Redox activation of Nox1 (NADPH Oxidase 1) involves an intermolecular disulfide bond between protein disulfide isomerase and p47phox in vascular smooth muscle cells

Gimenez, M. et al. (2019) Redox activation of Nox1 (NADPH Oxidase 1) involves an intermolecular disulfide bond between protein disulfide isomerase and p47phox in vascular smooth muscle cells. Arteriosclerosis, Thrombosis, and Vascular Biology, 39(2), pp. 224-236. (doi: 10.1161/ATVBAHA.118.311038) (PMID:30580571) (PMCID:PMC6344286)

Abstract

Objective— PDI (protein disulfide isomerase A1) was reported to support Nox1 (NADPH oxidase) activation mediated by growth factors in vascular smooth muscle cells. Our aim was to investigate the molecular mechanism by which PDI activates Nox1 and the functional implications of PDI in Nox1 activation in vascular disease. Approach and Results— Using recombinant proteins, we identified a redox interaction between PDI and the cytosolic subunit p47phox in vitro. Mass spectrometry of crosslinked peptides confirmed redox-dependent disulfide bonds between cysteines of p47phox and PDI and an intramolecular bond between Cys 196 and 378 in p47phox. PDI catalytic Cys 400 and p47phox Cys 196 were essential for the activation of Nox1 by PDI in vascular smooth muscle cells. Transfection of PDI resulted in the rapid oxidation of a redox-sensitive protein linked to p47phox, whereas PDI mutant did not promote this effect. Mutation of p47phox Cys 196, or the redox active cysteines of PDI, prevented Nox1 complex assembly and vascular smooth muscle cell migration. Proximity ligation assay confirmed the interaction of PDI and p47phox in murine carotid arteries after wire injury. Moreover, in human atheroma plaques, a positive correlation between the expression of PDI and p47phox occurred only in PDI family members with the a′ redox active site. Conclusions— PDI redox cysteines facilitate Nox1 complex assembly, thus identifying a new mechanism through which PDI regulates Nox activity in vascular disease

• Item Type: Articles
• Status: Published
• Refereed: Yes
• Glasgow Author(s) Enlighten ID: De Lucca Camargo, Ms Livia
• Authors: Gimenez, M., Veríssimo-Filho, S., Wittig, I., Schickling, B. M., Hahner, F., Schürmann, C., Netto, L. E.S., Rosa, J. C., Brandes, R. P., Sartoretto, S., De Lucca Camargo, L., Abdulkader, F., Miller, F. J., and Lopes, L. R.
• College/School: College of Medical Veterinary and Life Sciences > School of Cardiovascular & Metabolic Health
• Journal Name: Arteriosclerosis, Thrombosis, and Vascular Biology
• Publisher: American Heart Association
• ISSN: 1079-5642
• ISSN (Online): 1524-4636
• Published Online: 06 December 2018