Expression and function of Noxo1gamma, an alternative splicing form of the NADPH oxidase organizer 1.
Activation of the superoxide-producing NADPH oxidase Nox1 requires both the organizer protein Noxo1 and the activator protein Noxa1. Here we describe an alternative splicing form of Noxo1, Noxo1gamma, which is expressed in the testis and fetal brain. The Noxo1gamma protein contains an additional five amino acids in the N-terminal PX domain, a phosphoinositide-binding module; the domain plays an essential role in supporting superoxide production by NADPH oxidase (Nox) family oxidases including Nox1, gp91(phox)/Nox2, and Nox3, as shown in this study. The PX domain isolated from Noxo1gamma shows a lower affinity for phosphoinositides than that from the classical splicing form Noxo1beta. Consistent with this, in resting cells, Noxo1gamma is poorly localized to the membrane, and thus less effective in activating Nox1 than Noxo1beta, which is constitutively present at the membrane. On the other hand, cell stimulation with phorbol 12-myristate 13-acetate (PMA), an activator of Nox1-3, facilitates membrane translocation of Noxo1gamma; as a result, Noxo1gamma is equivalent to Noxo1beta in Nox1 activation in PMA-stimulated cells. The effect of the five-amino-acid insertion in the Noxo1 PX domain appears to depend on the type of Nox; in activation of gp91(phox)/Nox2, Noxo1gamma is less active than Noxo1beta even in the presence of PMA, whereas Noxo1gamma and Noxo1beta support the superoxide-producing activity of Nox3 to the same extent in a manner independent of cell stimulation.
Takeya
Ryu
R
Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan.
Taura
Masahiko
M
Yamasaki
Tomoko
T
Naito
Seiji
S
Sumimoto
Hideki
H
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism*; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000235:genetics; D006801:Humans; D008562:Membrane Glycoproteins / Q000187:drug effects / Q000378:metabolism; D008565:Membrane Proteins / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism*; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000187:drug effects / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism; D017434:Protein Structure, Tertiary; D013481:Superoxides / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D018076:DNA, Complementary; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C476419:NOXO1 protein, human; D010716:Phosphatidylinositols; D013481:Superoxides; D009247:NADH, NADPH Oxidoreductases; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; C431570:Nox3 protein, human