O2- production by B lymphocytes lacking the respiratory burst oxidase subunit p47phox after transfection with an expression vector containing a p47phox cDNA.
The respiratory burst oxidase of phagocytes and B lymphocytes is a complicated enzyme that catalyzes the one-electron reduction of oxygen by NADPH. It is responsible for the O2- production that occurs when these cells are exposed to phorbol 12-myristate 13-acetate or other appropriate stimuli. The activity of this enzyme is greatly decreased or absent in patients with chronic granulomatous disease, an inherited disorder characterized by a severe defect in host defense against bacteria and fungi. In every chronic granulomatous disease patient studied to date, an abnormality has been found in a gene encoding one of four components of the respiratory burst oxidase: the membrane protein p22phox or gp91phox, or the cytosolic protein p47phox or p67phox. We report here that O2- production was partly restored to phorbol 12-myristate 13-acetate-stimulated Epstein-Barr virus-transformed B lymphocytes from a patient with p47phox-deficient chronic granulomatous disease by transfection with an expression plasmid containing a p47phox cDNA inserted in the sense direction. No detectable O2- was produced by untransfected p47phox-deficient lymphocytes or by p47phox-deficient lymphocytes transfected with an antisense plasmid. The finding that O2- can be produced by p47phox-deficient B lymphocytes after the transfer of a p47phox cDNA into the deficient cells suggests that this system could be useful for studying the function of mutant p47phox proteins in whole cells.
Chanock
S J
SJ
Department of Molecular and Experimental Medicine, Scripps Research Institute, La Jolla, CA 92037.
Faust
L R
LR
Barrett
D
D
Bizal
C
C
Maly
F E
FE
Newburger
P E
PE
Ruedi
J M
JM
Smith
R M
RM
Babior
B M
BM
D000595:Amino Acid Sequence; D000906:Antibodies; D000956:Antigens, Viral / Q000235:genetics; D001402:B-Lymphocytes / Q000187:drug effects / Q000378:metabolism*; D001483:Base Sequence; D004268:DNA-Binding Proteins / Q000235:genetics; D019309:Epstein-Barr Virus Nuclear Antigens; D005822:Genetic Vectors; D004854:Herpesvirus 4, Human / Q000235:genetics / Q000276:immunology; D006801:Humans; D015151:Immunoblotting; D007700:Kinetics; D008163:Luminescent Measurements; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D009247:NADH, NADPH Oxidoreductases / Q000172:deficiency* / Q000235:genetics* / Q000378:metabolism; D019255:NADPH Oxidases; D009838:Oligodeoxyribonucleotides; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D016133:Polymerase Chain Reaction / Q000379:methods; D015183:Restriction Mapping; D013481:Superoxides / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection
D000906:Antibodies; D000956:Antigens, Viral; D004268:DNA-Binding Proteins; D019309:Epstein-Barr Virus Nuclear Antigens; D046911:Macromolecular Substances; D009838:Oligodeoxyribonucleotides; D010455:Peptides; D013481:Superoxides; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; C022709:superoxide-forming enzyme; D013755:Tetradecanoylphorbol Acetate