Characterization of two novel LPS-binding sites in leukocyte integrin betaA domain.
Lipopolysaccharide (LPS), a bacterial endotoxin, triggers deleterious systemic inflammatory responses when released into blood circulation, causing organ dysfunction and death. In response to LPS stimulation, CD14 and toll-like receptor (TLR)-4 elicit inflammatory signaling cascades. Although leukocyte integrins (CD11b/CD18 and CD11c/CD18) were reported to bind LPS and induce NF-kappaB translocation, the evidence on such epitope location remains elusive. The present study aims to delineate the LPS-binding sites on the integrin CD18 antigen and to design peptide(s) as potential prophylactic and/or therapeutic agents to modulate LPS effects in activated Jurkat cells. Epitope mapping analysis using a series of CD18 truncated variants revealed two putative LPS-binding sites within the betaA region (216-248 and 266-318 a.a.), which were further confirmed by point mutation studies. Inhibition assay demonstrated that the CD18-betaA(266-318) peptide could block LPS binding in a dose-dependent manner. Our data also indicated that treatment with the CD18-peptide modulated TNF-alpha mRNA transcription via the NF-kappaB signaling pathway in LPS-activated Jurkat cells. In conclusion, we have identified two novel LPS-binding sites located at the CD18 betaA domain of leukocyte integrin, and the integrin peptide betaA(266-318) is shown to inhibit LPS binding and subsequent inflammatory events, having therapeutic implications to cure gram-negative endotoxemia.
Wong
Kwong-Fai
KF
Department of Surgery, The University of Hong Kong, Jockey Club Clinical Research Center, 21 Sassoon Rd., Pokfulam, Hong Kong.
Luk
John M
JM
Cheng
R Holland
RH
Klickstein
Lloyd B
LB
Fan
Sheung-Tat
ST
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D018821:CD18 Antigens / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D007962:Leukocytes / Q000276:immunology*; D008070:Lipopolysaccharides / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016328:NF-kappa B / Q000378:metabolism; D010455:Peptides / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D017433:Protein Structure, Secondary; D016415:Sequence Alignment; D015398:Signal Transduction / Q000502:physiology; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000378:metabolism
D018821:CD18 Antigens; D008070:Lipopolysaccharides; D016328:NF-kappa B; D010455:Peptides; D014409:Tumor Necrosis Factor-alpha