title: Cellular metabolites enhance the light sensitivity of Arabidopsis cryptochrome through alternate electron transfer pathways.
abstract: Cryptochromes are blue light receptors with multiple signaling roles in plants and animals. Plant cryptochrome (cry1 and cry2) biological activity has been linked to flavin photoreduction via an electron transport chain comprising three evolutionarily conserved tryptophan residues known as the Trp triad. Recently, it has been reported that cry2 Trp triad mutants, which fail to undergo photoreduction in vitro, nonetheless show biological activity in vivo, raising the possibility of alternate signaling pathways. Here, we show that Arabidopsis thaliana cry2 proteins containing Trp triad mutations indeed undergo robust photoreduction in living cultured insect cells. UV/Vis and electron paramagnetic resonance spectroscopy resolves the discrepancy between in vivo and in vitro photochemical activity, as small metabolites, including NADPH, NADH, and ATP, were found to promote cry photoreduction even in mutants lacking the classic Trp triad electron transfer chain. These metabolites facilitate alternate electron transfer pathways and increase light-induced radical pair formation. We conclude that cryptochrome activation is consistent with a mechanism of light-induced electron transfer followed by flavin photoreduction in vivo. We further conclude that in vivo modulation by cellular compounds represents a feature of the cryptochrome signaling mechanism that has important consequences for light responsivity and activation.
authors.affiliation: Fachbereich Physik, Free University, 14195 Berlin, Germany.
authors.forename: Christopher
authors.initials: C
authors.lastname: Engelhard
authors.affiliation: University of Paris VI, UMR 8256 (B2A), IBPS, 75005 Paris, France.
authors.forename: Xuecong
authors.initials: X
authors.lastname: Wang
authors.affiliation: University of Paris VI, UMR 8256 (B2A), IBPS, 75005 Paris, France.
authors.forename: David
authors.initials: D
authors.lastname: Robles
authors.affiliation: Department of Plant Physiology and Photobiology, Faculty of Biology, Philipps-University, 35032 Marburg, Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Moldt
authors.affiliation: Biomedical Research Centre/Faculty of Chemistry, Philipps-University, 35032 Marburg, Germany.
authors.forename: Lars-Oliver
authors.initials: LO
authors.lastname: Essen
authors.affiliation: Department of Plant Physiology and Photobiology, Faculty of Biology, Philipps-University, 35032 Marburg, Germany.
authors.forename: Alfred
authors.initials: A
authors.lastname: Batschauer
authors.affiliation: Fachbereich Physik, Free University, 14195 Berlin, Germany.
authors.forename: Robert
authors.initials: R
authors.lastname: Bittl
authors.affiliation: University of Paris VI, UMR 8256 (B2A), IBPS, 75005 Paris, France Xavier University, Cincinatti, Ohio 45207 margaret.ahmad@upmc.fr.
authors.forename: Margaret
authors.initials: M
authors.lastname: Ahmad
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects*; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D056931:Cryptochromes / Q000235:genetics / Q000378:metabolism*; D004579:Electron Transport; D005415:Flavins / Q000378:metabolism*; D007313:Insecta; D008027:Light; D008958:Models, Molecular; D020125:Mutation, Missense; D010084:Oxidation-Reduction; D015398:Signal Transduction; D014364:Tryptophan / Q000737:chemistry
chemical_list: D029681:Arabidopsis Proteins; C103618:CRY2 protein, Arabidopsis; D056931:Cryptochromes; D005415:Flavins; D014364:Tryptophan