Lipopolysaccharide-mediated signal transduction through phospholipase D activation in monocytic cell lines
| Autor: | Gary K. Iwamoto, Viswanathan Natarajan |
|---|---|
| Rok vydání: | 1994 |
| Předmět: |
Lipopolysaccharides
Biophysics Phosphatidic Acids Glycerophospholipids Biochemistry Diglycerides chemistry.chemical_compound Alkaloids Endocrinology Phospholipase D Tumor Cells Cultured medicine Humans Staurosporine THP1 cell line Protein Kinase C Protein kinase C Diacylglycerol kinase Chemistry Molecular biology Enzyme Activation Tetradecanoylphorbol Acetate Ionomycin Calcium lipids (amino acids peptides and proteins) Intracellular Signal Transduction medicine.drug |
| Zdroj: | Biochimica et Biophysica Acta (BBA) - Lipids and Lipid Metabolism. 1213:14-20 |
| ISSN: | 0005-2760 |
| DOI: | 10.1016/0005-2760(94)90216-x |
| Popis: | Lipopolysaccharide (LPS)-induced phospholipase D (PLD) activation was investigated in undifferentiated monocytic leukemic cell lines THP-1 and U-937. Treatment of THP-1 or U-937 cells labelled with [32P]orthophosphate, [32P]acyl GPC or [3H]alkyl GPC with LPS, in the presence of 0.5% ethanol, resulted in the accumulation of labelled phosphatidylethanol (PEt) through PLD activation. LPS-mediated PLD activation of THP-1 or U-937 was inhibited by staurosporine (2 microM) and by protein kinase C (PKC) down-regulation with 12-O-tetradecanoylphorbol 13-acetate (TPA) suggesting a role for PKC. In addition to LPS, TPA, ionomycin and cell-permeant analogs of diacylglycerol also stimulated [3H]PEt accumulation. The TPA-induced PEt accumulation was also completely abolished by staurosporine or down-regulation of PKC (> 95% inhibition). Furthermore, the LPS-mediated [32P]PEt formation was attenuated by either depletion of extracellular Ca2+ with EGTA (5 mM) or chelation of intracellular Ca2+ by BAPTA (30 microM). These results indicate that an increase in intracellular Ca2+ is necessary for LPS-mediated PLD activation. Further support for PKC activation by LPS was obtained by determining PKC activity in an in vitro assay of histone H1 phosphorylation using [gamma-32P]ATP. In untreated THP-1 cells, approximately 64% of the PKC activity was localized in the cytosol and 36% in the membrane fraction. Treatment of the cells with LPS (10 micrograms/ml, for 2 h) resulted in an increase of 10% of the membrane-associated PKC activity and a corresponding decrease in the cytosol fraction. These data provide evidence that one of the mechanisms of LPS-mediated signal transduction in human monocytic cell lines involves activation of PLD. |
| Databáze: | OpenAIRE |
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