Abstract

Phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P2) has been known to serve as a substrate for phosphatidylinositol 3-kinase (PI3K) and phosphoinositide-specific phospholipase C (PI-PLC), which can produce PtdIns(3,4,5)P3 and inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) and diacylglycerol (DAG), respectively. In this study, we elucidated the role of PI-PLC during the LPS-activated mouse macrophages RAW264.7 treated with PI3K inhibitor wortmannin. First, wortmannin treatment enhanced Ins(1,4,5)P3 production and iNOS expression in LPS-activated macrophages. Inhibition of PI3K by p85 siRNA also showed an enhancement of iNOS expression. On the other hand, overexpression of PI3K by ras-p110 expression plasmid significantly decreased iNOS expression in LPS-activated macrophages. In addition, overexpression of wild-type or dominant-negative Akt expression plasmid did not affect the iNOS expression in LPS-activated macrophages. Second, treatment of PI-PLC inhibitor U73122 reversed the enhancement of iNOS expression, the increase of phosphorylation level of ERK, JNK and p38, and the increase of AP-1-dependent gene expression in wortmannin-treated and LPS-activated macrophages. However, NF-κB activity determined by EMSA assay and reporter plasmid assay did not change during LPS-activated macrophages with or without wortmannin. We propose that the inhibition of PI3K by wortmannin in mouse macrophages enhances the PI-PLC downstream signals, and subsequently increases the LPS induction of iNOS expression independently of Akt pathway.

Original languageEnglish
Pages (from-to)869-879
Number of pages11
JournalBiochimica et Biophysica Acta - Molecular Cell Research
Volume1773
Issue number6
DOIs
Publication statusPublished - Jun 2007

Fingerprint

Phosphoinositide Phospholipase C
Phosphatidylinositol 3-Kinase
Macrophages
Plasmids
Phosphatidylinositol 4,5-Diphosphate
Inositol 1,4,5-Trisphosphate
Diglycerides
Transcription Factor AP-1
Phosphatidylinositols
wortmannin
Small Interfering RNA
Phosphorylation
Gene Expression

Keywords

  • Inducible nitric oxide synthase
  • Lipopolysaccharide
  • Phosphatidylinositol 3-kinase
  • Phosphoinositide-specific phospholipase C
  • Wortmannin

ASJC Scopus subject areas

  • Cell Biology
  • Molecular Biology
  • Biophysics

Cite this

Switch activation of PI-PLC downstream signals in activated macrophages with wortmannin. / Liu, Der Zen; Liang, Hong Jen; Chen, Chien Ho; Lin, Shyr Yi; Zhong, Wen-Bin; Ho, Feng Ming; Hou, Wen Chi; Lo, Jui Lien; Ho, Yuan Soon; Lin, Pei Jung; Hung, Ling Fang; Liang, Yu Chih.

In: Biochimica et Biophysica Acta - Molecular Cell Research, Vol. 1773, No. 6, 06.2007, p. 869-879.

Research output: Contribution to journalArticle

Liu, Der Zen ; Liang, Hong Jen ; Chen, Chien Ho ; Lin, Shyr Yi ; Zhong, Wen-Bin ; Ho, Feng Ming ; Hou, Wen Chi ; Lo, Jui Lien ; Ho, Yuan Soon ; Lin, Pei Jung ; Hung, Ling Fang ; Liang, Yu Chih. / Switch activation of PI-PLC downstream signals in activated macrophages with wortmannin. In: Biochimica et Biophysica Acta - Molecular Cell Research. 2007 ; Vol. 1773, No. 6. pp. 869-879.
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abstract = "Phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P2) has been known to serve as a substrate for phosphatidylinositol 3-kinase (PI3K) and phosphoinositide-specific phospholipase C (PI-PLC), which can produce PtdIns(3,4,5)P3 and inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) and diacylglycerol (DAG), respectively. In this study, we elucidated the role of PI-PLC during the LPS-activated mouse macrophages RAW264.7 treated with PI3K inhibitor wortmannin. First, wortmannin treatment enhanced Ins(1,4,5)P3 production and iNOS expression in LPS-activated macrophages. Inhibition of PI3K by p85 siRNA also showed an enhancement of iNOS expression. On the other hand, overexpression of PI3K by ras-p110 expression plasmid significantly decreased iNOS expression in LPS-activated macrophages. In addition, overexpression of wild-type or dominant-negative Akt expression plasmid did not affect the iNOS expression in LPS-activated macrophages. Second, treatment of PI-PLC inhibitor U73122 reversed the enhancement of iNOS expression, the increase of phosphorylation level of ERK, JNK and p38, and the increase of AP-1-dependent gene expression in wortmannin-treated and LPS-activated macrophages. However, NF-κB activity determined by EMSA assay and reporter plasmid assay did not change during LPS-activated macrophages with or without wortmannin. We propose that the inhibition of PI3K by wortmannin in mouse macrophages enhances the PI-PLC downstream signals, and subsequently increases the LPS induction of iNOS expression independently of Akt pathway.",
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T1 - Switch activation of PI-PLC downstream signals in activated macrophages with wortmannin

AU - Liu, Der Zen

AU - Liang, Hong Jen

AU - Chen, Chien Ho

AU - Lin, Shyr Yi

AU - Zhong, Wen-Bin

AU - Ho, Feng Ming

AU - Hou, Wen Chi

AU - Lo, Jui Lien

AU - Ho, Yuan Soon

AU - Lin, Pei Jung

AU - Hung, Ling Fang

AU - Liang, Yu Chih

PY - 2007/6

Y1 - 2007/6

N2 - Phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P2) has been known to serve as a substrate for phosphatidylinositol 3-kinase (PI3K) and phosphoinositide-specific phospholipase C (PI-PLC), which can produce PtdIns(3,4,5)P3 and inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) and diacylglycerol (DAG), respectively. In this study, we elucidated the role of PI-PLC during the LPS-activated mouse macrophages RAW264.7 treated with PI3K inhibitor wortmannin. First, wortmannin treatment enhanced Ins(1,4,5)P3 production and iNOS expression in LPS-activated macrophages. Inhibition of PI3K by p85 siRNA also showed an enhancement of iNOS expression. On the other hand, overexpression of PI3K by ras-p110 expression plasmid significantly decreased iNOS expression in LPS-activated macrophages. In addition, overexpression of wild-type or dominant-negative Akt expression plasmid did not affect the iNOS expression in LPS-activated macrophages. Second, treatment of PI-PLC inhibitor U73122 reversed the enhancement of iNOS expression, the increase of phosphorylation level of ERK, JNK and p38, and the increase of AP-1-dependent gene expression in wortmannin-treated and LPS-activated macrophages. However, NF-κB activity determined by EMSA assay and reporter plasmid assay did not change during LPS-activated macrophages with or without wortmannin. We propose that the inhibition of PI3K by wortmannin in mouse macrophages enhances the PI-PLC downstream signals, and subsequently increases the LPS induction of iNOS expression independently of Akt pathway.

AB - Phosphatidylinositol (4,5)-bisphosphate (PtdIns(4,5)P2) has been known to serve as a substrate for phosphatidylinositol 3-kinase (PI3K) and phosphoinositide-specific phospholipase C (PI-PLC), which can produce PtdIns(3,4,5)P3 and inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) and diacylglycerol (DAG), respectively. In this study, we elucidated the role of PI-PLC during the LPS-activated mouse macrophages RAW264.7 treated with PI3K inhibitor wortmannin. First, wortmannin treatment enhanced Ins(1,4,5)P3 production and iNOS expression in LPS-activated macrophages. Inhibition of PI3K by p85 siRNA also showed an enhancement of iNOS expression. On the other hand, overexpression of PI3K by ras-p110 expression plasmid significantly decreased iNOS expression in LPS-activated macrophages. In addition, overexpression of wild-type or dominant-negative Akt expression plasmid did not affect the iNOS expression in LPS-activated macrophages. Second, treatment of PI-PLC inhibitor U73122 reversed the enhancement of iNOS expression, the increase of phosphorylation level of ERK, JNK and p38, and the increase of AP-1-dependent gene expression in wortmannin-treated and LPS-activated macrophages. However, NF-κB activity determined by EMSA assay and reporter plasmid assay did not change during LPS-activated macrophages with or without wortmannin. We propose that the inhibition of PI3K by wortmannin in mouse macrophages enhances the PI-PLC downstream signals, and subsequently increases the LPS induction of iNOS expression independently of Akt pathway.

KW - Inducible nitric oxide synthase

KW - Lipopolysaccharide

KW - Phosphatidylinositol 3-kinase

KW - Phosphoinositide-specific phospholipase C

KW - Wortmannin

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