Spatial compartmentalization of Tumor Necrosis Factor (TNF) Receptor 1-dependent signalling pathways in human airway smooth muscle cells: Lipid rafts are essential for TNF-a-mediated activation of RhoA but dispensible for the activation of the NF-¿B and MAPK pathways

Irene Hunter, Graeme Fleming Nixon

Research output: Contribution to journalArticle

74 Citations (Scopus)

Abstract

Tumor necrosis factor (TNF)-alpha-induced activation of RhoA, mediated by TNF receptor 1 (TNFR1), is a prerequisite step in a pathway that leads to increased 20-kDa light chain of myosin (MLC20) phosphorylation and airway smooth muscle contraction. In this study, we have investigated the proximal events in TNF-alpha-induced RhoA activation. TNFR1 is localized to both lipid raft and nonraft regions of the plasma membrane in primary human airway smooth muscle cells. TNF-alpha engagement of TNFR1 recruited the adaptor proteins TRADD, TRAF-2, and RIP into lipid rafts and activated RhoA, NF-kappa B, and MAPK pathways. Depletion of cholesterol from rafts with methyl-beta-cyclodextrin caused a redistribution of TNFR1 to nonraft plasma membrane and prevented ligand-induced RhoA activation. By contrast, TNF-alpha-induced activation of NF-kappa B and MAPKs was unaffected by methyl-beta-cyclodextrin indicating that, in airway smooth muscle cells, activation of these pathways occurred independently of lipid rafts. Targeted knockdown of caveolin-1 completely abrogated TNF-alpha-induced RhoA activation, identifying this raft-resident protein as a positive regulator of the activation process. The signaling adaptors TRADD and RIP were also found to be necessary for ligand-induced RhoA activation. Taken together, our results suggest that in airway smooth muscle cells, spatial compartmentalization of TNFR1 provides a mechanism for generating distinct signaling outcomes in response to ligand engagement and define a mechanistic role for lipid rafts and caveolin-1 in TNF-alpha-induced activation of RhoA.

Original languageEnglish
Pages (from-to)34705-34715
Number of pages10
JournalThe Journal of Biological Chemistry
Volume281
Issue number45
DOIs
Publication statusPublished - Nov 2006

Keywords

  • GROWTH-FACTOR RECEPTORS
  • PLASMA-MEMBRANE
  • CAVEOLAE
  • PROTEIN
  • COMPLEX
  • TRANSDUCTION
  • RECRUITMENT
  • MOLECULES
  • APOPTOSIS
  • DOMAIN

Cite this

@article{0652325c71bd490b80062e676f2edeb2,
title = "Spatial compartmentalization of Tumor Necrosis Factor (TNF) Receptor 1-dependent signalling pathways in human airway smooth muscle cells: Lipid rafts are essential for TNF-a-mediated activation of RhoA but dispensible for the activation of the NF-¿B and MAPK pathways",
abstract = "Tumor necrosis factor (TNF)-alpha-induced activation of RhoA, mediated by TNF receptor 1 (TNFR1), is a prerequisite step in a pathway that leads to increased 20-kDa light chain of myosin (MLC20) phosphorylation and airway smooth muscle contraction. In this study, we have investigated the proximal events in TNF-alpha-induced RhoA activation. TNFR1 is localized to both lipid raft and nonraft regions of the plasma membrane in primary human airway smooth muscle cells. TNF-alpha engagement of TNFR1 recruited the adaptor proteins TRADD, TRAF-2, and RIP into lipid rafts and activated RhoA, NF-kappa B, and MAPK pathways. Depletion of cholesterol from rafts with methyl-beta-cyclodextrin caused a redistribution of TNFR1 to nonraft plasma membrane and prevented ligand-induced RhoA activation. By contrast, TNF-alpha-induced activation of NF-kappa B and MAPKs was unaffected by methyl-beta-cyclodextrin indicating that, in airway smooth muscle cells, activation of these pathways occurred independently of lipid rafts. Targeted knockdown of caveolin-1 completely abrogated TNF-alpha-induced RhoA activation, identifying this raft-resident protein as a positive regulator of the activation process. The signaling adaptors TRADD and RIP were also found to be necessary for ligand-induced RhoA activation. Taken together, our results suggest that in airway smooth muscle cells, spatial compartmentalization of TNFR1 provides a mechanism for generating distinct signaling outcomes in response to ligand engagement and define a mechanistic role for lipid rafts and caveolin-1 in TNF-alpha-induced activation of RhoA.",
keywords = "GROWTH-FACTOR RECEPTORS, PLASMA-MEMBRANE, CAVEOLAE, PROTEIN, COMPLEX, TRANSDUCTION, RECRUITMENT, MOLECULES, APOPTOSIS, DOMAIN",
author = "Irene Hunter and Nixon, {Graeme Fleming}",
year = "2006",
month = "11",
doi = "10.1074/JBC.M605738200",
language = "English",
volume = "281",
pages = "34705--34715",
journal = "The Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC",
number = "45",

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TY - JOUR

T1 - Spatial compartmentalization of Tumor Necrosis Factor (TNF) Receptor 1-dependent signalling pathways in human airway smooth muscle cells: Lipid rafts are essential for TNF-a-mediated activation of RhoA but dispensible for the activation of the NF-¿B and MAPK pathways

AU - Hunter, Irene

AU - Nixon, Graeme Fleming

PY - 2006/11

Y1 - 2006/11

N2 - Tumor necrosis factor (TNF)-alpha-induced activation of RhoA, mediated by TNF receptor 1 (TNFR1), is a prerequisite step in a pathway that leads to increased 20-kDa light chain of myosin (MLC20) phosphorylation and airway smooth muscle contraction. In this study, we have investigated the proximal events in TNF-alpha-induced RhoA activation. TNFR1 is localized to both lipid raft and nonraft regions of the plasma membrane in primary human airway smooth muscle cells. TNF-alpha engagement of TNFR1 recruited the adaptor proteins TRADD, TRAF-2, and RIP into lipid rafts and activated RhoA, NF-kappa B, and MAPK pathways. Depletion of cholesterol from rafts with methyl-beta-cyclodextrin caused a redistribution of TNFR1 to nonraft plasma membrane and prevented ligand-induced RhoA activation. By contrast, TNF-alpha-induced activation of NF-kappa B and MAPKs was unaffected by methyl-beta-cyclodextrin indicating that, in airway smooth muscle cells, activation of these pathways occurred independently of lipid rafts. Targeted knockdown of caveolin-1 completely abrogated TNF-alpha-induced RhoA activation, identifying this raft-resident protein as a positive regulator of the activation process. The signaling adaptors TRADD and RIP were also found to be necessary for ligand-induced RhoA activation. Taken together, our results suggest that in airway smooth muscle cells, spatial compartmentalization of TNFR1 provides a mechanism for generating distinct signaling outcomes in response to ligand engagement and define a mechanistic role for lipid rafts and caveolin-1 in TNF-alpha-induced activation of RhoA.

AB - Tumor necrosis factor (TNF)-alpha-induced activation of RhoA, mediated by TNF receptor 1 (TNFR1), is a prerequisite step in a pathway that leads to increased 20-kDa light chain of myosin (MLC20) phosphorylation and airway smooth muscle contraction. In this study, we have investigated the proximal events in TNF-alpha-induced RhoA activation. TNFR1 is localized to both lipid raft and nonraft regions of the plasma membrane in primary human airway smooth muscle cells. TNF-alpha engagement of TNFR1 recruited the adaptor proteins TRADD, TRAF-2, and RIP into lipid rafts and activated RhoA, NF-kappa B, and MAPK pathways. Depletion of cholesterol from rafts with methyl-beta-cyclodextrin caused a redistribution of TNFR1 to nonraft plasma membrane and prevented ligand-induced RhoA activation. By contrast, TNF-alpha-induced activation of NF-kappa B and MAPKs was unaffected by methyl-beta-cyclodextrin indicating that, in airway smooth muscle cells, activation of these pathways occurred independently of lipid rafts. Targeted knockdown of caveolin-1 completely abrogated TNF-alpha-induced RhoA activation, identifying this raft-resident protein as a positive regulator of the activation process. The signaling adaptors TRADD and RIP were also found to be necessary for ligand-induced RhoA activation. Taken together, our results suggest that in airway smooth muscle cells, spatial compartmentalization of TNFR1 provides a mechanism for generating distinct signaling outcomes in response to ligand engagement and define a mechanistic role for lipid rafts and caveolin-1 in TNF-alpha-induced activation of RhoA.

KW - GROWTH-FACTOR RECEPTORS

KW - PLASMA-MEMBRANE

KW - CAVEOLAE

KW - PROTEIN

KW - COMPLEX

KW - TRANSDUCTION

KW - RECRUITMENT

KW - MOLECULES

KW - APOPTOSIS

KW - DOMAIN

U2 - 10.1074/JBC.M605738200

DO - 10.1074/JBC.M605738200

M3 - Article

VL - 281

SP - 34705

EP - 34715

JO - The Journal of Biological Chemistry

JF - The Journal of Biological Chemistry

SN - 0021-9258

IS - 45

ER -