Searching for the mechanism of signalling by plant photoreceptor cryptochrome

International audienceEven 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 or SPA1 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

Diatom molecular research comes of age: Model species for studying phytoplankton biology and diversity

Diatoms are the world’s most diverse group of algae, comprising at least 100,000 species. Contributing;20% of annual global carbon fixation, they underpin major aquatic food webs and drive global biogeochemical cycles. Over the past two decades, Thalassiosira pseudonana and Phaeodactylum tricornutum have become the most important model systems for diatom molecular research, ranging from cell biology to ecophysiology, due to their rapid growth rates, small genomes, and the cumulative wealth of associated genetic resources. To explore the evolutionary divergence of diatoms, additional model species are emerging, such as Fragilariopsis cylindrus and Pseudo-nitzschia multistriata. Here, we describe how functional genomics and reverse genetics have contributed to our understanding of this important class of microalgae in the context of evolution, cell biology, and metabolic adaptations. Our review will also highlight promising areas of investigation into the diversity of these photosynthetic organisms, including the discovery of new molecular pathways governing the life of secondary plastid-bearing organisms in aquatic environments

Correction: Human and Drosophila Cryptochromes Are Light Activated by Flavin Photoreduction in Living Cells

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

Multisignal control of expression of the LHCX protein family in the marine diatom Phaeodactylum tricornutum

Diatoms are phytoplanktonic organisms that grow successfully in the ocean where light conditions are highly variable. Studies of the molecular mechanisms of light acclimation in the marine diatom Phaeodactylum tricornutum show that carotenoid de-epoxidation enzymes and LHCX1, a member of the light-harvesting protein family, both contribute to dissipate excess light energy through non-photochemical quenching (NPQ). In this study, we investigate the role of the other members of the LHCX family in diatom stress responses. Our analysis of available genomic data shows that the presence of multiple LHCX genes is a conserved feature of diatom species living in different ecological niches. Moreover, an analysis of the levels of four P. tricornutum LHCX transcripts in relation to protein expression and photosynthetic activity indicates that LHCXs are differentially regulated under different light intensities and nutrient starvation, mostly modulating NPQ capacity. We conclude that multiple abiotic stress signals converge to regulate the LHCX content of cells, providing a way to fine-tune light harvesting and photoprotection. Moreover, our data indicate that the expansion of the LHCX gene family reflects functional diversification of its members which could benefit cells responding to highly variable ocean environments

Light-driven processes: key players of the functional biodiversity in microalgae

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Light sensing and responses in marine microalgae.

International audienceMarine eukaryotic phytoplankton are major contributors to global primary production. To adapt and thrive in the oceans, phytoplankton relies on a variety of light-regulated responses and light-acclimation capacities probably driven by sophisticated photoregulatory mechanisms. A plethora of photoreceptor-like sequences from marine microalgae have been identified in omics approaches. Initial studies have revealed that some algal photoreceptors are similar to those known in plants. In addition, new variants with different spectral tuning and algal-specific light sensors have also been found, changing current views and perspectives on how photoreceptor structure and function have diversified in phototrophs experiencing different environmental conditions

Dealing with light: The widespread and multitasking cryptochrome/photolyase family in photosynthetic organisms

International audienceLight is essential for the life of photosynthetic organisms as it is a source of energy and information from the environment. Light excess or limitation can be a cause of stress however. Photosynthetic organisms exhibit sophisticated mechanisms to adjust their physiology and growth to the local environmental light conditions. The cryptochrome/photolyase family (CPF) is composed of flavoproteins with similar structures that display a variety of light-dependent functions. This family encompasses photolyases, blue-light activated enzymes that repair ultraviolet-light induced DNA damage, and cryptochromes, known for their photoreceptor functions in terrestrial plants. For this review, we searched extensively for CPFs in the available genome databases to trace the distribution and evolution of this protein family in photosynthetic organisms. By merging molecular data with current knowledge from the functional characterization of CPFs from terrestrial and aquatic organisms, we discuss their roles in (i) photoperception, (ii) biological rhythm regulation and (iii) light-induced stress responses. We also explore their possible implication in light-related physiological acclimation and their distribution in phototrophs living in different environments. The outcome of this structure-function analysis reconstructs the complex scenarios in which CPFs have evolved, as highlighted by the novel functions and biochemical properties of the most recently described family members in algae. (C) 2014 Elsevier GmbH. All rights reserved

AKINbeta3, a plant specific SnRK1 protein, is lacking domains present in yeast and mammals non-catalytic ?-subunits

International audienceThe SNF1/AMPK/SnRK1 heterotrimeric kinase complex is involved in the adaptation of cellular metabolism in response to diverse stresses in yeast, mammals and plants. Following a model proposed in yeast, the kinase targets are likely to bind the complex via the non-catalytic β-subunits. These proteins currently identified in yeast, mammals and plants present a common structure with two conserved interacting domains named Kinase Interacting Sequence (KIS) and Association with SNF1 Complex (ASC), and a highly variable N-terminal domain. In this paper we describe the characterisation of AKINβ3, a novel protein related to AKINβ subunits of Arabidopsis thaliana, containing a truncated KIS domain and no N-terminal extension. Interestingly the missing region of the KIS domain corresponds to the glycogen-binding domain (β-GBD) identified in the mammalian AMPKβ1. In spite of its unusual features, AKINβ3 complements the yeast sip1Δsip2Δgal83Δ mutant. Moreover, interactions between AKINβ3 and other AKIN complex subunits from A. thaliana were detected by two-hybrid experiments and in vitro binding assays. Taken together these data demonstrate that AKINβ3 is a β-type subunit. A search for β-type subunits revealed the existence of β3-type proteins in other plant species. Furthermore, we suggest that the AKINβ3-type subunits could be plant specific since no related sequences have been found in any of the other completely sequenced genomes. These data suggest the existence of novel SnRK1 complexes including AKINβ3-type subunits, involved in several functions among which some could be plant specific

Biochemical and molecular properties of LHCX1, the essential regulator of dynamic photoprotection in diatoms

International audienceAbstract Light harvesting is regulated by a process triggered by the acidification of the thylakoid lumen, known as nonphotochemical “energy-dependent quenching” (qE). In diatoms, qE is controlled by the light-harvesting complex (LHC) protein LHCX1, while the LHC stress-related (LHCSR) and photosystem II subunit S proteins are essential for green algae and plants, respectively. Here, we report a biochemical and molecular characterization of LHCX1 to investigate its role in qE. We found that, when grown under intermittent light, Phaeodactylum tricornutum forms very large qE, due to LHCX1 constitutive upregulation. This “super qE” is abolished in LHCX1 knockout mutants. Biochemical and spectroscopic analyses of LHCX1 reveal that this protein might differ in the character of binding pigments relative to the major pool of light-harvesting antenna proteins. The possibility of transient pigment binding or not binding pigments at all is discussed. Targeted mutagenesis of putative protonatable residues (D95 and E205) in transgenic P. tricornutum lines does not alter qE capacity, showing that they are not involved in sensing lumen pH, differently from residues conserved in LHCSR3. Our results suggest functional divergence between LHCX1 and LHCSR3 in qE modulation. We propose that LHCX1 evolved independently to facilitate dynamic tracking of light fluctuations in turbulent waters. The evolution of LHCX(-like) proteins in organisms with secondary red plastids, such as diatoms, might have conferred a selective advantage in the control of dynamic photoprotection, ultimately resulting in their ecological success

AKINβγ Contributes to SnRK1 Heterotrimeric Complexes and Interacts with Two Proteins Implicated in Plant Pathogen Resistance through Its KIS/GBD Sequence

The sucrose nonfermenting-1 protein kinase (SNF1)/AMP-activated protein kinase subfamily plays a central role in metabolic responses to nutritional and environmental stresses. In yeast (Saccharomyces cerevisiae) and mammals, the β- and γ-noncatalytic subunits are implicated in substrate specificity and subcellular localization, respectively, and regulation of the kinase activity. The atypical βγ-subunit has been previously described in maize (Zea mays), presenting at its N-terminal end a sequence related to the KIS (kinase interacting sequence) domain specific to the β-subunits (Lumbreras et al., 2001). The existence of two components, SNF1-related protein kinase (SnRK1) complexes containing the βγ-subunit and one SnRK1 kinase, had been proposed. In this work, we show that, despite its unusual features, the Arabidopsis (Arabidopsis thaliana) homolog AKINβγ clearly interacts with AKINβ-subunits in vitro and in vivo, suggesting its involvement in heterotrimeric complexes located in both cytoplasm and nucleus. Unexpectedly, a transcriptional analysis of AKINβγ gene expression highlighted the implication of alternative splicing mechanisms in the regulation of AKINβγ expression. A two-hybrid screen performed with AKINβγ as bait, together with in planta bimolecular fluorescence complementation experiments, suggests the existence of interactions in the cytosol between AKINβγ and two leucine-rich repeats related to pathogen resistance proteins. Interestingly, this interaction occurs through the truncated KIS domain that corresponds exactly to a GBD (glycogen-binding domain) recently described in mammals and yeast. A phylogenetic study suggests that AKINβγ-related proteins are restricted to the plant kingdom. Altogether, these data suggest the existence of plant-specific SnRK1 trimeric complexes putatively involved in a plant-specific function such as plant-pathogen interactions