Source:http://linkedlifedata.com/resource/pubmed/id/11929750
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rdf:type | |
lifeskim:mentions | |
pubmed:issue |
8
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pubmed:dateCreated |
2002-4-3
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pubmed:abstractText |
The phagocyte nicotinamide adenine dinucleotide phosphate (reduced form) (NADPH) oxidase was functionally reconstituted in monkey kidney COS-7 cells by transfection of essential subunits, gp91(phox), p22(phox), p47(phox), and p67(phox). COS-7 cells express the essential small guanosine 5'-triphosphatase, Rac1. Transgenic COS-phox cells were capable of arachidonic acid-induced NADPH oxidase activity up to 80% of that of human neutrophils, and of phorbol myristate acetate (PMA)-induced activity up to 20% of that of neutrophils. Expression of all 4 phox components was required for enzyme activity, and enzyme activation was associated with membrane translocation of p47(phox), p67(phox), and Rac1. Expression of p47(phox) Ser303Ala/Ser304Ala or Ser379Ala phosphorylation-deficient mutants resulted in significantly impaired NAPDH oxidase activity, compared with expression of wild-type p47(phox) or the p47(phox) Ser303Glu/Ser304Glu phosphorylation mimic, suggesting that p47(phox) phosphorylation contributes to enzyme activity in the COS system, as is the case in neutrophils. Hence, COS-phox cells should be useful as a new whole-cell model that is both capable of high-level superoxide production and readily amenable to genetic manipulation for investigation of NADPH oxidase function. PMA-elicited superoxide production in COS-phox cells was regulated by activation of protein kinase C (PKC) and Rac. Although COS-7 cells differ from human neutrophils in PKC isoform expression, transient expression of major neutrophil isoforms in COS-phox cells did not increase PMA-induced superoxide production, suggesting that endogenous isoforms were not rate limiting. Val204 in p67(phox), previously shown to be required for NADPH oxidase activity under cell-free conditions, was found to be essential for superoxide production by intact COS-phox cells, on the basis of transfection studies using a p67(phox) (Val204Ala) mutant.
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pubmed:grant |
http://linkedlifedata.com/resource/pubmed/grant/2RO1AI22564,
http://linkedlifedata.com/resource/pubmed/grant/AI35947,
http://linkedlifedata.com/resource/pubmed/grant/CA84138,
http://linkedlifedata.com/resource/pubmed/grant/GM37696,
http://linkedlifedata.com/resource/pubmed/grant/P50DK49218,
http://linkedlifedata.com/resource/pubmed/grant/R01 AI022564-14,
http://linkedlifedata.com/resource/pubmed/grant/R01 AI022564-15,
http://linkedlifedata.com/resource/pubmed/grant/R01 AI022564-16A1,
http://linkedlifedata.com/resource/pubmed/grant/R01HL45635
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pubmed:language |
eng
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pubmed:journal | |
pubmed:citationSubset |
AIM
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pubmed:chemical |
http://linkedlifedata.com/resource/pubmed/chemical/NADPH Oxidase,
http://linkedlifedata.com/resource/pubmed/chemical/Phosphoproteins,
http://linkedlifedata.com/resource/pubmed/chemical/Protein Kinase C,
http://linkedlifedata.com/resource/pubmed/chemical/Protein Subunits,
http://linkedlifedata.com/resource/pubmed/chemical/Superoxides,
http://linkedlifedata.com/resource/pubmed/chemical/Tetradecanoylphorbol Acetate,
http://linkedlifedata.com/resource/pubmed/chemical/neutrophil cytosol factor 67K,
http://linkedlifedata.com/resource/pubmed/chemical/neutrophil cytosolic factor 1,
http://linkedlifedata.com/resource/pubmed/chemical/rac1 GTP-Binding Protein
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pubmed:status |
MEDLINE
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pubmed:month |
Apr
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pubmed:issn |
0006-4971
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pubmed:author | |
pubmed:issnType |
Print
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pubmed:day |
15
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pubmed:volume |
99
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
2653-61
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pubmed:dateRevised |
2010-12-3
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pubmed:meshHeading |
pubmed-meshheading:11929750-Animals,
pubmed-meshheading:11929750-COS Cells,
pubmed-meshheading:11929750-Humans,
pubmed-meshheading:11929750-NADPH Oxidase,
pubmed-meshheading:11929750-Neutrophils,
pubmed-meshheading:11929750-Organisms, Genetically Modified,
pubmed-meshheading:11929750-Phagocytes,
pubmed-meshheading:11929750-Phosphoproteins,
pubmed-meshheading:11929750-Protein Kinase C,
pubmed-meshheading:11929750-Protein Subunits,
pubmed-meshheading:11929750-Respiratory Burst,
pubmed-meshheading:11929750-Superoxides,
pubmed-meshheading:11929750-Tetradecanoylphorbol Acetate,
pubmed-meshheading:11929750-Transfection,
pubmed-meshheading:11929750-rac1 GTP-Binding Protein
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pubmed:year |
2002
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pubmed:articleTitle |
Creation of a genetic system for analysis of the phagocyte respiratory burst: high-level reconstitution of the NADPH oxidase in a nonhematopoietic system.
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pubmed:affiliation |
Herman B Wells Center for Pediatric Research, Department of Pediatrics, James Whitcomb Riley Hospital for Children, Indiana University Medical Center, Indianapolis 46202, USA.
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pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.
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