Murine TOLL-like receptor 4 confers lipopolysaccharide responsiveness as determined by activation of NFκB and expression of the inducible cyclooxygenase

Sang H. Rhee, Daniel Hwang

Research output: Contribution to journalArticle

224 Citations (Scopus)

Abstract

Genetic evidence indicating that TOLL-like receptor 4 (Tlr4) is the lipopolysaccharide (LPS) receptor in mice was reported. However, biochemical evidence that murine Tlr4 confers LPS responsiveness has not been convincingly demonstrated. Inducible cyclooxygenase (COX-2) is selectively expressed in LPS-stimulated macrophages in part mediated through the activation of NFκB. Thus, we determined whether murine Tlr4 confers LPS responsiveness as evaluated by the activation of NFκB and COX-2 expression. Transfection of a murine macrophage-like cell line (RAW264.7) with the constitutively active form (ΔTlr4) of Tlr4 is sufficient to activate NFκB and COX-2 expression. However, the truncated form (ΔTlr4(P712H)) of the missense mutant Tlr4(P712H) found in LPS-hyporesponsive mouse strain (C3H/HeJ) inhibits LPS-induced NFκB activation and COX-2 expression. The inability of ΔTlr4(P712H) to activate NFκB and induce COX-2 expression is rescued by a constitutively active adapter protein myeloid differentiation factor 88 (MyD88), which interacts directly with the cytoplasmic domain of Tlr proteins. Furthermore, MyD88 is co-immunoprecipitated with the wild-type ΔTlr4 but not with the ΔTlr4(P712H) mutant. Together, these results indicate that Tlr4 confers LPS responsiveness in RAW264.7 cells and suggest that hyporesponsiveness of C3H/HeJ mice to LPS is attributed to the disruption of Tlr4-mediated signaling pathways that results from the inability of the mutant Tlr4(P712H) to interact with MyD88.

Original languageEnglish (US)
Pages (from-to)34035-34040
Number of pages6
JournalJournal of Biological Chemistry
Volume275
Issue number44
StatePublished - Nov 3 2000
Externally publishedYes

Fingerprint

Prostaglandin-Endoperoxide Synthases
Lipopolysaccharides
Chemical activation
Myeloid Differentiation Factor 88
Inbred C3H Mouse
Macrophages
CD14 Antigens
Cyclooxygenase 2
Transfection
Proteins
Cells
Cell Line

ASJC Scopus subject areas

  • Biochemistry

Cite this

Murine TOLL-like receptor 4 confers lipopolysaccharide responsiveness as determined by activation of NFκB and expression of the inducible cyclooxygenase. / Rhee, Sang H.; Hwang, Daniel.

In: Journal of Biological Chemistry, Vol. 275, No. 44, 03.11.2000, p. 34035-34040.

Research output: Contribution to journalArticle

@article{a951e88512ed4a1e9631f8de03084f59,
title = "Murine TOLL-like receptor 4 confers lipopolysaccharide responsiveness as determined by activation of NFκB and expression of the inducible cyclooxygenase",
abstract = "Genetic evidence indicating that TOLL-like receptor 4 (Tlr4) is the lipopolysaccharide (LPS) receptor in mice was reported. However, biochemical evidence that murine Tlr4 confers LPS responsiveness has not been convincingly demonstrated. Inducible cyclooxygenase (COX-2) is selectively expressed in LPS-stimulated macrophages in part mediated through the activation of NFκB. Thus, we determined whether murine Tlr4 confers LPS responsiveness as evaluated by the activation of NFκB and COX-2 expression. Transfection of a murine macrophage-like cell line (RAW264.7) with the constitutively active form (ΔTlr4) of Tlr4 is sufficient to activate NFκB and COX-2 expression. However, the truncated form (ΔTlr4(P712H)) of the missense mutant Tlr4(P712H) found in LPS-hyporesponsive mouse strain (C3H/HeJ) inhibits LPS-induced NFκB activation and COX-2 expression. The inability of ΔTlr4(P712H) to activate NFκB and induce COX-2 expression is rescued by a constitutively active adapter protein myeloid differentiation factor 88 (MyD88), which interacts directly with the cytoplasmic domain of Tlr proteins. Furthermore, MyD88 is co-immunoprecipitated with the wild-type ΔTlr4 but not with the ΔTlr4(P712H) mutant. Together, these results indicate that Tlr4 confers LPS responsiveness in RAW264.7 cells and suggest that hyporesponsiveness of C3H/HeJ mice to LPS is attributed to the disruption of Tlr4-mediated signaling pathways that results from the inability of the mutant Tlr4(P712H) to interact with MyD88.",
author = "Rhee, {Sang H.} and Daniel Hwang",
year = "2000",
month = "11",
day = "3",
language = "English (US)",
volume = "275",
pages = "34035--34040",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "44",

}

TY - JOUR

T1 - Murine TOLL-like receptor 4 confers lipopolysaccharide responsiveness as determined by activation of NFκB and expression of the inducible cyclooxygenase

AU - Rhee, Sang H.

AU - Hwang, Daniel

PY - 2000/11/3

Y1 - 2000/11/3

N2 - Genetic evidence indicating that TOLL-like receptor 4 (Tlr4) is the lipopolysaccharide (LPS) receptor in mice was reported. However, biochemical evidence that murine Tlr4 confers LPS responsiveness has not been convincingly demonstrated. Inducible cyclooxygenase (COX-2) is selectively expressed in LPS-stimulated macrophages in part mediated through the activation of NFκB. Thus, we determined whether murine Tlr4 confers LPS responsiveness as evaluated by the activation of NFκB and COX-2 expression. Transfection of a murine macrophage-like cell line (RAW264.7) with the constitutively active form (ΔTlr4) of Tlr4 is sufficient to activate NFκB and COX-2 expression. However, the truncated form (ΔTlr4(P712H)) of the missense mutant Tlr4(P712H) found in LPS-hyporesponsive mouse strain (C3H/HeJ) inhibits LPS-induced NFκB activation and COX-2 expression. The inability of ΔTlr4(P712H) to activate NFκB and induce COX-2 expression is rescued by a constitutively active adapter protein myeloid differentiation factor 88 (MyD88), which interacts directly with the cytoplasmic domain of Tlr proteins. Furthermore, MyD88 is co-immunoprecipitated with the wild-type ΔTlr4 but not with the ΔTlr4(P712H) mutant. Together, these results indicate that Tlr4 confers LPS responsiveness in RAW264.7 cells and suggest that hyporesponsiveness of C3H/HeJ mice to LPS is attributed to the disruption of Tlr4-mediated signaling pathways that results from the inability of the mutant Tlr4(P712H) to interact with MyD88.

AB - Genetic evidence indicating that TOLL-like receptor 4 (Tlr4) is the lipopolysaccharide (LPS) receptor in mice was reported. However, biochemical evidence that murine Tlr4 confers LPS responsiveness has not been convincingly demonstrated. Inducible cyclooxygenase (COX-2) is selectively expressed in LPS-stimulated macrophages in part mediated through the activation of NFκB. Thus, we determined whether murine Tlr4 confers LPS responsiveness as evaluated by the activation of NFκB and COX-2 expression. Transfection of a murine macrophage-like cell line (RAW264.7) with the constitutively active form (ΔTlr4) of Tlr4 is sufficient to activate NFκB and COX-2 expression. However, the truncated form (ΔTlr4(P712H)) of the missense mutant Tlr4(P712H) found in LPS-hyporesponsive mouse strain (C3H/HeJ) inhibits LPS-induced NFκB activation and COX-2 expression. The inability of ΔTlr4(P712H) to activate NFκB and induce COX-2 expression is rescued by a constitutively active adapter protein myeloid differentiation factor 88 (MyD88), which interacts directly with the cytoplasmic domain of Tlr proteins. Furthermore, MyD88 is co-immunoprecipitated with the wild-type ΔTlr4 but not with the ΔTlr4(P712H) mutant. Together, these results indicate that Tlr4 confers LPS responsiveness in RAW264.7 cells and suggest that hyporesponsiveness of C3H/HeJ mice to LPS is attributed to the disruption of Tlr4-mediated signaling pathways that results from the inability of the mutant Tlr4(P712H) to interact with MyD88.

UR - http://www.scopus.com/inward/record.url?scp=0034602150&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034602150&partnerID=8YFLogxK

M3 - Article

C2 - 10952994

AN - SCOPUS:0034602150

VL - 275

SP - 34035

EP - 34040

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 44

ER -