Light‐driven Activation of NADP(H) Oxidase 5

Abstract

Ultraviolet band A (UVA) light irradiation has been shown to induce cytosolic calcium oscillations in the isolated rat peritoneal mast cells via activation of NADPH oxidase (NOX). It has been clarified that UVA-triggered calcium oscillations in mast cells (isolated rat peritoneal mast cells, rat mast cell line RBL-2H3, mouse bone marrow-derived mast cells) was due to NOX2 activation. To elucidate whether UVA light irradiation would also activate NOX5, a monomeric isoform of the NOX family of oxidases not requiring transmembrane p22 or cytosolic subunits for activity and not endogenously expressed in rodents, NOX5 was expressed ectopically in CHO-K1 cells, and the effect of UVA irradiation or photodynamic action on cytosolic calcium concentration was monitored with Fura-2 fluorescent calcium imaging. It was found that NOX5 expressed ectopically in CHO-K1 cells was activated by UVA (LED 380 nm) light irradiation to induce persistent cytosolic calcium oscillations. Blue LED (450 nm) light irradiation of CHO-K1 cells expressing protein photosensitizer miniSOG-NOX5 fusion protein triggered also persistent cytosolic calcium oscillations which were completely blocked by NOX inhibitor diphenyleneiodonium (DPI), and inhibited by singlet oxygen quencher Trolox C. Sulphonated aluminium phthalocyanine (SALPC) photodynamically activated the heterologously expressed NOX5 in CHO-K1 cells to trigger similarly cytosolic calcium oscillations, which were completely inhibited by DPI. These data suggest that both UVA light irradiation and 1O2 generation with photodynamic action could activate NOX5, to ultimately trigger calcium oscillations. It is therefore possible now to realize light-driven NOX5 activation for cell-type or subcellular organelle specific generation of superoxide anion.