title: Subcellular localization and function of alternatively spliced Noxo1 isoforms.
abstract: Nox organizer 1 (Noxo1), a p47(phox) homolog, is produced as four isoforms with unique N-terminal PX domains derived by alternative mRNA splicing. We compared the subcellular distribution of these isoforms or their isolated PX domains produced as GFP fusion proteins, as well as their ability to support Nox1 activity in several transfected models. Noxo1alpha, beta, gamma, and delta show different subcellular localization patterns, determined by their PX domains. In HEK293 cells, Noxo1beta exhibits prominent plasma membrane binding, Noxo1gamma shows plasma membrane and nuclear associations, and Noxo1alpha and delta localize primarily on intracellular vesicles or cytoplasmic aggregates, but not the plasma membrane. Nox1 activity correlates with Noxo1 plasma membrane binding in HEK293 cells, since Noxo1beta supports the highest activity and Noxo1gamma and Noxo1alpha support moderate or low activities, respectively. In COS-7 cells, where Noxo1alpha localizes on the plasma membrane, the activities supported by the three isoforms (alpha, beta, and gamma) do not differ significantly. The PX domains of beta and gamma bind the same phospholipids, including phosphatidic acid. These results indicate that the variant PX domains are unique determinants of Noxo1 localization and Nox1 function. Finally, the overexpressed Noxo1 isoforms do not affect p22(phox) localization, although Nox1 is needed to transport p22(phox) to the plasma membrane.
authors.affiliation: Molecular Defenses Section, Laboratory of Host Defenses, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA. tueyama@kobe-u.ac.jp
authors.forename: Takehiko
authors.initials: T
authors.lastname: Ueyama
authors.forename: Kristen
authors.initials: K
authors.lastname: Lekstrom
authors.forename: Satoshi
authors.initials: S
authors.lastname: Tsujibe
authors.forename: Naoaki
authors.initials: N
authors.lastname: Saito
authors.forename: Thomas L
authors.initials: TL
authors.lastname: Leto
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport / Q000235:genetics* / Q000378:metabolism* / Q000648:ultrastructure*; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D004789:Enzyme Activation / Q000502:physiology; D006801:Humans; D015151:Immunoblotting; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism / Q000648:ultrastructure; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013347:Subcellular Fractions / Q000201:enzymology* / Q000648:ultrastructure*; D014162:Transfection
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D007527:Isoenzymes; C476419:NOXO1 protein, human; D019255:NADPH Oxidases