Dok-1 and Dok-2 are negative regulators of lipopolysaccharide-induced signaling.
Endotoxin, a bacterial lipopolysaccharide (LPS), causes fatal septic shock via Toll-like receptor (TLR)4 on effector cells of innate immunity like macrophages, where it activates nuclear factor kappaB (NF-kappaB) and mitogen-activated protein (MAP) kinases to induce proinflammatory cytokines such as tumor necrosis factor (TNF)-alpha. Dok-1 and Dok-2 are adaptor proteins that negatively regulate Ras-Erk signaling downstream of protein tyrosine kinases (PTKs). Here, we demonstrate that LPS rapidly induced the tyrosine phosphorylation and adaptor function of these proteins. The stimulation with LPS of macrophages from mice lacking Dok-1 or Dok-2 induced elevated Erk activation, but not the other MAP kinases or NF-kappaB, resulting in hyperproduction of TNF-alpha and nitric oxide. Furthermore, the mutant mice showed hyperproduction of TNF-alpha and hypersensitivity to LPS. However, macrophages from these mutant mice reacted normally to other pathogenic molecules, CpG oligodeoxynucleotides, poly(I:C) ribonucleotides, or Pam3CSK4 lipopeptide, which activated cognate TLRs but induced no tyrosine phosphorylation of Dok-1 or Dok-2. Forced expression of either adaptor, but not a mutant having a Tyr/Phe substitution, in macrophages inhibited LPS-induced Erk activation and TNF-alpha production. Thus, Dok-1 and Dok-2 are essential negative regulators downstream of TLR4, implying a novel PTK-dependent pathway in innate immunity.
Shinohara
Hisaaki
H
Department of Cell Regulation, Medical Research Institute, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
Inoue
Akane
A
Toyama-Sorimachi
Noriko
N
Nagai
Yoshinori
Y
Yasuda
Tomoharu
T
Suzuki
Hiromi
H
Horai
Reiko
R
Iwakura
Yoichiro
Y
Yamamoto
Tadashi
T
Karasuyama
Hajime
H
Miyake
Kensuke
K
Yamanashi
Yuji
Y
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D007113:Immunity, Innate / Q000502:physiology; D008070:Lipopolysaccharides / Q000378:metabolism / Q000494:pharmacology*; D008264:Macrophages / Q000187:drug effects* / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009569:Nitric Oxide / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051217:Toll-Like Receptor 9; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D004268:DNA-Binding Proteins; C489561:Dok1 protein, mouse; C498444:Dok2 protein, mouse; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D010750:Phosphoproteins; D016601:RNA-Binding Proteins; D011956:Receptors, Cell Surface; C493486:Tlr2 protein, mouse; C493487:Tlr4 protein, mouse; C495370:Tlr9 protein, mouse; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D051217:Toll-Like Receptor 9; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide; D048049:Extracellular Signal-Regulated MAP Kinases