Funiculosin variants and phosphorylated derivatives promote innate immune responses via the Toll-like receptor 4/myeloid differentiation factor-2 complex

Naoki Okamoto, Keisuke Mizote, Hiroe Honda, Akinori Saeki, Yasuharu Watanabe, Tomomi Yamaguchi-Miyamoto, Ryutaro Fukui, Natsuko Tanimura, Yuji Motoi, Sachiko Akashi-Takamura, Tatsuhisa Kato, Shigeto Fujishita, Takahito Kimura, B. Umeharu Ohto, Toshiyuki Shimizu, Takatsugu Hirokawa, Kensuke Miyake, Koichi Fukase, Yukari Fujimoto, Yoshinori NagaiKiyoshi Takatsu

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

The Toll-like receptor 4 (TLR4)/myeloid differentiation factor- 2 (MD-2) complex is essential for LPS recognition and induces innate immune responses against Gram-negative bacteria. As activation of TLR4/MD-2 is also critical for the induction of adaptive immune responses, TLR4/MD-2 agonists have been developed as vaccine adjuvants, but their efficacy has not yet been ascertained. Here, we demonstrate that a funiculosin (FNC) variant, FNC-RED, and FNC-RED and FNC derivatives are agonists for both murine and human TLR4/MD-2. FNCRED induced nuclear factor-B (NF-B) activation via murine TLR4/MD-2, whereas FNC had no TLR4/MD-2 stimulatory activity. Biacore analysis revealed that FNC-RED binds to murine TLR4/MD-2 but not murine radioprotective 105 (RP105)/myeloid differentiation factor-1 (MD-1), another LPS sensor. FNC-RED induced CD14-independent expressions of pro-inflammatory cytokines and co-stimulatory molecules in murine macrophages and dendritic cells. In contrast, FNC-RED stimulation was reduced in CD14-dependent LPS responses, including dimerization and internalization of TLR4/MD-2 and IFN-κ expression. FNC-RED-induced IL-12p40 production from murine dendritic cells was dependent on NF-κ B but not MAPK pathway. In addition, fetal bovine serum augmented lipid A-induced NF-κ B activation but blocked FNC-RED-mediated responses. Two synthetic phosphate group-containing FNC-RED and FNC derivatives, FNC-RED-P01 and FNC-P01, respectively, activated human TLR4/MD-2, unlike FNC-RED. Finally, computational analysis revealed that this species-specific activation by FNC-REDandFNC-RED-P01resulted from differences in electrostatic surface potentials between murine and human TLR4/MD-2. We conclude that FNC-RED and its synthetic derivative represent a novel category of murine and human TLR4/MD-2 agonist.

Original languageEnglish
Pages (from-to)15378-15394
Number of pages17
JournalJournal of Biological Chemistry
Volume292
Issue number37
DOIs
Publication statusPublished - 2017

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Toll-Like Receptor 4
Innate Immunity
Derivatives
Chemical activation
Dendritic Cells
Interleukin-12 Subunit p40
Lipid A
Dimerization
Macrophages
Adaptive Immunity
Gram-Negative Bacteria
Static Electricity
Electrostatics
Bacteria
Vaccines
Phosphates
Cytokines
Molecules
human TLR4 protein
Sensors

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Funiculosin variants and phosphorylated derivatives promote innate immune responses via the Toll-like receptor 4/myeloid differentiation factor-2 complex. / Okamoto, Naoki; Mizote, Keisuke; Honda, Hiroe; Saeki, Akinori; Watanabe, Yasuharu; Yamaguchi-Miyamoto, Tomomi; Fukui, Ryutaro; Tanimura, Natsuko; Motoi, Yuji; Akashi-Takamura, Sachiko; Kato, Tatsuhisa; Fujishita, Shigeto; Kimura, Takahito; Ohto, B. Umeharu; Shimizu, Toshiyuki; Hirokawa, Takatsugu; Miyake, Kensuke; Fukase, Koichi; Fujimoto, Yukari; Nagai, Yoshinori; Takatsu, Kiyoshi.

In: Journal of Biological Chemistry, Vol. 292, No. 37, 2017, p. 15378-15394.

Research output: Contribution to journalArticle

Okamoto, N, Mizote, K, Honda, H, Saeki, A, Watanabe, Y, Yamaguchi-Miyamoto, T, Fukui, R, Tanimura, N, Motoi, Y, Akashi-Takamura, S, Kato, T, Fujishita, S, Kimura, T, Ohto, BU, Shimizu, T, Hirokawa, T, Miyake, K, Fukase, K, Fujimoto, Y, Nagai, Y & Takatsu, K 2017, 'Funiculosin variants and phosphorylated derivatives promote innate immune responses via the Toll-like receptor 4/myeloid differentiation factor-2 complex', Journal of Biological Chemistry, vol. 292, no. 37, pp. 15378-15394. https://doi.org/10.1074/jbc.M117.791780
Okamoto, Naoki ; Mizote, Keisuke ; Honda, Hiroe ; Saeki, Akinori ; Watanabe, Yasuharu ; Yamaguchi-Miyamoto, Tomomi ; Fukui, Ryutaro ; Tanimura, Natsuko ; Motoi, Yuji ; Akashi-Takamura, Sachiko ; Kato, Tatsuhisa ; Fujishita, Shigeto ; Kimura, Takahito ; Ohto, B. Umeharu ; Shimizu, Toshiyuki ; Hirokawa, Takatsugu ; Miyake, Kensuke ; Fukase, Koichi ; Fujimoto, Yukari ; Nagai, Yoshinori ; Takatsu, Kiyoshi. / Funiculosin variants and phosphorylated derivatives promote innate immune responses via the Toll-like receptor 4/myeloid differentiation factor-2 complex. In: Journal of Biological Chemistry. 2017 ; Vol. 292, No. 37. pp. 15378-15394.
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abstract = "The Toll-like receptor 4 (TLR4)/myeloid differentiation factor- 2 (MD-2) complex is essential for LPS recognition and induces innate immune responses against Gram-negative bacteria. As activation of TLR4/MD-2 is also critical for the induction of adaptive immune responses, TLR4/MD-2 agonists have been developed as vaccine adjuvants, but their efficacy has not yet been ascertained. Here, we demonstrate that a funiculosin (FNC) variant, FNC-RED, and FNC-RED and FNC derivatives are agonists for both murine and human TLR4/MD-2. FNCRED induced nuclear factor-B (NF-B) activation via murine TLR4/MD-2, whereas FNC had no TLR4/MD-2 stimulatory activity. Biacore analysis revealed that FNC-RED binds to murine TLR4/MD-2 but not murine radioprotective 105 (RP105)/myeloid differentiation factor-1 (MD-1), another LPS sensor. FNC-RED induced CD14-independent expressions of pro-inflammatory cytokines and co-stimulatory molecules in murine macrophages and dendritic cells. In contrast, FNC-RED stimulation was reduced in CD14-dependent LPS responses, including dimerization and internalization of TLR4/MD-2 and IFN-κ expression. FNC-RED-induced IL-12p40 production from murine dendritic cells was dependent on NF-κ B but not MAPK pathway. In addition, fetal bovine serum augmented lipid A-induced NF-κ B activation but blocked FNC-RED-mediated responses. Two synthetic phosphate group-containing FNC-RED and FNC derivatives, FNC-RED-P01 and FNC-P01, respectively, activated human TLR4/MD-2, unlike FNC-RED. Finally, computational analysis revealed that this species-specific activation by FNC-REDandFNC-RED-P01resulted from differences in electrostatic surface potentials between murine and human TLR4/MD-2. We conclude that FNC-RED and its synthetic derivative represent a novel category of murine and human TLR4/MD-2 agonist.",
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T1 - Funiculosin variants and phosphorylated derivatives promote innate immune responses via the Toll-like receptor 4/myeloid differentiation factor-2 complex

AU - Okamoto, Naoki

AU - Mizote, Keisuke

AU - Honda, Hiroe

AU - Saeki, Akinori

AU - Watanabe, Yasuharu

AU - Yamaguchi-Miyamoto, Tomomi

AU - Fukui, Ryutaro

AU - Tanimura, Natsuko

AU - Motoi, Yuji

AU - Akashi-Takamura, Sachiko

AU - Kato, Tatsuhisa

AU - Fujishita, Shigeto

AU - Kimura, Takahito

AU - Ohto, B. Umeharu

AU - Shimizu, Toshiyuki

AU - Hirokawa, Takatsugu

AU - Miyake, Kensuke

AU - Fukase, Koichi

AU - Fujimoto, Yukari

AU - Nagai, Yoshinori

AU - Takatsu, Kiyoshi

PY - 2017

Y1 - 2017

N2 - The Toll-like receptor 4 (TLR4)/myeloid differentiation factor- 2 (MD-2) complex is essential for LPS recognition and induces innate immune responses against Gram-negative bacteria. As activation of TLR4/MD-2 is also critical for the induction of adaptive immune responses, TLR4/MD-2 agonists have been developed as vaccine adjuvants, but their efficacy has not yet been ascertained. Here, we demonstrate that a funiculosin (FNC) variant, FNC-RED, and FNC-RED and FNC derivatives are agonists for both murine and human TLR4/MD-2. FNCRED induced nuclear factor-B (NF-B) activation via murine TLR4/MD-2, whereas FNC had no TLR4/MD-2 stimulatory activity. Biacore analysis revealed that FNC-RED binds to murine TLR4/MD-2 but not murine radioprotective 105 (RP105)/myeloid differentiation factor-1 (MD-1), another LPS sensor. FNC-RED induced CD14-independent expressions of pro-inflammatory cytokines and co-stimulatory molecules in murine macrophages and dendritic cells. In contrast, FNC-RED stimulation was reduced in CD14-dependent LPS responses, including dimerization and internalization of TLR4/MD-2 and IFN-κ expression. FNC-RED-induced IL-12p40 production from murine dendritic cells was dependent on NF-κ B but not MAPK pathway. In addition, fetal bovine serum augmented lipid A-induced NF-κ B activation but blocked FNC-RED-mediated responses. Two synthetic phosphate group-containing FNC-RED and FNC derivatives, FNC-RED-P01 and FNC-P01, respectively, activated human TLR4/MD-2, unlike FNC-RED. Finally, computational analysis revealed that this species-specific activation by FNC-REDandFNC-RED-P01resulted from differences in electrostatic surface potentials between murine and human TLR4/MD-2. We conclude that FNC-RED and its synthetic derivative represent a novel category of murine and human TLR4/MD-2 agonist.

AB - The Toll-like receptor 4 (TLR4)/myeloid differentiation factor- 2 (MD-2) complex is essential for LPS recognition and induces innate immune responses against Gram-negative bacteria. As activation of TLR4/MD-2 is also critical for the induction of adaptive immune responses, TLR4/MD-2 agonists have been developed as vaccine adjuvants, but their efficacy has not yet been ascertained. Here, we demonstrate that a funiculosin (FNC) variant, FNC-RED, and FNC-RED and FNC derivatives are agonists for both murine and human TLR4/MD-2. FNCRED induced nuclear factor-B (NF-B) activation via murine TLR4/MD-2, whereas FNC had no TLR4/MD-2 stimulatory activity. Biacore analysis revealed that FNC-RED binds to murine TLR4/MD-2 but not murine radioprotective 105 (RP105)/myeloid differentiation factor-1 (MD-1), another LPS sensor. FNC-RED induced CD14-independent expressions of pro-inflammatory cytokines and co-stimulatory molecules in murine macrophages and dendritic cells. In contrast, FNC-RED stimulation was reduced in CD14-dependent LPS responses, including dimerization and internalization of TLR4/MD-2 and IFN-κ expression. FNC-RED-induced IL-12p40 production from murine dendritic cells was dependent on NF-κ B but not MAPK pathway. In addition, fetal bovine serum augmented lipid A-induced NF-κ B activation but blocked FNC-RED-mediated responses. Two synthetic phosphate group-containing FNC-RED and FNC derivatives, FNC-RED-P01 and FNC-P01, respectively, activated human TLR4/MD-2, unlike FNC-RED. Finally, computational analysis revealed that this species-specific activation by FNC-REDandFNC-RED-P01resulted from differences in electrostatic surface potentials between murine and human TLR4/MD-2. We conclude that FNC-RED and its synthetic derivative represent a novel category of murine and human TLR4/MD-2 agonist.

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