title: Blue-light induced accumulation of reactive oxygen species is a consequence of the Drosophila cryptochrome photocycle.
abstract: Cryptochromes are evolutionarily conserved blue-light absorbing flavoproteins which participate in many important cellular processes including in entrainment of the circadian clock in plants, Drosophila and humans. Drosophila melanogaster cryptochrome (DmCry) absorbs light through a flavin (FAD) cofactor that undergoes photoreduction to the anionic radical (FAD•-) redox state both in vitro and in vivo. However, recent efforts to link this photoconversion to the initiation of a biological response have remained controversial. Here, we show by kinetic modeling of the DmCry photocycle that the fluence dependence, quantum yield, and half-life of flavin redox state interconversion are consistent with the anionic radical (FAD•-) as the signaling state in vivo. We show by fluorescence detection techniques that illumination of purified DmCry results in enzymatic conversion of molecular oxygen (O2) to reactive oxygen species (ROS). We extend these observations in living cells to demonstrate transient formation of superoxide (O2•-), and accumulation of hydrogen peroxide (H2O2) in the nucleus of insect cell cultures upon DmCry illumination. These results define the kinetic parameters of the Drosophila cryptochrome photocycle and support light-driven electron transfer to the flavin in DmCry signaling. They furthermore raise the intriguing possibility that light-dependent formation of ROS as a byproduct of the cryptochrome photocycle may contribute to its signaling role.
authors.affiliation: UMR CNRS 8256 (B2A), IBPS, Université Paris VI, Paris, France.|Department of Biomedical Engineering, Florida Institute of Technology, Melbourne, Florida, United States of America.
authors.forename: Louis-David
authors.initials: LD
authors.lastname: Arthaut
authors.affiliation: UMR CNRS 8256 (B2A), IBPS, Université Paris VI, Paris, France.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Jourdan
authors.affiliation: GCRN team, Brain Plasticity Unit, UMR 8249 CNRS/ESPCI Paris, PSL Research University, Paris, France.
authors.forename: Ali
authors.initials: A
authors.lastname: Mteyrek
authors.affiliation: UMR CNRS 8256 (B2A), IBPS, Université Paris VI, Paris, France.|Department of Physics and Astronomy, University of California, Irvine, California, United States of America.
authors.forename: Maria
authors.initials: M
authors.lastname: Procopio
authors.affiliation: UMR CNRS 8256 (B2A), IBPS, Université Paris VI, Paris, France.|Botany Department, Faculty of Science, Tanta University, Tanta, Egypt.
authors.forename: Mohamed
authors.initials: M
authors.lastname: El-Esawi
authors.affiliation: UMR CNRS 8256 (B2A), IBPS, Université Paris VI, Paris, France.
authors.forename: Alain
authors.initials: A
authors.lastname: d'Harlingue
authors.affiliation: UMR CNRS 8256 (B2A), IBPS, Université Paris VI, Paris, France.
authors.forename: Pierre-Etienne
authors.initials: PE
authors.lastname: Bouchet
authors.affiliation: UMR CNRS 8256 (B2A), IBPS, Université Paris VI, Paris, France.
authors.forename: Jacques
authors.initials: J
authors.lastname: Witczak
authors.affiliation: Department of Physics and Astronomy, University of California, Irvine, California, United States of America.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Ritz
authors.affiliation: GCRN team, Brain Plasticity Unit, UMR 8249 CNRS/ESPCI Paris, PSL Research University, Paris, France.
authors.forename: André
authors.initials: A
authors.lastname: Klarsfeld
authors.affiliation: GCRN team, Brain Plasticity Unit, UMR 8249 CNRS/ESPCI Paris, PSL Research University, Paris, France.
authors.forename: Serge
authors.initials: S
authors.lastname: Birman
authors.affiliation: Department of Chemistry and Biochemistry, Montana State University, Bozeman, Montana, United States of America.
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Usselman
authors.affiliation: Botanical Institute and Cluster of Excellence on Plant Sciences (CEPLAS), Biocenter, University of Cologne, Cologne, Germany.
authors.forename: Ute
authors.initials: U
authors.lastname: Hoecker
authors.affiliation: Department of Biomedical Engineering, Florida Institute of Technology, Melbourne, Florida, United States of America.
authors.forename: Carlos F
authors.initials: CF
authors.lastname: Martino
authors.affiliation: UMR CNRS 8256 (B2A), IBPS, Université Paris VI, Paris, France.|Department of Biology, Xavier University, Cincinnati, Ohio, United States of America.
authors.forename: Margaret
authors.initials: M
authors.lastname: Ahmad
mesh_terms: D000818:Animals; D002460:Cell Line; D056931:Cryptochromes / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008027:Light; D017440:Photoperiod; D011789:Quantum Theory; D017382:Reactive Oxygen Species / Q000378:metabolism*; D018411:Spodoptera
chemical_list: D056931:Cryptochromes; D017382:Reactive Oxygen Species