeeta S
authors.initials: SS
authors.lastname: Chavan
authors.affiliation: Department of Medicine and Department of Women's and Children's Health, Karolinska Institutet, Karolinska University Hospital, SE-171 77 Stockholm, Sweden Department of Medicine and Department of Women's and Children's Health, Karolinska Institutet, Karolinska University Hospital, SE-171 77 Stockholm, Sweden.
authors.forename: Ulf
authors.initials: U
authors.lastname: Andersson
authors.affiliation: Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213.
authors.forename: Timothy R
authors.initials: TR
authors.lastname: Billiar
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030 kjtracey@nshs.edu hyang@nshs.edu.
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Tracey
authors.affiliation: Department of Biomedical Science and Department of Medicinal Chemistry, The Feinstein Institute for Medical Research, Manhasset, NY 11030.
authors.forename: Yousef
authors.initials: Y
authors.lastname: Al-Abed
mesh_terms: D000082:Acetaminophen; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D056486:Chemical and Drug Induced Liver Injury / Q000097:blood / Q000209:etiology / Q000378:metabolism; D016207:Cytokines / Q000097:blood / Q000494:pharmacology; D004220:Disulfides / Q000378:metabolism; D024243:HMGB1 Protein / Q000378:metabolism* / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D051218:Lymphocyte Antigen 96 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D008297:Male; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008958:Models, Molecular; D010455:Peptides / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D011485:Protein Binding / Q000187:drug effects; D017434:Protein Structure, Tertiary; D034622:RNA Interference; D015427:Reperfusion Injury / Q000097:blood / Q000378:metabolism; D015398:Signal Transduction; D016019:Survival Analysis; D051197:Toll-Like Receptor 4 / Q000378:metabolism*
chemical_list: D016207:Cytokines; D004220:Disulfides; D024243:HMGB1 Protein; D008070:Lipopolysaccharides; D051218:Lymphocyte Antigen 96; D010455:Peptides; D051197:Toll-Like Receptor 4; D000082:Acetaminophen