Functional association of nox1 with p22phox in vascular smooth muscle cells.
The vascular NAD(P)H oxidases constitute important sources of ROS in the vessel wall and have been implicated in vascular disease. Vascular smooth muscle cells (VSMCs) from conduit arteries express two gp91phox homologs, Nox1 and Nox4, of which Nox1 is agonist-sensitive. Because p22phox has been shown to be functionally important in vascular cells stimulated with vasoactive hormones, the relationship of Nox1 and p22phox was investigated in VSMCs from rat and human aortas. Coimmunoprecipitation studies demonstrated that p22phox and hemagglutinin-tagged Nox1 associate in unstimulated VSMCs. These findings were confirmed by confocal microscopy, showing colocalization of the two proteins in their native states in the plasma membrane and submembrane areas of the cell. NADPH-driven superoxide production, as measured by electron spin resonance using 1-hydroxy-3-carboxypyrrolidine as a spin probe, is dependent on the coexpression of both subunits, suggesting the importance of the association for the functional integrity of the enzyme. These results indicate that in contrast to the neutrophil enzyme, VSMCs can use Nox1 rather than gp91phox as a catalytic center in the p22phox-based oxidase and that these two proteins are preassembled at or near the plasma membrane and submembrane vesicular structures in unstimulated cells.
Division of Cardiology, Department of Medicine, Emory University, 1639 Pierce Drive, Atlanta, GA 30322, USA.
Ibrahim R
IR
Hanna
Lula L
LL
Hilenski
Anna
A
Dikalova
Yoshihiro
Y
Taniyama
Sergey
S
Dikalov
Alicia
A
Lyle
Mark T
MT
Quinn
Bernard
B
Lassègue
Kathy K
KK
Griendling
D000818:Animals; D001011:Aorta / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004578:Electron Spin Resonance Spectroscopy; D026901:Membrane Transport Proteins / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D009252:NADPH Dehydrogenase / Q000378:metabolism*; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases; D010750:Phosphoproteins / Q000378:metabolism*; D011485:Protein Binding; D051381:Rats
D026901:Membrane Transport Proteins; D010750:Phosphoproteins; D009247:NADH, NADPH Oxidoreductases; D000074624:NADPH Oxidase 1; D019255:NADPH Oxidases; C000614712:NOX1 protein, rat; C086986:CYBA protein, human; D009252:NADPH Dehydrogenase