Activation mechanism of NADPH oxidase by SDS in intact guinea pig neutrophils.
It is well known that sodium dodecyl sulfate (SDS) activates NADPH oxidase in a cell-free system independently of protein kinase C (PKC). However, in intact neutrophils, direct evidence has never been presented to show that O2- production by SDS is actually due to the NADPH oxidase activation observed in the cell-free system. So, in this paper, we investigated the activation mechanism by SDS in intact guinea pig neutrophils. We previously reported that hypotonic treatment reversibly enhanced O2- production stimulated by PKC activators in intact neutrophils (M. Hiura et al., 1991, Arch. Biochem. Biophys. 291, 31-37). In this paper, SDS also significantly stimulated O2- production in the intact cells under the hypotonic condition. This enhancement was gradual and was PKC inhibitor resistant. Furthermore, phosphorylation of the 46-kDa protein, one of cytosolic activation factors, was not detected by autoradiography of two-dimensional electrophoresis. Translocation of cytosolic activation factors was demonstrated by a decrease in the activity of the factors remained in the cytosol. In the presence of SDS, addition of 1-oleoyl-2-acetylglycerol, a PKC activator, further enhanced O2- production and translocation of the cytosolic activation factors. On the other hand, SDS remarkably increased membrane fluidity in intact neutrophils as well as in the cell-free system. These results indicate that activation of NADPH oxidase by SDS in intact neutrophils seems to be partly due to the same mechanism observed in cell-free activation, and that SDS alone slightly activates the oxidase and other stimulation, such as hypotonic and/or PKC activator treatments, is required for significant activation. The increase in the membrane fluidity may be one of the activation mechanisms of NADPH oxidase by SDS.
Department of Physiological Chemistry, Hiroshima University School of Medicine, Japan.
J
J
Sasaki
M
M
Hiura
M
M
Yamaguchi
M
M
Sakai
K
K
Aoki
H
H
Abe
N
N
Okamura
S
S
Ishibashi
D000818:Animals; D001692:Biological Transport; D003600:Cytosol / Q000378:metabolism; D004075:Diglycerides / Q000494:pharmacology; D004789:Enzyme Activation; D005260:Female; D006168:Guinea Pigs; D008565:Membrane Proteins / Q000187:drug effects; D009247:NADH, NADPH Oxidoreductases / Q000187:drug effects*; D009252:NADPH Dehydrogenase / Q000032:analysis; D019255:NADPH Oxidases; D009504:Neutrophils / Q000201:enzymology*; D009997:Osmotic Pressure; D010750:Phosphoproteins / Q000032:analysis; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology*; D013481:Superoxides
D004075:Diglycerides; D008565:Membrane Proteins; D010750:Phosphoproteins; C079789:neutrophil cytosol factor 67K; D013481:Superoxides; D012967:Sodium Dodecyl Sulfate; C038237:1-oleoyl-2-acetylglycerol; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; C060505:neutrophil cytosolic factor 1; D009252:NADPH Dehydrogenase; D011493:Protein Kinase C