Cellular Metabolites Enhance the Light Sensitivity of Arabidopsis Cryptochrome through Alternate Electron Transfer Pathways

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

Photoreaction of Plant and DASH Cryptochromes Probed by Infrared Spectroscopy: The Neutral Radical State of Flavoproteins

Immeln D, Pokorny R, Herman E, Moldt J, Batschauer A, Kottke T. Photoreaction of Plant and DASH Cryptochromes Probed by Infrared Spectroscopy: The Neutral Radical State of Flavoproteins. Journal of Physical Chemistry B. 2010;114(51):17155-17161

Ordnungsbildung und Erkenntnisprozesse

Ordnungsbildung und Erkenntnis bedingen einander. Erkenntnis setzt die Beobachtung von Ordnungsstrukturen voraus oder deren Schöpfung durch Abstraktion und Modellbildung. Beiträge aus unterschiedlichen Bereichen universitärer Forschung untersuchen Strukturen, die einen Bezug haben zu (partiell) autonomen Akteuren (beziehungsweise Agierenden, Agenten) und den dynamischen Prozessen, in denen sie entwickelt werden. Dabei ablaufende Erkenntnisprozesse erfordern interobjektiv erfahrbare, teilweise auch in Symbolik und Ritualen fassbare Ordnungsstrukturen, auch wenn diese erst simultan mit Handlungs- oder Erkenntnisprozessen entstehen. Rekursive Bezüge können zu Formen der Selbstorganisation führen. Bei höher entwickelten Strukturen können Aspekte des Wissens, Lernens (und Vergessens) einbezogen werden und zusätzlich durch emotionale Zustände verstärkt oder abgeschwächt werden. • Wie entstehen Struktur und Ordnung? • Wie werden sie stabilisiert, modifiziert, revolutioniert, restabilisiert? • Wie werden sie zerstört und aufgelöst? • Wie lässt sich das Verhältnis von Ordnung/Struktur und Wandel/Prozess fassen und (gegebenenfalls formal oder im Rechner) modellieren? • Welche institutionalisierten Mechanismen spielen dabei welche Rolle? • Wie prägen diese Mechanismen die Auseinandersetzungen zwischen Akteuren um "richtiges" und "falsches" Handeln und " richtiges" und "falsches" Wissen von diesem Handeln (Realitätsdefinitionen, Ordnungs- und Zielvorstellungen, Legitimationen)? • Welche Wechselwirkungen bestehen zwischen "stummen" Verhaltensordnungen (Handeln) und "beredter" symbolvermittelter Reflexion eben dieser Verhaltensordnungen (Reden und Wissen)? Wissenschaftlerinnen und Wissenschaftler hatten sich am 5.9.2005 an der Universität Hamburg zu einem Workshop über "Ordnungsbildung und Erkenntnisprozesse" zusammengefunden. In diesem Band sind zahlreiche der gehaltenen Beiträge gesammeltFormation of order and cognition are interdependent. Knowledge presupposes the observation of order structures or their creation through abstraction and modelling. Contributions from different areas of university research examine structures that relate to (partially) autonomous actors (or agents) and the dynamic processes in which they are developed. Processes of knowledge that take place in this context require structures of order that can be experienced interobjectively, and in some cases can also be grasped in symbolism and rituals, even if these structures are created simultaneously with processes of action or knowledge. Recursive references can lead to forms of self-organization. In more highly developed structures, aspects of knowledge, learning (and forgetting) can be included and additionally strengthened or weakened by emotional states. On November 5, 2005, scientists came together for a workshop on "Formation of Order and Knowledge Processes" at the University of Hamburg. In this volume, numerous of the given talks and articles are collected

Photoreduction of the Folate Cofactor in Members of the Photolyase Family*

Cryptochromes and DNA photolyases are related flavoproteins with flavin adenine dinucleotide as the common cofactor. Whereas photolyases repair DNA lesions caused by UV radiation, cryptochromes generally lack repair activity but act as UV-A/blue light photoreceptors. Two distinct electron transfer (ET) pathways have been identified in DNA photolyases. One pathway uses within its catalytic cycle, light-driven electron transfer from FADH−* to the DNA lesion and electron back-transfer to semireduced FADHo after photoproduct cleavage. This cyclic ET pathway seems to be unique for the photolyase subfamily. The second ET pathway mediates photoreduction of semireduced or fully oxidized FAD via a triad of aromatic residues that is conserved in photolyases and cryptochromes. The 5,10-methenyltetrahydrofolate (5,10-methenylTHF) antenna cofactor in members of the photolyase family is bleached upon light excitation. This process has been described as photodecomposition of 5,10-methenylTHF. We show that photobleaching of 5,10-methenylTHF in Arabidopsis cry3, a member of the cryptochrome DASH family, with repair activity for cyclobutane pyrimidine dimer lesions in single-stranded DNA and in Escherichia coli photolyase results from reduction of 5,10-methenylTHF to 5,10-methyleneTHF that requires the intact tryptophan triad. Thus, a third ET pathway exists in members of the photolyase family that remained undiscovered so far