The activity of gp91phox, the catalytic subunit of the superoxide- generating respiratory burst oxidase, is stimulated by the regulatory subunits p47phox, p67phox and the small GTPase Rac. Novel homologs of p47phox and p67phox (NOXO1 and NOXA1, respectively) were recently identified and are implicated in the regulation of the gp91phox homologs Nox1 and Nox3. Herein, we report four splice forms of human NOXO1. NOXO1β is the major mRNA splice form in human colon and fetal liver while NOXO1γ was the majority species in testis. Neither the α nor δ forms were expressed in significant amounts in any tissue tested. Splice forms were generated by alternative splicing of the two ends of exon 3 of the NOXO1 gene, and resulted in differences in the PX domain. The PX domain is known to bind inositol lipids, but the expressed, purified PX domains from NOXO1β and NOXO1γ bound these lipids with the same specificity and affinity. NOXO1β and NOXO1γ both activated Nox1, but NOXO1γ showed a poorer ability to activate Nox3 compared with NOXO1β. These data suggest different tissue localizations and functions for NOXO1β and NOXO1γ in regulating Nox family members. © 2005 Elsevier B.V. All rights reserved.