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 language | English (US) |
|---|---|
| Pages (from-to) | 34035-34040 |
| Number of pages | 6 |
| Journal | Journal of Biological Chemistry |
| Volume | 275 |
| Issue number | 44 |
| State | Published - Nov 3 2000 |
| Externally published | Yes |
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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 journal › Article
}
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.
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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 -