Báo cáo khoa học: Expression and function of Noxo1c, an alternative splicing form of the NADPH oxidase organizer 1

Expression and Function of Noxo1γ, an Alternative Splicing Form of the NADPH Oxidase Organizer 1

Authors: Ryu Takeya, Masahiko Taura, Tomoko Yamasaki, Seiji Naito, Hideki Sumimoto

Field: Biochemistry, Molecular Biology

Document Content: This study investigates Noxo1γ, an alternative splicing variant of the NADPH oxidase organizer 1 (Noxo1). Noxo1γ is expressed in the testis and fetal brain and possesses an additional five amino acids in its N-terminal PX domain, a key module for phosphoinositide binding. While Noxo1γ exhibits a lower affinity for phosphoinositides compared to the classical Noxo1β form, leading to reduced membrane localization in resting cells, it effectively supports Nox1 activation in PMA-stimulated cells. The study further explores the differential roles of Noxo1γ in activating various NADPH oxidase (Nox) family members, including Nox1, Nox2 (gp91phox), and Nox3, highlighting how the insertion in the PX domain influences their respective activation mechanisms and dependencies.

Detailed Table of Contents:

• Abstract
• Keywords
• Correspondence
• Introduction
• Results and Discussion
• Alternative splice forms of human Noxo1
• Expression of Noxo1β and Noxo1γ in various human tissues
• Activation of Nox1 by Noxo1β and Noxo1γ in Chinese hamster ovary (CHO) cells
• Intracellular localization of Noxo1β and Noxo1γ
• Phosphoinositide-binding activity of the PX domains of Noxo1β and Noxo1γ
• Activation of gp91phox and Nox3 by Noxo1γ
• Role of the interaction between Noxo1y and p22phox in Nox1-dependent and Nox3-dependent superoxide production
• Role of phosphoinositide-binding activity of Noxo1 in Nox3 activation
• Concluding remarks
• Experimental procedures
• Isolation of cDNA for splice variants of human NOXO1 gene
• Superoxide-producing activity of CHO cells expressing Nox1, gp91phox, or Nox3
• Measurement of superoxide production using adherent cells undetached from culture dishes
• Estimation of expression of cytosolic regulatory proteins
• Localization of Noxo1y and Noxo1β in CHO cells
• Membrane translocation of Noxo1
• Lipid-binding assay using recombinant GST-fusion proteins
• Two-hybrid experiments
• Acknowledgements
• References