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Margaret Ahmad - One of the best experts on this subject based on the ideXlab platform.
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Arabidopsis Cryptochrome is responsive to Radiofrequency (RF) electromagnetic fields.
Scientific Reports, 2020Co-Authors: Maria Albaqami, Margaret Ahmad, Thorsten Ritz, Marootpong Pooam, Merfat Hammad, Maria Procopio, Mahyar Sameti, Carlos F MartinoAbstract:How living systems respond to weak electromagnetic fields represents one of the major unsolved challenges in sensory biology. Recent evidence has implicated Cryptochrome, an evolutionarily conserved flavoprotein receptor, in magnetic field responses of organisms ranging from plants to migratory birds. However, whether Cryptochromes fulfill the criteria to function as biological magnetosensors remains to be established. Currently, theoretical predictions on the underlying mechanism of chemical magnetoreception have been supported by experimental observations that exposure to radiofrequency (RF) in the MHz range disrupt bird orientation and mammalian cellular respiration. Here we show that, in keeping with certain quantum physical hypotheses, a weak 7 MHz radiofrequency magnetic field significantly reduces the biological responsivity to blue light of the Cryptochrome receptor cry1 in Arabidopsis seedlings. Using an in vivo phosphorylation assay that specifically detects activated Cryptochrome, we demonstrate that RF exposure reduces conformational changes associated with biological activity. RF exposure furthermore alters Cryptochrome-dependent plant growth responses and gene expression to a degree consistent with theoretical predictions. To our knowledge this represents the first demonstration of a biological receptor responding to RF exposure, providing important new implications for magnetosensing as well as possible future applications in biotechnology and medicine.
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Cryptochrome: A photoreceptor with the properties of a magnetoreceptor?
Communicative & integrative biology, 2020Co-Authors: Thorsten Ritz, C Helfrich-foerster, T Yoshii, Margaret AhmadAbstract:It was recently discovered that the photoreceptor Cryptochrome is involved in mediating magnetosensitive entrainment of the internal clock of fruit flies.1 This discovery follows other recent studies implicating a role of Cryptochrome in mediating magnetic sensitivity in orientation responses of fruit flies2,3 and growth responses of plants.4 Such widespread use of the same molecule for mediating magnetic sensitivity might suggest that Cryptochrome is in some way optimal for detecting the magnetic field of the earth and that the magnetoreception function cannot be easily taken over by other molecules. This raises the question what properties might set Cryptochromes apart from other molecules in terms of their ability to detect the geomagnetic field. Here, we will discuss possible answers to this question. We will first review the likely biophysical mechanism by which magnetic fields can be detected by a photoreceptor and discuss what constitutes an optimal photo-magneto-receptor. We will then discuss in how far Cryptochrome matches the profile of an optimal molecule and what further steps are required for more conclusive answers.
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magnetic sensitivity mediated by the arabidopsis blue light receptor Cryptochrome occurs during flavin reoxidation in the dark
Planta, 2019Co-Authors: Marootpong Pooam, Margaret Ahmad, Justin Link, Louisdavid Arthaut, Derek Burdick, Carlos F MartinoAbstract:MAIN CONCLUSION: Arabidopsis Cryptochrome mediates responses to magnetic fields that have been applied in the absence of light, consistent with flavin reoxidation as the primary detection mechanism. Cryptochromes are highly conserved blue-light-absorbing flavoproteins which have been linked to the perception of electromagnetic stimuli in numerous organisms. These include sensing the direction of the earth's magnetic field in migratory birds and the intensity of magnetic fields in insects and plants. When exposed to light, Cryptochromes undergo flavin reduction/reoxidation redox cycles leading to biological activation which generate radical pairs thought to be the basis for magnetic sensitivity. However, the nature of the magnetically sensitive radical pairs and the steps at which they act during the Cryptochrome redox cycle are currently a matter of debate. Here, we investigate the response of Arabidopsis Cryptochrome-1 in vivo to a static magnetic field of 500 μT (10 × earth's field) using both plant growth and light-dependent phosphorylation as an assay. Cryptochrome responses to light were enhanced by the magnetic field, as indicated by increased inhibition of hypocotyl elongation and increased Cryptochrome phosphorylation. However, when light and dark intervals were given intermittently, a plant response to the magnetic field was observed even when the magnetic field was given exclusively during the dark intervals between light exposures. This indicates that the magnetically sensitive reaction step in the Cryptochrome photocycle must occur during flavin reoxidation, and likely involves the formation of reactive oxygen species.
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blue light induced biosynthesis of ros contributes to the signaling mechanism of arabidopsis Cryptochrome
Scientific Reports, 2017Co-Authors: Mohamed A Elesawi, Margaret Ahmad, Justin Link, Louisdavid Arthaut, Carlos F Martino, Nathalie Jourdan, Alain DharlingueAbstract:Cryptochromes are evolutionarily conserved blue light receptors with many roles throughout plant growth and development. They undergo conformational changes in response to light enabling interaction with multiple downstream signaling partners. Recently, it has been shown that Cryptochromes also synthesize reactive oxygen species (ROS) in response to light, suggesting the possibility of an alternate signaling mechanism. Here we show by fluorescence imaging and microscopy that H202 and ROS accumulate in the plant nucleus after Cryptochrome activation. They induce ROS-regulated transcripts including for genes implicated in pathogen defense, biotic and abiotic stress. Mutant Cryptochrome alleles that are non-functional in photomorphogenesis retain the capacity to induce ROS-responsive phenotypes. We conclude that nuclear biosynthesis of ROS by Cryptochromes represents a new signaling paradigm that complements currently known mechanisms. This may lead to novel applications using blue light induced oxidative bursts to prime crop plants against the deleterious effects of environmental stresses and toxins.
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photocycle and signaling mechanisms of plant Cryptochromes
Current Opinion in Plant Biology, 2016Co-Authors: Margaret AhmadAbstract:Cryptochromes are flavoprotein blue light receptors that control many aspects of plant growth and development including seedling de-etiolation, elongation growth, the initiation of flowering, and entrainment of the circadian clock. Photon absorption by Arabidopsis Cryptochromes cry1 and cry2 initiates electron transfer to the oxidized flavin cofactor (FADox) and formation of the presumed biological signaling state FADH°. Current literature on the nature and formation of the signaling state is reviewed, and potential novel roles for Cryptochromes in oxidative stress and as magnetosensors are discussed in light of the Cryptochrome photocycle.
Aziz Sancar - One of the best experts on this subject based on the ideXlab platform.
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Photolyase and Cryptochrome Blue-Light Photoreceptors
Advances in Protein Chemistry, 2020Co-Authors: Aziz SancarAbstract:Publisher Summary This chapter focuses on the photolyase/Cryptochrome blue-light photoreceptor family that encompasses a large group of proteins from all three biological kingdoms. These proteins absorb near-ultraviolet (UV)/blue light and use the light energy to repair far UV-induced DNA damage to reset the circadian clock. The photolyase/Cryptochrome family has three members: photolyase (cyclobutane pyrimidine dimer photolyase), (6–4) photolyase, and Cryptochrome. Both photolyase and Cryptochrome also perform light-independent functions in DNA repair and in generating the molecular circadian clock, respectively. Photolyases are monomeric proteins of 450–550 amino acids and two noncovalently bound cofactors. Escherichia coli (E. coli) photolyase is made up of two well-defined domains: an N terminal α/β domain (residues 1–132) and a C-terminal α-helical domain. Cryptochromes regulate blue-light-dependent growth and development in plants; Cryptochromes have now been identified in the nonphotosynthetic bacterium Vibrio cholerae (V. cholerae), which has no known photoresponses. Cryptochromes exhibit 20–40% sequence identities to photolyases.
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The Second Chromophore in Drosophila Photolyase/Cryptochrome Family Photoreceptors
Biochemistry, 2011Co-Authors: Christopher P Selby, Aziz SancarAbstract:The photolyase/Cryptochrome family of proteins are FAD-containing flavoproteins which carry out blue-light-dependent functions including DNA repair, plant growth and development, and regulation of the circadian clock. In addition to FAD, many members of the family contain a second chromophore which functions as a photoantenna, harvesting light and transferring the excitation energy to FAD and thus increasing the efficiency of the system. The second chromophore is methenyltetrahydrofolate (MTHF) in most photolyases characterized to date and FAD, FMN, or 5-deazariboflavin in others. To date, no second chromophore has been identified in Cryptochromes. Drosophila contains three members of the Cryptochrome/photolyase family: cyclobutane pyrimidine dimer (CPD) photolyase, (6–4) photoproduct photolyase, and Cryptochrome. We developed an expression system capable of incorporating all known second chromophores into the cognate Cryptochrome/photolyase family members. Using this system, we demonstrate that Drosophil...
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ultrafast dynamics and anionic active states of the flavin cofactor in Cryptochrome and photolyase
Journal of the American Chemical Society, 2008Co-Authors: Sang-hun Song, Aziz Sancar, Nuri Ozturk, Jiang Li, Lijuan Wang, Dongping ZhongAbstract:We report here our systematic studies of the dynamics of four redox states of the flavin cofactor in both photolyases and insect type 1 Cryptochromes. With femtosecond resolution, we observed ultrafast photoreduction of oxidized state flavin adenine dinucleotide (FAD) in subpicosecond and of neutral radical semiquinone (FADH•) in tens of picoseconds through intraprotein electron transfer mainly with a neighboring conserved tryptophan triad. Such ultrafast dynamics make these forms of flavin unlikely to be the functional states of the photolyase/Cryptochrome family. In contrast, we find that upon excitation the anionic semiquinone (FAD•−) and hydroquinone (FADH−) have longer lifetimes that are compatible with high-efficiency intermolecular electron transfer reactions. In photolyases, the excited active state (FADH−*) has a long (nanosecond) lifetime optimal for DNA-repair function. In insect type 1 Cryptochromes known to be blue-light photoreceptors the excited active form (FAD•−*) has complex deactivation...
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formation and function of flavin anion radical in Cryptochrome 1 blue light photoreceptor of monarch butterfly
Journal of Biological Chemistry, 2007Co-Authors: Sang-hun Song, Tracy R Denaro, Dongping Zhong, Nuri Ozturk, Ozlem N Arat, Steven M Reppert, Aziz SancarAbstract:Abstract The monarch butterfly (Danaus plexippus) Cryptochrome 1 (DpCry1) belongs in the class of photosensitive insect Cryptochromes. Here we purified DpCry1 expressed in a bacterial host and obtained the protein with a stoichiometric amount of the flavin cofactor in the two-electron oxidized, FADox, form. Exposure of the purified protein to light converts the FADox to the flavin anion radical by intraprotein electron transfer from a Trp residue in the apoenzyme. To test whether this novel photoreduction reaction is part of the DpCry1 physiological photocycle, we mutated the Trp residue that acts as the ultimate electron donor in flavin photoreduction. The mutation, W328F, blocked the photoreduction entirely but had no measurable effect on the light-induced degradation of DpCry1 in vivo. In light of this finding and the recently published action spectrum of this class of Crys, we conclude that DpCry1 and similar insect Cryptochromes do not contain flavin in the FADox form in vivo and that, most likely, the photoreduction reaction is not part of the insect Cryptochrome photoreaction that results in proteolytic degradation of the photopigment.
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a Cryptochrome photolyase class of enzymes with single stranded dna specific photolyase activity
Proceedings of the National Academy of Sciences of the United States of America, 2006Co-Authors: Christopher P Selby, Aziz SancarAbstract:Photolyases and Cryptochrome blue-light photoreceptors are evolutionarily related flavoproteins that perform distinct functions. Photolyases repair UV-damaged DNA in many species from bacteria to plants and animals. Cryptochromes regulate growth and development in plants and the circadian clock in animals. Recently, a new branch of the photolyase/Cryptochrome family was identified. Members of this branch exhibited no or trace levels of DNA repair activity in vivo and in vitro and, therefore, were considered to be Cryptochromes, and they were named Cryptochrome-DASH. Here, we show that Cry-DASH proteins from bacterial, plant, and animal sources actually are photolyases with high degree of specificity for cyclobutane pyrimidine dimers in ssDNA.
Qin Wang - One of the best experts on this subject based on the ideXlab platform.
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the universally conserved residues are not universally required for stable protein expression or functions of Cryptochromes
Molecular Biology and Evolution, 2020Co-Authors: Wenjin He, Tiantian Su, Qin WangAbstract:Universally conserved residues (UCRs) are invariable amino acids evolutionarily conserved among members of a protein family across diverse kingdoms of life. UCRs are considered important for stability and/or function of protein families, but it has not been experimentally examined systematically. Cryptochromes are photoreceptors in plants or light-independent components of the circadian clocks in mammals. We experimentally analyzed 51 UCRs of Arabidopsis Cryptochrome 2 (CRY2) that are universally conserved in eukaryotic Cryptochromes from Arabidopsis to human. Surprisingly, we found that UCRs required for stable protein expression of CRY2 in plants are not similarly required for stable protein expression of human hCRY1 in human cells. Moreover, 74% of the stably expressed CRY2 proteins mutated in UCRs retained wild-type-like activities for at least one photoresponses analyzed. Our finding suggests that the evolutionary mechanisms underlying conservation of UCRs or that distinguish UCRs from non-UCRs determining the same functions of individual Cryptochromes remain to be investigated.
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New insights into the mechanisms of phytochrome–Cryptochrome coaction
New Phytologist, 2017Co-Authors: Qin Wang, Xu WangAbstract:Contents Summary 547 I. Introduction 547 II. Phytochromes mediate light-induced transcription of BICs to inactivate Cryptochromes 548 III. PPKs phosphorylate light-signaling proteins and histones to affect plant development 548 IV. Prospect 550 Acknowledgements 550 References 550 SUMMARY: Plants perceive and respond to light signals by multiple sensory photoreceptors, including phytochromes and Cryptochromes, which absorb different wavelengths of light to regulate genome expression and plant development. Photophysiological analyses have long revealed the coordinated actions of different photoreceptors, a phenomenon referred to as the photoreceptor coaction. The mechanistic explanations of photoreceptor coactions are not fully understood. The function of direct protein-protein interaction of phytochromes and Cryptochromes and common signaling molecules of these photoreceptors, such as SPA1/COP1 E3 ubiquitin ligase complex and bHLH transcription factors PIFs, would partially explain phytochrome-Cryptochrome coactions. In addition, newly discovered proteins that block Cryptochrome photodimerization or catalyze Cryptochrome phosphorylation may also participate in the phytochrome and Cryptochrome coaction. This Tansley insight, which is not intended to make a comprehensive review of the studies of photoreceptor coactions, attempts to highlight those recent findings and their possible roles in the photoreceptor coaction.
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New insights into the mechanisms of phytochrome-Cryptochrome coaction.
New Phytologist, 2017Co-Authors: Qin Wang, Xu WangAbstract:Contents Summary 547 I. Introduction 547 II. Phytochromes mediate light-induced transcription of BICs to inactivate Cryptochromes 548 III. PPKs phosphorylate light-signaling proteins and histones to affect plant development 548 IV. Prospect 550 Acknowledgements 550 References 550 SUMMARY: Plants perceive and respond to light signals by multiple sensory photoreceptors, including phytochromes and Cryptochromes, which absorb different wavelengths of light to regulate genome expression and plant development. Photophysiological analyses have long revealed the coordinated actions of different photoreceptors, a phenomenon referred to as the photoreceptor coaction. The mechanistic explanations of photoreceptor coactions are not fully understood. The function of direct protein-protein interaction of phytochromes and Cryptochromes and common signaling molecules of these photoreceptors, such as SPA1/COP1 E3 ubiquitin ligase complex and bHLH transcription factors PIFs, would partially explain phytochrome-Cryptochrome coactions. In addition, newly discovered proteins that block Cryptochrome photodimerization or catalyze Cryptochrome phosphorylation may also participate in the phytochrome and Cryptochrome coaction. This Tansley insight, which is not intended to make a comprehensive review of the studies of photoreceptor coactions, attempts to highlight those recent findings and their possible roles in the photoreceptor coaction.
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molecular basis for blue light dependent phosphorylation of arabidopsis Cryptochrome 2
Nature Communications, 2017Co-Authors: Weixian Deng, Qin Wang, Xu Wang, Mingxin Piao, Yaxing Li, William D BarshopAbstract:Plant Cryptochromes undergo blue light-dependent phosphorylation to regulate their activity and abundance, but the protein kinases that phosphorylate plant Cryptochromes have remained unclear. Here we show that photoexcited Arabidopsis Cryptochrome 2 (CRY2) is phosphorylated in vivo on as many as 24 different residues, including 7 major phosphoserines. We demonstrate that four closely related Photoregulatory Protein Kinases (previously referred to as MUT9-like kinases) interact with and phosphorylate photoexcited CRY2. Analyses of the ppk123 and ppk124 triple mutants and amiR4k artificial microRNA-expressing lines demonstrate that PPKs catalyse blue light-dependent CRY2 phosphorylation to both activate and destabilize the photoreceptor. Phenotypic analyses of these mutant lines indicate that PPKs may have additional substrates, including those involved in the phytochrome signal transduction pathway. These results reveal a mechanism underlying the co-action of Cryptochromes and phytochromes to coordinate plant growth and development in response to different wavelengths of solar radiation in nature.
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Photoactivation and inactivation of Arabidopsis Cryptochrome 2
Science, 2016Co-Authors: Qin Wang, Lianfeng Gu, Zhaohe Yang, Takeshi Yoshizumi, Xu Wang, Liang YangAbstract:Cryptochromes are blue-light receptors that regulate development and the circadian clock in plants and animals. We found that Arabidopsis Cryptochrome 2 (CRY2) undergoes blue light–dependent homodimerization to become physiologically active. We identified BIC1 (blue-light inhibitor of Cryptochromes 1) as an inhibitor of plant Cryptochromes that binds to CRY2 to suppress the blue light–dependent dimerization, photobody formation, phosphorylation, degradation, and physiological activities of CRY2. We hypothesize that regulated dimerization governs homeostasis of the active Cryptochromes in plants and other evolutionary lineages.
Carrie L Partch - One of the best experts on this subject based on the ideXlab platform.
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animal Cryptochromes divergent roles in light perception circadian timekeeping and beyond
Photochemistry and Photobiology, 2017Co-Authors: Alicia K Michael, Carrie L Partch, Jennifer L Fribourgh, Russell N Van GelderAbstract:Cryptochromes are evolutionarily related to the light-dependent DNA repair enzyme photolyase, serving as major regulators of circadian rhythms in insects and vertebrate animals. There are two types of Cryptochromes in the animal kingdom: Drosophila-like CRYs that act as non-visual photopigments linking circadian rhythms to the environmental light/dark cycle, and vertebrate-like CRYs that do not appear to sense light directly, but control the generation of circadian rhythms by acting as transcriptional repressors. Some animals have both types of CRYs, while others possess only one. Cryptochromes have two domains, the photolyase homology region (PHR) and an extended, intrinsically disordered C-terminus. While all animal CRYs share a high degree of sequence and structural homology in their PHR domains, the C-termini are divergent in both length and sequence identity. Recently, Cryptochrome function has been shown to extend beyond its pivotal role in circadian clocks, participating in regulation of the DNA damage response, cancer progression, and glucocorticoid signaling, as well as being implicated as possible magnetoreceptors. In this review, we provide a historical perspective on the discovery of animal Cryptochromes, examine similarities and differences of the two types of animal Cryptochromes, and explore some of the divergent roles for this class of proteins. This article is protected by copyright. All rights reserved.
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crystal structure of Cryptochrome 3 from arabidopsis thaliana and its implications for photolyase activity
Proceedings of the National Academy of Sciences of the United States of America, 2006Co-Authors: Yihua Huang, Carrie L Partch, Richard H G Baxter, B S Smith, Christopher L Colbert, Johann DeisenhoferAbstract:Cryptochromes use near-UV/blue light to regulate a variety of growth and adaptive process. Recent biochemical studies demonstrate that the Cryptochrome-Drosophila, Arabidopsis, Synechocystis, Human (Cry-DASH) subfamily of Cryptochromes have photolyase activity exclusively for single-stranded cyclobutane pyrimidine dimer (CPD)-containing DNA substrate [Selby C, Sancar A (2006) Proc Natl Acad Sci USA 103:17696–17700]. The crystal structure of Cryptochrome 3 from Arabidopsis thaliana (At-Cry3), a member of the Cry-DASH proteins, at 2.1 A resolution, reveals that both the light-harvesting cofactor 5,10-methenyl-tetrahydrofolyl-polyglutamate (MTHF) and the catalytic cofactor flavin adenine dinucleotide (FAD) are noncovalently bound to the protein. The residues responsible for binding of MTHF in At-Cry3 are not conserved in Escherichia coli photolyase but are strongly conserved in the Cry-DASH subfamily of Cryptochromes. The distance and orientation between MTHF and flavin adenine dinucleotide in At-Cry3 is similar to that of E. coli photolyase, in conjunction with the presence of electron transfer chain, suggesting the conservation of redox activity in At-Cry3. Two amino acid substitutions and the penetration of three charged side chains into the CPD-binding cavity in At-Cry3 alter the hydrophobic environment that is accommodating the hydrophobic sugar ring and thymine base moieties in class I CPD photolyases. These changes most likely make CPD binding less energetically favorable and, hence, insufficient to compete with pairing and stacking interactions between the CPD and the duplex DNA substrate. Thus, Cry-DASH subfamily proteins may be unable to stabilize CPD flipped out from the duplex DNA substrate but may be able to preserve the DNA repair activity toward single-stranded CPD-containing DNA substrate.
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Photochemistry and photobiology of Cryptochrome blue-light photopigments : The search for a photocycle
Photochemistry and Photobiology, 2005Co-Authors: Carrie L Partch, Aziz SancarAbstract:Cryptochromes are flavoproteins that exhibit high sequence and structural similarity to the light-dependent DNA-repair enzyme, photolyase. Cryptochromes have lost the ability to repair DNA; instead, they use the energy from near-UV/blue light to regulate a variety of growth and adaptive processes in organisms ranging from bacteria to humans. The photocycle of Cryptochrome is not yet known, although it is hypothesized that it may share some similarity to that of photolyase, which utilizes light-driven electron transfer from the catalytic flavin chromophore. In this review, we present genetic evidence for the photoreceptive role of Cryptochromes and discuss recent biochemical studies that have furthered our understanding of the Cryptochrome photocycle. In particular, the role of the unique C-terminal domain in Cryptochrome phototransduction is discussed.
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role of structural plasticity in signal transduction by the Cryptochrome blue light photoreceptor
Biochemistry, 2005Co-Authors: Carrie L Partch, Sezgin Özgür, Michael W Clarkson, Aziz SancarAbstract:Cryptochromes are blue-light photoreceptors that regulate a variety of responses such as growth and circadian rhythms in organisms ranging from bacteria to humans. Cryptochromes share a high level of sequence identity with the light-activated DNA repair enzyme photolyase. Photolyase uses energy from blue light to repair UV-induced photoproducts in DNA through cyclic electron transfer between the catalytic flavin adenine dinucleotide cofactor and the damaged DNA. Cryptochromes lack DNA repair activity, and their mechanism of signal transduction is not known. It is hypothesized that a light-dependent signaling state in Cryptochromes is created as a result of an intramolecular redox reaction, resulting in conformational rearrangement and effector binding. Plant and animal Cryptochromes possess 30-250 amino acid carboxy-terminal extensions beyond the photolyase-homology region that have been shown to mediate phototransduction. We analyzed the structures of C-terminal domains from an animal and a plant Cryptochrome by computational, biophysical, and biochemical methods and found these domains to be intrinsically unstructured. We show that the photolyase-homology region interacts with the C-terminal domain, inducing stable tertiary structure in the C-terminal domain. Importantly, we demonstrate a light- dependent conformational change in the C-terminal domain of Arabidopsis Cry1. Collectively, these findings provide the first biochemical evidence for the proposed conformational rearrangement of Cryptochromes upon light exposure.
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further evidence for the role of Cryptochromes in retinohypothalamic photoreception phototransduction
Molecular Brain Research, 2004Co-Authors: Carol L Thompson, David T Plante, Randy J Thresher, Francisco Araujo, Christopher P Selby, Carrie L Partch, Aziz SancarAbstract:Cryptochrome is a blue-light absorbing photopigment that has been proposed to act as a photoreceptor for a variety of nonvisual light-responsive tasks. While mouse models have suggested an important role for Cryptochrome in nonvisual photoreception, there are no biochemical data demonstrating the functional photoreceptive capability of Cryptochrome in mice. There are two models that describe the effect of Cryptochrome on light responsive events: (1) Cryptochrome is a photoreceptor or (2) Cryptochrome is required for either normal phototransduction from the retina to the brain or for normal transcriptional regulation in the brain, irrespective of light. To differentiate between these two models, we have examined the integrity of the regulatory mechanism of c-fos in Cryptochromeless cell lines and in the suprachiasmatic nucleus (SCN) of Cryptochromeless mice. Photoinduction of c-fos mRNA in the SCN can be used as a marker for circadian photoreception/phototransduction and it is drastically reduced in mice lacking Cryptochromes. Our results indicate that light-independent transcription regulatory system of c-fos is normal in Cryptochromeless mice and that the reduced c-fos light responsiveness in the absence of Cryptochromes is due to a loss of photoreceptor function.
Jeanpierre Bouly - One of the best experts on this subject based on the ideXlab platform.
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atp binding turns plant Cryptochrome into an efficient natural photoswitch
Scientific Reports, 2015Co-Authors: Jeanpierre Bouly, Kenichi Hitomi, Pavel Muller, Thorsten Ritz, Elizabeth D. Getzoff, Veronique Balland, Klaus BrettelAbstract:Cryptochromes are flavoproteins that drive diverse developmental light-responses in plants and participate in the circadian clock in animals. Plant Cryptochromes have found application as photoswitches in optogenetics. We have studied effects of pH and ATP on the functionally relevant photoreduction of the oxidized FAD cofactor to the semi-reduced FADH ? radical in isolated Arabidopsis Cryptochrome 1 by transient absorption spectroscopy on nanosecond to millisecond timescales. In the absence of ATP, the yield of light-induced radicals strongly decreased with increasing pH from 6.5 to 8.5. With ATP present, these yields were significantly higher and virtually pH-independent up to pH 9. Analysis of our data in light of the crystallographic structure suggests that ATP-binding shifts the pK a of aspartic acid D396, the putative proton donor to FAD? 2 , from ,7.4 to .9, and favours a reaction pathway yielding long-lived aspartate D396 2. Its negative charge could trigger conformational changes necessary for signal transduction.
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Searching for the mechanism of signalling by plant photoreceptor Cryptochrome
FEBS Letters, 2014Co-Authors: Pavel Muller, Jeanpierre BoulyAbstract:Abstract Even though the plant photoreceptors Cryptochromes were discovered more than 20 years ago, the mechanism through which they transduce light signals to their partner molecules such as COP1 (Constitutive Photomorphogenic 1) or SPA1 (Suppressor of Phytochrome A) still remains to be established. We propose that a negative charge induced by light in the vicinity of the flavin chromophore initiates Cryptochrome 1 signalling. This negative charge might expel the protein-bound ATP from the binding pocket, thereby pushing off the C-terminus that covers the ATP pocket in the dark state of the protein. This conformational change should allow for phosphorylation of previously inaccessible amino acids. A partially phosphorylated ‘ESSSSGRR–VPE’ fragment of the C-terminus could mimic the sequence of the transcription factor HY5 that is essential for binding to the negative regulator of photomorphogenesis COP1. HY5 release through competition for the COP1 binding site could represent the long-sought connection between light activation of Cryptochrome and modulation of photomorphogenesis.
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human and drosophila Cryptochromes are light activated by flavin photoreduction in living cells
PLOS Biology, 2008Co-Authors: Nathalie Hoang, Jeanpierre Bouly, Marie Picot, Albrecht Berndt, Erik Schleicher, Eva Wolf, Sylwia Kacprzak, William Wu, Robert Bittl, Margaret AhmadAbstract:Cryptochromes are a class of flavoprotein blue-light signaling receptors found in plants, animals, and humans that control plant development and the entrainment of circadian rhythms. In plant Cryptochromes, light activation is proposed to result from photoreduction of a protein-bound flavin chromophore through intramolecular electron transfer. However, although similar in structure to plant Cryptochromes, the light-response mechanism of animal Cryptochromes remains entirely unknown. To complicate matters further, there is currently a debate on whether mammalian Cryptochromes respond to light at all or are instead activated by non–light-dependent mechanisms. To resolve these questions, we have expressed both human and Drosophila Cryptochrome proteins to high levels in living Sf21 insect cells using a baculovirus-derived expression system. Intact cells are irradiated with blue light, and the resulting Cryptochrome photoconversion is monitored by fluorescence and electron paramagnetic resonance spectroscopic techniques. We demonstrate that light induces a change in the redox state of flavin bound to the receptor in both human and Drosophila Cryptochromes. Photoreduction from oxidized flavin and subsequent accumulation of a semiquinone intermediate signaling state occurs by a conserved mechanism that has been previously identified for plant Cryptochromes. These results provide the first evidence of how animal-type Cryptochromes are activated by light in living cells. Furthermore, human Cryptochrome is also shown to undergo this light response. Therefore, human Cryptochromes in exposed peripheral and/or visual tissues may have novel light-sensing roles that remain to be elucidated.
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Cryptochrome blue light photoreceptors are activated through interconversion of flavin redox states
Journal of Biological Chemistry, 2007Co-Authors: Jeanpierre Bouly, Nadia Bakrim, Erik Schleicher, Alfred Batschauer, Stefan Meier, Paul Galland, Maribel L Dionisiosese, Filip Vandenbussche, Dominique Van Der Straeten, Robert BittlAbstract:Abstract Cryptochromes are blue light-sensing photoreceptors found in plants, animals, and humans. They are known to play key roles in the regulation of the circadian clock and in development. However, despite striking structural similarities to photolyase DNA repair enzymes, Cryptochromes do not repair double-stranded DNA, and their mechanism of action is unknown. Recently, a blue light-dependent intramolecular electron transfer to the excited state flavin was characterized and proposed as the primary mechanism of light activation. The resulting formation of a stable neutral flavin semiquinone intermediate enables the photoreceptor to absorb green/yellow light (500–630 nm) in addition to blue light in vitro. Here, we demonstrate that Arabidopsis Cryptochrome activation by blue light can be inhibited by green light in vivo consistent with a change of the cofactor redox state. We further characterize light-dependent changes in the Cryptochrome1 (cry1) protein in living cells, which match photoreduction of the purified cry1 in vitro. These experiments were performed using fluorescence absorption/emission and EPR on whole cells and thereby represent one of the few examples of the active state of a known photoreceptor being monitored in vivo. These results indicate that cry1 activation via blue light initiates formation of a flavosemiquinone signaling state that can be converted by green light to an inactive form. In summary, Cryptochrome activation via flavin photoreduction is a reversible mechanism novel to blue light photoreceptors. This photocycle may have adaptive significance for sensing the quality of the light environment in multiple organisms.
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Cryptochrome photoreceptors cry1 and cry2 antagonistically regulate primary root elongation in Arabidopsis thaliana
Planta, 2006Co-Authors: Roberto C. Canamero, Jeanpierre Bouly, Nadia Bakrim, Alvaro Garay, Elizabeth E. Dudkin, Yvette Habricot, Margaret AhmadAbstract:Cryptochromes are blue-light receptors controlling multiple aspects of plant growth and development. They are flavoproteins with significant homology to photolyases, but instead of repairing DNA they function by transducing blue light energy into a signal that can be recognized by the cellular signaling machinery. Here we report the effect of cry1 and cry2 blue light receptors on primary root growth in Arabidopsis thaliana seedlings, through analysis of both Cryptochrome-mutant and Cryptochrome-overexpressing lines. Cry1 mutant seedlings show reduced root elongation in blue light while overexpressing seedlings show significantly increased elongation as compared to wild type controls. By contrast, the cry2 mutation has the opposite effect on root elongation growth as does cry1 , demonstrating that cry1 and cry2 act antagonistically in this response pathway. The site of Cryptochrome signal perception is within the shoot, and the inhibitor of auxin transport, 1- N -naphthylphthalamic acid, abolishes the differential effect of Cryptochromes on root growth, suggesting the blue-light signal is transmitted from the shoot to the root by a mechanism that involves auxin. Primary root elongation in blue light may thereby involve interaction between Cryptochrome and auxin signaling pathways.