Binding of lipopeptide to CD14 induces physical proximity of CD14, TLR2 and TLR1.
Lipoproteins or lipopeptides (LP) are bacterial cell wall components detected by the innate immune system. For LP, it has been shown that TLR2 is the essential receptor in cellular activation. However, molecular mechanisms of LP recognition are not yet clear. We used a FLAG-labeled derivative of the synthetic lipopeptide N-palmitoyl-S-[2,3-bis(palmitoyloxy)-(2R,S)-propyl]-(R)-cysteinyl-seryl-(lysyl)(3)-lysine (Pam(3)CSK(4)) to study the roles of CD14, TLR2 and TLR1 in binding and signaling of LP and their molecular interactions in human cells. The activity of Pam(3)CSK(4)-FLAG was TLR2 dependent, whereas the binding was enabled by CD14, as evaluated by flow cytometry and confocal microscopy. Using FRET and FRAP imaging techniques to study molecular associations, we could show that after Pam(3)CSK(4)-FLAG binding, CD14 and Pam(3)CSK(4)-FLAG associate with TLR2 and TLR1, and TLR2 is targeted to a low-mobility complex. Thus, LP binding to CD14 is the first step in the LP recognition, inducing physical proximity of CD14 and LP with TLR2/TLR1 and formation of the TLR2 signaling complex.
Manukyan
Maria
M
Department of Immunology and Cell Biology, Research Center Borstel, Borstel, Germany.
Triantafilou
Kathy
K
Triantafilou
Martha
M
Mackie
Alan
A
Nilsen
Nadra
N
Espevik
Terje
T
Wiesmüller
Karl-Heinz
KH
Ulmer
Artur J
AJ
Heine
Holger
H
D000818:Animals; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D005434:Flow Cytometry; D036681:Fluorescence Recovery After Photobleaching; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000378:metabolism*; D018950:Lipopolysaccharide Receptors / Q000276:immunology / Q000378:metabolism*; D008074:Lipoproteins / Q000276:immunology / Q000378:metabolism*; D008213:Lymphocyte Activation / Q000276:immunology; D008562:Membrane Glycoproteins / Q000276:immunology / Q000378:metabolism*; D018613:Microscopy, Confocal; D011485:Protein Binding / Q000502:physiology; D011956:Receptors, Cell Surface / Q000276:immunology / Q000378:metabolism*; D051194:Toll-Like Receptor 1; D051195:Toll-Like Receptor 2; D051193:Toll-Like Receptors; D014162:Transfection
D018950:Lipopolysaccharide Receptors; D008074:Lipoproteins; D008562:Membrane Glycoproteins; D011956:Receptors, Cell Surface; C495342:TLR2 protein, human; D051194:Toll-Like Receptor 1; D051195:Toll-Like Receptor 2; D051193:Toll-Like Receptors