TY - JOUR
T1 - Essential roles of CD14 and lipopolysaccharide-binding protein for activation of toll-like receptor (TLR)2 as well as TLR4
T2 - Reconstitution of TLR2- and TLR4-activation by distinguishable ligands in LPS preparations
AU - Muta, Tatsushi
AU - Takeshige, Koichiro
PY - 2001
Y1 - 2001
N2 - Although genetic studies have revealed a critical role for the toll-like receptor (TLR) 4 in the biological response to lipopolysaccharide (LPS), the activities of ectopically expressed TLR4 and TLR2 are controversial. We have found that under appropriate transfection conditions, both TLR2 and TLR4 mediate LPS-induced NF-κB activation in human embryonic kidney 293 cells. The reconstitution systems we established here allow direct biochemical characterization and comparison of activation of each receptor. TLR4 is ≈ 100-fold more sensitive to LPS than TLR2. In contrast to the response to commercial LPS preparations, TLR2 is unresponsive to repurified LPS or synthetic lipid A, indicating the requirement for an additional molecule(s). On the other hand, a lipid A-neutralizing reagent, polymyxin B, blocks the ability of the LPS preparation to stimulate both receptors, suggesting that lipid A is also involved in the activation of TLR2. Mutant TLRs harboring a point mutation in the cytoplasmic domain is inactive in transducing the signal upon stimulation, and act as dominant-negative mutants specifically inhibiting the activation of corresponding type of the receptor but not the other type. Thus, the two receptors are independently activated by distinguishable ligands. Nevertheless, the responses of both TLRs to the LPS preparation are strongly dependent on serum and CD14 and LPS-binding protein are essential for the activation of both of the two receptors. Supporting its functional significance, both receptors are found to associate with CD14.
AB - Although genetic studies have revealed a critical role for the toll-like receptor (TLR) 4 in the biological response to lipopolysaccharide (LPS), the activities of ectopically expressed TLR4 and TLR2 are controversial. We have found that under appropriate transfection conditions, both TLR2 and TLR4 mediate LPS-induced NF-κB activation in human embryonic kidney 293 cells. The reconstitution systems we established here allow direct biochemical characterization and comparison of activation of each receptor. TLR4 is ≈ 100-fold more sensitive to LPS than TLR2. In contrast to the response to commercial LPS preparations, TLR2 is unresponsive to repurified LPS or synthetic lipid A, indicating the requirement for an additional molecule(s). On the other hand, a lipid A-neutralizing reagent, polymyxin B, blocks the ability of the LPS preparation to stimulate both receptors, suggesting that lipid A is also involved in the activation of TLR2. Mutant TLRs harboring a point mutation in the cytoplasmic domain is inactive in transducing the signal upon stimulation, and act as dominant-negative mutants specifically inhibiting the activation of corresponding type of the receptor but not the other type. Thus, the two receptors are independently activated by distinguishable ligands. Nevertheless, the responses of both TLRs to the LPS preparation are strongly dependent on serum and CD14 and LPS-binding protein are essential for the activation of both of the two receptors. Supporting its functional significance, both receptors are found to associate with CD14.
KW - CD14
KW - Innate immunity
KW - Lipopolysaccharide
KW - Lipopolysaccharide-binding protein
KW - Toll-like receptor
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U2 - 10.1046/j.1432-1327.2001.02385.x
DO - 10.1046/j.1432-1327.2001.02385.x
M3 - Article
C2 - 11502220
AN - SCOPUS:0034819316
VL - 268
SP - 4580
EP - 4589
JO - FEBS Journal
JF - FEBS Journal
SN - 1742-464X
IS - 16
ER -