A rise in ionized calcium activates the neutrophil NADPH-oxidase but is not sufficient to directly translocate cytosolic p47phox or p67phox to b cytochrome containing membranes.
Neutrophil production of reactive oxygen species is dependent on an assembly process that involves a translocation of the cytosolic NADPH-oxidase components (p47phox; p67phox; Rac2) to a b cytochrome containing membrane. Based on the fact that an intracellular Ca2+ rise can activate the oxidase without any extracellular release of reactive oxygen species, we suggest that the oxidase can be assembled in a membrane distinct from the plasma membrane. Disintegrated cells were used to monitor Ca2+ dependent membrane binding of neutrophil cytosolic proteins. Membranes containing the b cytochrome part of the oxidase, i.e., specific granules and plasma membranes/secretory vesicles, were used in the translocation experiments. Several cytosolic proteins were found to translocate to specific granules as well as the plasma membranes/secretory vesicles, one of them being annexin I. Using antibodies in the blotting assay against the cytosolic oxidase components p47phox and p67phox, we could show that no Ca2+ dependent translocation of these cytosolic proteins occur to neither of the b cytochrome containing membranes.
Movitz
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Department of Medical Microbiology and Immunology, University of Göteborg, Sweden.
Sjölin
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Dahlgren
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D017305:Annexin A1 / Q000378:metabolism; D001693:Biological Transport, Active; D002118:Calcium / Q000378:metabolism*; D003573:Cytochrome b Group / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D066298:In Vitro Techniques; D007425:Intracellular Membranes / Q000378:metabolism; D015759:Ionomycin / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D019255:NADPH Oxidases / Q000378:metabolism*; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism
D017305:Annexin A1; D003573:Cytochrome b Group; D007476:Ionophores; D010750:Phosphoproteins; D017382:Reactive Oxygen Species; C079789:neutrophil cytosol factor 67K; D015759:Ionomycin; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1; D002118:Calcium