The role of MyD88 and TLR4 in the LPS‐mimetic activity of Taxol

Taxol can mimic bacterial lipopolysaccharide (LPS) by activating mouse macrophages in a cell cycle‐independent, LPS antagonist‐inhibitable manner. Macrophages from C3H/HeJ mice, which have a spontaneous mutation in Toll‐like receptor 4 (TLR4), are hyporesponsive to both LPS and Taxol, suggesting that LPS and Taxol may share a signaling pathway involving TLR4. To determine whether TLR4 and its interacting adaptor molecule MyD88 are necessary for Taxol's LPS mimetic actions, we examined Taxol responses of primary macrophages from genetically defective mice lacking either TLR4 (C57BL/10ScNCr) or MyD88 (MyD88 knockout). When stimulated with Taxol, macrophages from wild‐type mice responded robustly by secreting both TNF and NO, while macrophages from either TLR4‐deficient C57BL/10ScNCr mice or MyD88 knockout mice produced only minimal amounts of TNF and NO. Taxol‐induced NF‐κB‐driven luciferase activity was reduced after transfection of RAW 264.7 macrophages with a dominant negative version of mouse MyD88. Taxol‐induced microtubule‐associated protein kinase (MAPK) activation and NF‐κB nuclear translocation were absent from TLR4‐null macrophages, but were preserved in MyD88 knockout macrophages with a slight delay in kinetics. Neither Taxol‐induced NF‐κB activation, nor IκB degradation was affected by the presence of phosphatidylinositol 3‐kinaseinhibitors. These results suggest that Taxol and LPS not only share a TLR4/MyD88‐dependent pathway in generating inflammatory mediators, but also share a TLR4‐dependent/MyD88‐independent pathway leading to activation of MAPK and NF‐κB.

[1]  P. Schiff,et al.  Promotion of microtubule assembly in vitro by taxol , 1979, Nature.

[2]  H. Chapman,et al.  Macrophage tumor killing: influence of the local environment. , 1977, Science.

[3]  C. Janeway,et al.  Self-defense: the fruit fly style. , 1998, Proceedings of the National Academy of Sciences of the United States of America.

[4]  P. Moseley.  Heat Shock Proteins and the Inflammatory Response , 1998, Annals of the New York Academy of Sciences.

[5]  A. DeFranco,et al.  Phosphatidylinositol 3‐kinase and mTOR mediate lipopolysaccharide‐stimulated nitric oxide production in macrophages via interferon‐β , 2000, Journal of leukocyte biology.

[6]  R. Ulevitch,et al.  Lipopolysaccharide binding protein and CD14 in LPS dependent macrophage activation. , 1993, Immunobiology.

[7]  David Baltimore,et al.  Inducibility of κ immunoglobulin enhancer-binding protein NF-κB by a posttranslational mechanism , 1986, Cell.

[8]  M. Nishijima,et al.  Cutting Edge: Gln of Mouse MD-2 Is Essential for Species-Specific Lipopolysaccharide Mimetic Action of Taxol , 2000 .

[9]  T. Yoshida,et al.  Mouse Toll-like Receptor 4·MD-2 Complex Mediates Lipopolysaccharide-mimetic Signal Transduction by Taxol* , 2000, The Journal of Biological Chemistry.

[10]  D. Baltimore,et al.  Inducibility of kappa immunoglobulin enhancer-binding protein Nf-kappa B by a posttranslational mechanism. , 1986, Cell.

[11]  P. Ricciardi-Castagnoli,et al.  Physical contact between lipopolysaccharide and toll-like receptor 4 revealed by genetic complementation. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[12]  B. Lemaître,et al.  Drosophila host defense: differential induction of antimicrobial peptide genes after infection by various classes of microorganisms. , 1997, Proceedings of the National Academy of Sciences of the United States of America.

[13]  C. Jefferies,et al.  Rac1 Regulates Interleukin 1-induced Nuclear Factor κB Activation in an Inhibitory Protein κBα-independent Manner by Enhancing the Ability of the p65 Subunit to Transactivate Gene Expression* , 2000, The Journal of Biological Chemistry.

[14]  C. Colaco.  Towards a unified theory of immunity: dendritic cells, stress proteins and antigen capture. , 1998, Cellular and molecular biology.

[15]  P. Feng,et al.  IRAK (Pelle) family member IRAK-2 and MyD88 as proximal mediators of IL-1 signaling. , 1997, Science.

[16]  C. Janeway,et al.  MyD88 is an adaptor protein in the hToll/IL-1 receptor family signaling pathways. , 1998, Molecular cell.

[17]  M. Rothe,et al.  Bacterial Lipopolysaccharide Activates Nuclear Factor-κB through Interleukin-1 Signaling Mediators in Cultured Human Dermal Endothelial Cells and Mononuclear Phagocytes* , 1999, The Journal of Biological Chemistry.

[18]  T. Hamilton,et al.  The cell biology of macrophage activation. , 1984, Annual review of immunology.

[19]  W. Mellado,et al.  Taxol: Mechanisms of Action and Resistance a , 1986, Journal of the National Cancer Institute. Monographs.

[20]  J. Manfredi,et al.  Taxol binds to cellular microtubules , 1982, The Journal of cell biology.

[21]  L. Larivière,et al.  Endotoxin-tolerant Mice Have Mutations in Toll-like Receptor 4 (Tlr4) , 1999, The Journal of experimental medicine.

[22]  S. Akira,et al.  Targeted disruption of the MyD88 gene results in loss of IL-1- and IL-18-mediated function. , 1998, Immunity.

[23]  Fuchs Da,et al.  Cytologic evidence that taxol, an antineoplastic agent from Taxus brevifolia, acts as a mitotic spindle poison. , 1978 .

[24]  P. Godowski,et al.  Toll-like receptor 2–mediated NF-κB activation requires a Rac1-dependent pathway , 2000, Nature Immunology.

[25]  D. Klinman,et al.  Contribution of CpG motifs to the immunogenicity of DNA vaccines. , 1997, Journal of immunology.

[26]  S. Saccani,et al.  The Human Toll Signaling Pathway: Divergence of Nuclear Factor κB and JNK/SAPK Activation Upstream of Tumor Necrosis Factor Receptor–associated Factor 6 (TRAF6) , 1998, The Journal of experimental medicine.

[27]  S M Teutsch,et al.  Trends in infectious diseases mortality in the United States. , 1996, JAMA.

[28]  B. Monks,et al.  Toll-like receptor 4 imparts ligand-specific recognition of bacterial lipopolysaccharide. , 2000, The Journal of clinical investigation.

[29]  Z. Cao,et al.  MyD88: an adapter that recruits IRAK to the IL-1 receptor complex. , 1997, Immunity.

[30]  M. McCarron,et al.  Septic Shock , 1964, International anesthesiology clinics.

[31]  J. Berman,et al.  Relationship between the structure of taxol and other taxanes on induction of tumor necrosis factor-alpha gene expression and cytotoxicity. , 1994, Cancer research.

[32]  K. Anderson,et al.  The Toll gene of drosophila, required for dorsal-ventral embryonic polarity, appears to encode a transmembrane protein , 1988, Cell.

[33]  L. Wahl,et al.  Macrophage sensitivity to endotoxin: genetic control by a single codominant gene. , 1978, Journal of immunology.

[34]  C. Nathan,et al.  Shared actions of endotoxin and taxol on TNF receptors and TNF release. , 1990, Science.

[35]  P. Ricciardi-Castagnoli,et al.  Defective LPS signaling in C3H/HeJ and C57BL/10ScCr mice: mutations in Tlr4 gene. , 1998, Science.

[36]  R. Johnson,et al.  Cytologic evidence that taxol, an antineoplastic agent from Taxus brevifolia, acts as a mitotic spindle poison. , 1978, Cancer treatment reports.

[37]  L. Tong,et al.  Structural basis for signal transduction by the Toll/interleukin-1 receptor domains , 2000, Nature.

[38]  S. Akira,et al.  Cellular responses to bacterial cell wall components are mediated through MyD88-dependent signaling cascades. , 2000, International immunology.

[39]  H. Petty,et al.  Cutting Edge: Lipopolysaccharide Induces Physical Proximity Between CD14 and Toll-Like Receptor 4 (TLR4) Prior to Nuclear Translocation of NF-κB1 , 2000, The Journal of Immunology.

[40]  A. Aderem,et al.  Toll-like receptor-2 mediates mycobacteria-induced proinflammatory signaling in macrophages. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[41]  L. Larivière,et al.  Cutting edge: functional characterization of the effect of the C3H/HeJ defect in mice that lack an Lpsn gene: in vivo evidence for a dominant negative mutation. , 1999, Journal of immunology.

[42]  F. Martinon,et al.  MyD88, an Adapter Protein Involved in Interleukin-1 Signaling* , 1998, The Journal of Biological Chemistry.

[43]  S. Akira,et al.  Cutting Edge: Preferentially the R-Stereoisomer of the Mycoplasmal Lipopeptide Macrophage-Activating Lipopeptide-2 Activates Immune Cells Through a Toll-Like Receptor 2- and MyD88-Dependent Signaling Pathway1 , 2000, The Journal of Immunology.

[44]  S. Akira,et al.  Unresponsiveness of MyD88-deficient mice to endotoxin. , 1999, Immunity.

[45]  D. Morrison,et al.  Endotoxins and disease mechanisms. , 1987, Annual review of medicine.

[46]  M. Rothe,et al.  Human Toll-like Receptor 2 Confers Responsiveness to Bacterial Lipopolysaccharide , 1998, The Journal of experimental medicine.

[47]  J. Watson,et al.  The genetic mapping of a defective LPS response gene in C3H/HeJ mice. , 1978, Journal of immunology.

[48]  A. McPhail,et al.  Plant antitumor agents. VI. The isolation and structure of taxol, a novel antileukemic and antitumor agent from Taxus brevifolia. , 1971, Journal of the American Chemical Society.

[49]  C. Manthey,et al.  Lipopolysaccharide antagonists block taxol-induced signaling in murine macrophages , 1993, The Journal of experimental medicine.

[50]  Michael W. Peterson,et al.  Lipopolysaccharide Activates Akt in Human Alveolar Macrophages Resulting in Nuclear Accumulation and Transcriptional Activity of β-Catenin1 , 2001, The Journal of Immunology.

[51]  C. Nathan,et al.  Secretory products of macrophages. , 1987, The Journal of clinical investigation.

[52]  T. Kirikae,et al.  CD14 is not involved in Rhodobacter sphaeroides diphosphoryl lipid A inhibition of tumor necrosis factor alpha and nitric oxide induction by taxol in murine macrophages , 1995, Infection and immunity.

[53]  C. Manthey,et al.  Taxol provides a second signal for murine macrophage tumoricidal activity. , 1994, Journal of immunology.

[54]  M. Nishijima,et al.  Cutting Edge: Gln22 of Mouse MD-2 Is Essential for Species-Specific Lipopolysaccharide Mimetic Action of Taxol1 , 2001, The Journal of Immunology.

[55]  N. Rosen,et al.  Heat shock protein 90 mediates macrophage activation by Taxol and bacterial lipopolysaccharide. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[56]  S. Akira,et al.  Cutting edge: Toll-like receptor 4 (TLR4)-deficient mice are hyporesponsive to lipopolysaccharide: evidence for TLR4 as the Lps gene product. , 1999, Journal of immunology.

[57]  B. Beutler,et al.  Analysis of Tlr4-mediated LPS signal transduction in macrophages by mutational modification of the receptor. , 1999, Blood cells, molecules & diseases.

[58]  D. Morrison,et al.  Macrophage stimulation by bacterial lipopolysaccharides. II. Evidence for differentiation signals delivered by lipid A and by a protein rich fraction of lipopolysaccharides , 1978, The Journal of experimental medicine.

[59]  S. Rhee,et al.  Murine TOLL-like Receptor 4 Confers Lipopolysaccharide Responsiveness as Determined by Activation of NFκB and Expression of the Inducible Cyclooxygenase* , 2000, The Journal of Biological Chemistry.

[60]  H. Kolb,et al.  Cutting Edge: Heat Shock Protein 60 Is a Putative Endogenous Ligand of the Toll-Like Receptor-4 Complex1 , 2000, The Journal of Immunology.

[61]  R. Johnston,et al.  Increased production of superoxide anion by macrophages exposed in vitro to muramyl dipeptide or lipopolysaccharide , 1980, The Journal of experimental medicine.

[62]  F. Ognibene,et al.  Septic shock in humans. Advances in the understanding of pathogenesis, cardiovascular dysfunction, and therapy. , 1990, Annals of internal medicine.

[63]  C. Nathan,et al.  Taxol shares the ability of bacterial lipopolysaccharide to induce tyrosine phosphorylation of microtubule-associated protein kinase. , 1993, Journal of Immunology.

[64]  R. Ulevitch,et al.  Isolation of a lipopolysaccharide-binding acute phase reactant from rabbit serum , 1986, The Journal of experimental medicine.

[65]  H. Petty,et al.  Lipopolysaccharide induces physical proximity between CD14 and toll-like receptor 4 (TLR4) prior to nuclear translocation of NF-kappa B. , 2000, Journal of immunology.

[66]  R. Ulevitch,et al.  CD14, a receptor for complexes of lipopolysaccharide (LPS) and LPS binding protein. , 1990, Science.

[67]  G. Stark,et al.  Activation of Phosphatidylinositol 3-Kinase in Response to Interleukin-1 Leads to Phosphorylation and Activation of the NF-κB p65/RelA Subunit , 1999, Molecular and Cellular Biology.

[68]  A. Ding,et al.  Activation of NF-kappa B in murine macrophages by taxol. , 1995, Cancer biochemistry biophysics.

[69]  B. M. Sultzer.  Genetic Control of Leucocyte Responses to Endotoxin , 1968, Nature.

[70]  C. Janeway,et al.  A human homologue of the Drosophila Toll protein signals activation of adaptive immunity , 1997, Nature.

[71]  Zhonghe Zhou,et al.  The smallest known non-avian theropod dinosaur , 2000, Nature.