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Evidence for protein-catalyzed transfer of platelet activating factor by macrophage cytosol
J B Banks1, R L Wykle, J T O'Flaherty
1Biochemistry Group, Western Carolina University, Cullowhee, NC 28723.
Abstract:
Platelet activating factor (PAF) is a potent, proinflammatory lipid. PAF is synthesized in response to stimuli and is rapidly destroyed by specific acetylhydrolases. In order to express its biological activity, PAF and its metabolites are transported among subcellular membranes by as yet unexplained mechanisms. We report here an assay system using methylcarbamyl-PAF (CPAF, 1-O-hexadecyl-2-O-(N-methylcarbamyl)-sn-glycero-3-phosphocholine) and a vesicle-extrusion technique for examining protein-catalyzed intermembrane transfer of CPAF, and demonstrate the presence of proteins catalyzing the separate transfer of CPAF and diacyl phosphatidylcholine in macrophage cytosol. The CPAF transfer activity is heat- and trypsin-sensitive and elutes from gel-filtration columns well separated from proteins catalyzing the transfer of phosphatidylcholine.
Insights
Researchers identified proteins in macrophages that transfer platelet-activating factor (PAF) between membranes. This finding helps explain how this potent inflammatory lipid moves within cells.
Area of Science:
- Lipid signaling
- Cellular biology
- Biochemistry
Background:
- Platelet-activating factor (PAF) is a potent pro-inflammatory lipid mediator.
- PAF synthesis and degradation are rapid, necessitating efficient intracellular transport mechanisms.
- The mechanisms governing PAF and its metabolite transport between subcellular membranes remain largely unexplained.
Purpose of the Study:
- To develop an assay system for studying protein-catalyzed intermembrane transfer of PAF.
- To investigate the presence and characteristics of proteins involved in CPAF transfer in macrophages.
Main Methods:
- Development of an assay using methylcarbamyl-PAF (CPAF) and a vesicle-extrusion technique.
- Examination of protein-catalyzed intermembrane transfer of CPAF in macrophage cytosol.
- Biochemical characterization of CPAF transfer activity, including heat and trypsin sensitivity, and gel-filtration chromatography.
Main Results:
- Demonstration of proteins in macrophage cytosol that catalyze the transfer of CPAF.
- Identification of separate proteins responsible for the transfer of CPAF and diacyl phosphatidylcholine.
- Characterization of CPAF transfer activity as heat- and trypsin-sensitive.
Conclusions:
- Macrophage cytosol contains specific proteins that facilitate the intermembrane transfer of CPAF.
- These transfer proteins are distinct from those involved in phosphatidylcholine transport.
- The findings provide insight into the mechanisms of intracellular lipid mediator trafficking.