Publications by authors named "Angela Falciatore"

41 Publications

Engineering the Unicellular Alga for Enhancing Carotenoid Production.

Antioxidants (Basel) 2020 Aug 16;9(8). Epub 2020 Aug 16.

Stazione Zoologica Anton Dohrn, Villa Comunale, 80121 Naples, Italy.

Microalgae represent a promising resource for the production of beneficial natural compounds due to their richness in secondary metabolites and easy cultivation. Carotenoids feature among distinctive compounds of many microalgae, including diatoms, which owe their golden color to the xanthophyll fucoxanthin. Carotenoids have antioxidant, anti-obesity and anti-inflammatory properties, and there is a considerable market demand for these compounds. Here, with the aim to increase the carotenoid content in the model diatom , we exploited genetic transformation to overexpress genes involved in the carotenoid biosynthetic pathway. We produced transgenic lines over-expressing simultaneously one, two or three carotenoid biosynthetic genes, and evaluated changes in pigment content with high-performance liquid chromatography. Two triple transformants over-expressing the genes Violaxanthin de-epoxidase (), Vde-related () and Zeaxanthin epoxidase 3 () showed an accumulation of carotenoids, with an increase in the fucoxanthin content up to four fold. , and mRNA and protein levels in the triple transformants were coherently increased. The exact role of these enzymes in the diatom carotenoid biosynthetic pathway is not completely elucidated nevertheless our strategy successfully modulated the carotenoid metabolism leading to an accumulation of valuable compounds, leading the way toward improved utilization of microalgae in the field of antioxidants.
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http://dx.doi.org/10.3390/antiox9080757DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7465010PMC
August 2020

Genome-Scale Metabolic Reconstruction and Perturbation Analysis of the Polar Diatom Predicts High Metabolic Robustness.

Biology (Basel) 2020 Feb 17;9(2). Epub 2020 Feb 17.

Unité Mixte Internationale 3376 Takuvik, CNRS-ULaval, Département de Biologie and Québec-Océan, Université Laval, Québec, QC G1V 0A6, Canada.

Diatoms are major primary producers in polar environments where they can actively grow under extremely variable conditions. Integrative modeling using a genome-scale model (GSM) is a powerful approach to decipher the complex interactions between components of diatom metabolism and can provide insights into metabolic mechanisms underlying their evolutionary success in polar ecosystems. We developed the first GSM for a polar diatom, , which enabled us to study its metabolic robustness using sensitivity analysis. We find that the predicted growth rate was robust to changes in all model parameters (i.e., cell biochemical composition) except the carbon uptake rate. Constraints on total cellular carbon buffer the effect of changes in the input parameters on reaction fluxes and growth rate. We also show that single reaction deletion of 20% to 32% of active (nonzero flux) reactions and single gene deletion of 44% to 55% of genes associated with active reactions affected the growth rate, as well as the production fluxes of total protein, lipid, carbohydrate, DNA, RNA, and pigments by less than 1%, which was due to the activation of compensatory reactions (e.g., analogous enzymes and alternative pathways) with more highly connected metabolites involved in the reactions that were robust to deletion. Interestingly, including highly divergent alleles unique for increased its metabolic robustness to cellular perturbations even more. Overall, our results underscore the high robustness of metabolism in , a feature that likely helps to maintain cell homeostasis under polar conditions.
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http://dx.doi.org/10.3390/biology9020030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7168318PMC
February 2020

Diatom Molecular Research Comes of Age: Model Species for Studying Phytoplankton Biology and Diversity.

Plant Cell 2020 03 18;32(3):547-572. Epub 2019 Dec 18.

School of Environmental Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, United Kingdom

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, and 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 and 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.
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http://dx.doi.org/10.1105/tpc.19.00158DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7054031PMC
March 2020

Lhcx proteins provide photoprotection via thermal dissipation of absorbed light in the diatom Phaeodactylum tricornutum.

Nat Commun 2019 09 13;10(1):4167. Epub 2019 Sep 13.

Plant Ecophysiology, Department of Biology, University of Konstanz, 78457, Konstanz, Germany.

Diatoms possess an impressive capacity for rapidly inducible thermal dissipation of excess absorbed energy (qE), provided by the xanthophyll diatoxanthin and Lhcx proteins. By knocking out the Lhcx1 and Lhcx2 genes individually in Phaeodactylum tricornutum strain 4 and complementing the knockout lines with different Lhcx proteins, multiple mutants with varying qE capacities are obtained, ranging from zero to high values. We demonstrate that qE is entirely dependent on the concerted action of diatoxanthin and Lhcx proteins, with Lhcx1, Lhcx2 and Lhcx3 having similar functions. Moreover, we establish a clear link between Lhcx1/2/3 mediated inducible thermal energy dissipation and a reduction in the functional absorption cross-section of photosystem II. This regulation of the functional absorption cross-section can be tuned by altered Lhcx protein expression in response to environmental conditions. Our results provide a holistic understanding of the rapidly inducible thermal energy dissipation process and its mechanistic implications in diatoms.
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http://dx.doi.org/10.1038/s41467-019-12043-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6744471PMC
September 2019

Phaeodactylum tricornutum.

Trends Genet 2019 09 15;35(9):706-707. Epub 2019 Jul 15.

Institut de Biologie Physico-Chimique, Laboratoire du Biologie du Chloroplaste et Perception de la Lumière chez les Microalgues, UMR7141 CNRS, Sorbonne Université, Paris, France.

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http://dx.doi.org/10.1016/j.tig.2019.05.007DOI Listing
September 2019

bHLH-PAS protein RITMO1 regulates diel biological rhythms in the marine diatom .

Proc Natl Acad Sci U S A 2019 06 6;116(26):13137-13142. Epub 2019 Jun 6.

Laboratory of Computational and Quantitative Biology, Sorbonne Université, CNRS, Institut de Biologie Paris-Seine, F-75005 Paris, France;

Periodic light-dark cycles govern the timing of basic biological processes in organisms inhabiting land as well as the sea, where life evolved. Although prominent marine phytoplanktonic organisms such as diatoms show robust diel rhythms, the mechanisms regulating these processes are still obscure. By characterizing a bHLH-PAS nuclear protein, hereby named RITMO1, we shed light on the regulation of the daily life of diatoms. Alteration of RITMO1 expression levels and timing by ectopic overexpression results in lines with deregulated diurnal gene expression profiles compared with the wild-type cells. Reduced gene expression oscillations are also observed in these lines in continuous darkness, showing that the regulation of rhythmicity by RITMO1 is not directly dependent on light inputs. We also describe strong diurnal rhythms of cellular fluorescence in wild-type cells, which persist in continuous light conditions, indicating the existence of an endogenous circadian clock in diatoms. The altered rhythmicity observed in RITMO1 overexpression lines in continuous light supports the involvement of this protein in circadian rhythm regulation. Phylogenetic analysis reveals a wide distribution of RITMO1-like proteins in the genomes of diatoms as well as in other marine algae, which may indicate a common function in these phototrophs. This study adds elements to our understanding of diatom biology and offers perspectives to elucidate timekeeping mechanisms in marine organisms belonging to a major, but under-investigated, branch of the tree of life.
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http://dx.doi.org/10.1073/pnas.1819660116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6600994PMC
June 2019

Genome editing in diatoms: achievements and goals.

Plant Cell Rep 2018 Oct 23;37(10):1401-1408. Epub 2018 Aug 23.

Laboratoire de Biologie Computationnelle et Quantitative, Institut de Biologie Paris-Seine, Sorbonne Université, CNRS, 75005, Paris, France.

Diatoms are major components of phytoplankton and play a key role in the ecology of aquatic ecosystems. These algae are of great scientific importance for a wide variety of research areas, ranging from marine ecology and oceanography to biotechnology. During the last 20 years, the availability of genomic information on selected diatom species and a substantial progress in genetic manipulation, strongly contributed to establishing diatoms as molecular model organisms for marine biology research. Recently, tailored TALEN endonucleases and the CRISPR/Cas9 system were utilized in diatoms, allowing targeted genetic modifications and the generation of knockout strains. These approaches are extremely valuable for diatom research because breeding, forward genetic screens by random insertion, and chemical mutagenesis are not applicable to the available model species Phaeodactylum tricornutum and Thalassiosira pseudonana, which do not cross sexually in the lab. Here, we provide an overview of the genetic toolbox that is currently available for performing stable genetic modifications in diatoms. We also discuss novel challenges that need to be addressed to fully exploit the potential of these technologies for the characterization of diatom biology and for metabolic engineering.
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http://dx.doi.org/10.1007/s00299-018-2334-1DOI Listing
October 2018

Dynamic Changes between Two LHCX-Related Energy Quenching Sites Control Diatom Photoacclimation.

Plant Physiol 2018 07 17;177(3):953-965. Epub 2018 May 17.

Sorbonne Université, Centre National de la Recherche Scientifique, Institut de Biologie Paris-Seine, Laboratory of Computational and Quantitative Biology, F-75005 Paris, France

Marine diatoms are prominent phytoplankton organisms that perform photosynthesis in extremely variable environments. Diatoms possess a strong ability to dissipate excess absorbed energy as heat via nonphotochemical quenching (NPQ). This process relies on changes in carotenoid pigment composition (xanthophyll cycle) and on specific members of the light-harvesting complex family specialized in photoprotection (LHCXs), which potentially act as NPQ effectors. However, the link between light stress, NPQ, and the existence of different LHCX isoforms is not understood in these organisms. Using picosecond fluorescence analysis, we observed two types of NPQ in the pennate diatom that were dependent on light conditions. Short exposure of low-light-acclimated cells to high light triggers the onset of energy quenching close to the core of photosystem II, while prolonged light stress activates NPQ in the antenna. Biochemical analysis indicated a link between the changes in the NPQ site/mechanism and the induction of different LHCX isoforms, which accumulate either in the antenna complexes or in the core complex. By comparing the responses of wild-type cells and transgenic lines with a reduced expression of the major LHCX isoform, LHCX1, we conclude that core complex-associated NPQ is more effective in photoprotection than is the antenna complex. Overall, our data clarify the complex molecular scenario of light responses in diatoms and provide a rationale for the existence of a degenerate family of LHCX proteins in these algae.
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http://dx.doi.org/10.1104/pp.18.00448DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6053010PMC
July 2018

A systems-wide understanding of photosynthetic acclimation in algae and higher plants.

J Exp Bot 2017 05;68(11):2667-2681

Cluster of Excellence on Plant Sciences (CEPLAS), Institute of Quantitative and Theoretical Biology, Heinrich Heine University Düsseldorf, Germany.

The ability of phototrophs to colonise different environments relies on robust protection against oxidative stress, a critical requirement for the successful evolutionary transition from water to land. Photosynthetic organisms have developed numerous strategies to adapt their photosynthetic apparatus to changing light conditions in order to optimise their photosynthetic yield, which is crucial for life on Earth to exist. Photosynthetic acclimation is an excellent example of the complexity of biological systems, where highly diverse processes, ranging from electron excitation over protein protonation to enzymatic processes coupling ion gradients with biosynthetic activity, interact on drastically different timescales from picoseconds to hours. Efficient functioning of the photosynthetic apparatus and its protection is paramount for efficient downstream processes, including metabolism and growth. Modern experimental techniques can be successfully integrated with theoretical and mathematical models to promote our understanding of underlying mechanisms and principles. This review aims to provide a retrospective analysis of multidisciplinary photosynthetic acclimation research carried out by members of the Marie Curie Initial Training Project, AccliPhot, placing the results in a wider context. The review also highlights the applicability of photosynthetic organisms for industry, particularly with regards to the cultivation of microalgae. It intends to demonstrate how theoretical concepts can successfully complement experimental studies broadening our knowledge of common principles in acclimation processes in photosynthetic organisms, as well as in the field of applied microalgal biotechnology.
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http://dx.doi.org/10.1093/jxb/erx137DOI Listing
May 2017

A role for the cell-wall protein silacidin in cell size of the diatom Thalassiosira pseudonana.

ISME J 2017 11 21;11(11):2452-2464. Epub 2017 Jul 21.

School of Environmental Sciences, University of East Anglia, Norwich Research Park, Norwich, UK.

Diatoms contribute 20% of global primary production and form the basis of many marine food webs. Although their species diversity correlates with broad diversity in cell size, there is also an intraspecific cell-size plasticity owing to sexual reproduction and varying environmental conditions. However, despite the ecological significance of the diatom cell size for food-web structure and global biogeochemical cycles, our knowledge about genes underpinning the size of diatom cells remains elusive. Here, a combination of reverse genetics, experimental evolution and comparative RNA-sequencing analyses enabled us to identify a previously unknown genetic control of cell size in the diatom Thalassiosira pseudonana. In particular, the targeted deregulation of the expression of the cell-wall protein silacidin caused a significant increase in valve diameter. Remarkably, the natural downregulation of the silacidin gene transcript due to experimental evolution under low temperature also correlated with cell-size increase. Our data give first evidence for a genetically controlled regulation of cell size in T. pseudonana and possibly other centric diatoms as they also encode the silacidin gene in their genomes.
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http://dx.doi.org/10.1038/ismej.2017.100DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5649158PMC
November 2017

Investigating mixotrophic metabolism in the model diatom .

Philos Trans R Soc Lond B Biol Sci 2017 Sep;372(1728)

Laboratoire de Physiologie Cellulaire et Végétale, Université Grenoble Alpes (UGA), Centre National de la Recherche Scientifique (CNRS), Commissariat à l'Energie Atomique et aux Energies Alternatives (CEA), Institut National Recherche Agronomique (INRA), Institut de Biosciences et Biotechnologies de Grenoble (BIG), CEA Grenoble, F-38000 Grenoble, France

Diatoms are prominent marine microalgae, interesting not only from an ecological point of view, but also for their possible use in biotechnology applications. They can be cultivated in phototrophic conditions, using sunlight as the sole energy source. Some diatoms, however, can also grow in a mixotrophic mode, wherein both light and external reduced carbon contribute to biomass accumulation. In this study, we investigated the consequences of mixotrophy on the growth and metabolism of the pennate diatom , using glycerol as the source of reduced carbon. Transcriptomics, metabolomics, metabolic modelling and physiological data combine to indicate that glycerol affects the central-carbon, carbon-storage and lipid metabolism of the diatom. In particular, provision of glycerol mimics typical responses of nitrogen limitation on lipid metabolism at the level of triacylglycerol accumulation and fatty acid composition. The presence of glycerol, despite provoking features reminiscent of nutrient limitation, neither diminishes photosynthetic activity nor cell growth, revealing essential aspects of the metabolic flexibility of these microalgae and suggesting possible biotechnological applications of mixotrophy.This article is part of the themed issue 'The peculiar carbon metabolism in diatoms'.
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http://dx.doi.org/10.1098/rstb.2016.0404DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5516113PMC
September 2017

Light sensing and responses in marine microalgae.

Curr Opin Plant Biol 2017 06 26;37:70-77. Epub 2017 Apr 26.

Sorbonne Universités, UPMC, Institut de Biologie Paris-Seine, CNRS, Laboratoire de Biologie Computationnelle et Quantitative, 4, Place de Jussieu, 75005 Paris, France. Electronic address:

Marine 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.
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http://dx.doi.org/10.1016/j.pbi.2017.03.005DOI Listing
June 2017

Evolutionary genomics of the cold-adapted diatom Fragilariopsis cylindrus.

Nature 2017 01 16;541(7638):536-540. Epub 2017 Jan 16.

Department of Energy Joint Genome Institute, 2800 Mitchell Drive, Walnut Creek, California 94598, USA.

The Southern Ocean houses a diverse and productive community of organisms. Unicellular eukaryotic diatoms are the main primary producers in this environment, where photosynthesis is limited by low concentrations of dissolved iron and large seasonal fluctuations in light, temperature and the extent of sea ice. How diatoms have adapted to this extreme environment is largely unknown. Here we present insights into the genome evolution of a cold-adapted diatom from the Southern Ocean, Fragilariopsis cylindrus, based on a comparison with temperate diatoms. We find that approximately 24.7 per cent of the diploid F. cylindrus genome consists of genetic loci with alleles that are highly divergent (15.1 megabases of the total genome size of 61.1 megabases). These divergent alleles were differentially expressed across environmental conditions, including darkness, low iron, freezing, elevated temperature and increased CO. Alleles with the largest ratio of non-synonymous to synonymous nucleotide substitutions also show the most pronounced condition-dependent expression, suggesting a correlation between diversifying selection and allelic differentiation. Divergent alleles may be involved in adaptation to environmental fluctuations in the Southern Ocean.
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http://dx.doi.org/10.1038/nature20803DOI Listing
January 2017

The diatom Phaeodactylum tricornutum adjusts nonphotochemical fluorescence quenching capacity in response to dynamic light via fine-tuned Lhcx and xanthophyll cycle pigment synthesis.

New Phytol 2017 Apr 21;214(1):205-218. Epub 2016 Nov 21.

UMR7266 'LIENSs', CNRS Université de La Rochelle, Institut du Littoral et de l'Environnement, 2 rue Olympe de Gouges, La Rochelle, 17000, France.

Diatoms contain a highly flexible capacity to dissipate excessively absorbed light by nonphotochemical fluorescence quenching (NPQ) based on the light-induced conversion of diadinoxanthin (Dd) into diatoxanthin (Dt) and the presence of Lhcx proteins. Their NPQ fine regulation on the molecular level upon a shift to dynamic light conditions is unknown. We investigated the regulation of Dd + Dt amount, Lhcx gene and protein synthesis and NPQ capacity in the diatom Phaeodactylum tricornutum after a change from continuous low light to 3 d of sine (SL) or fluctuating (FL) light conditions. Four P. tricornutum strains with different NPQ capacities due to different expression of Lhcx1 were included. All strains responded to dynamic light comparably, independently of initial NPQ capacity. During SL, NPQ capacity was strongly enhanced due to a gradual increase of Lhcx2 and Dd + Dt amount. During FL, cells enhanced their NPQ capacity on the first day due to increased Dd + Dt, Lhcx2 and Lhcx3; already by the second day light acclimation was accomplished. While quenching efficiency of Dt was strongly lowered during SL conditions, it remained high throughout the whole FL exposure. Our results highlight a more balanced and cost-effective photoacclimation strategy of P. tricornutum under FL than under SL conditions.
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http://dx.doi.org/10.1111/nph.14337DOI Listing
April 2017

Regulation of chain length in two diatoms as a growth-fragmentation process.

Phys Rev E 2016 Aug 23;94(2-1):022418. Epub 2016 Aug 23.

Sorbonne Universités, UPMC Université Paris 6, UMR 7238, Computational and Quantitative Biology, 15 rue de l'École de Médecine, Paris, France.

Chain formation in diatoms is relevant because of several aspects of their adaptation to the ecosystem. However, the tools to quantify the regulation of their assemblage and infer specific mechanisms in a laboratory setting are scarce. To address this problem, we define an approach based on a statistical physics model of chain growth and separation in combination with experimental evaluation of chain-length distributions. Applying this combined analysis to data from Chaetoceros decipiens and Phaeodactylum tricornutum, we find that cells of the first species control chain separation, likely through a cell-to-cell communication process, while the second species only modulates the separation rate. These results promote quantitative methods for characterizing chain formation in several chain-forming species and in diatoms in particular.
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http://dx.doi.org/10.1103/PhysRevE.94.022418DOI Listing
August 2016

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

J Exp Bot 2016 06 25;67(13):3939-51. Epub 2016 May 25.

Sorbonne Universités, UPMC, Institut de Biologie Paris-Seine, CNRS, Laboratoire de Biologie Computationnelle et Quantitative, 15 rue de l'Ecole de Médecine, 75006 Paris, France

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.
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http://dx.doi.org/10.1093/jxb/erw198DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4915529PMC
June 2016

Diatom Phytochromes Reveal the Existence of Far-Red-Light-Based Sensing in the Ocean.

Plant Cell 2016 Mar 3;28(3):616-28. Epub 2016 Mar 3.

Sorbonne Universités, UPMC, Institut de Biologie Paris-Seine, CNRS, Laboratoire de Biologie Computationnelle et Quantitative UMR 7238, 75006 Paris, France

The absorption of visible light in aquatic environments has led to the common assumption that aquatic organisms sense and adapt to penetrative blue/green light wavelengths but show little or no response to the more attenuated red/far-red wavelengths. Here, we show that two marine diatom species, Phaeodactylum tricornutum and Thalassiosira pseudonana, possess a bona fide red/far-red light sensing phytochrome (DPH) that uses biliverdin as a chromophore and displays accentuated red-shifted absorbance peaks compared with other characterized plant and algal phytochromes. Exposure to both red and far-red light causes changes in gene expression in P. tricornutum, and the responses to far-red light disappear in DPH knockout cells, demonstrating that P. tricornutum DPH mediates far-red light signaling. The identification of DPH genes in diverse diatom species widely distributed along the water column further emphasizes the ecological significance of far-red light sensing, raising questions about the sources of far-red light. Our analyses indicate that, although far-red wavelengths from sunlight are only detectable at the ocean surface, chlorophyll fluorescence and Raman scattering can generate red/far-red photons in deeper layers. This study opens up novel perspectives on phytochrome-mediated far-red light signaling in the ocean and on the light sensing and adaptive capabilities of marine phototrophs.
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http://dx.doi.org/10.1105/tpc.15.00928DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4826011PMC
March 2016

Characterization of marine diatom-infecting virus promoters in the model diatom Phaeodactylum tricornutum.

Sci Rep 2015 Dec 22;5:18708. Epub 2015 Dec 22.

Laboratory of Aquatic Environmental Science (LAQUES), Faculty of Agriculture, Kochi University, Otsu-200, Monobe, Nankoku, Kochi 783-8502, Japan.

Viruses are considered key players in phytoplankton population control in oceans. However, mechanisms that control viral gene expression in prominent microalgae such as diatoms remain largely unknown. In this study, potential promoter regions isolated from several marine diatom-infecting viruses (DIVs) were linked to the egfp reporter gene and transformed into the Pennales diatom Phaeodactylum tricornutum. We analysed their activity in cells grown under different conditions. Compared to diatom endogenous promoters, novel DIV promoter (ClP1) mediated a significantly higher degree of reporter transcription and translation. Stable expression levels were observed in transformants grown under both light and dark conditions, and high levels of expression were reported in cells in the stationary phase compared to the exponential phase of growth. Conserved motifs in the sequence of DIV promoters were also found. These results allow the identification of novel regulatory regions that drive DIV gene expression and further examinations of the mechanisms that control virus-mediated bloom control in diatoms. Moreover, the identified ClP1 promoter can serve as a novel tool for metabolic engineering of diatoms. This is the first report describing a promoter of DIVs that may be of use in basic and applied diatom research.
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http://dx.doi.org/10.1038/srep18708DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4686930PMC
December 2015

The upstream regulatory sequence of the light harvesting complex Lhcf2 gene of the marine diatom Phaeodactylum tricornutum enhances transcription in an orientation- and distance-independent fashion.

Mar Genomics 2015 Dec 25;24 Pt 1:69-79. Epub 2015 Jun 25.

Sorbonne Universités, UPMC Univ Paris 06, Institut de Biologie Paris-Seine, UMR 7238, F-75006 Paris, France; CNRS, UMR 7238, F-75006 Paris, France. Electronic address:

Diatoms are a key phytoplankton group in the contemporary ocean, showing extraordinary adaptation capacities to rapidly changing environments. The recent availability of whole genome sequences from representative species has revealed distinct features in their genomes, like novel combinations of genes encoding distinct metabolisms and a significant number of diatom-specific genes. However, the regulatory mechanisms driving diatom gene expression are still largely uncharacterized. Considering the wide variety of fields of study orbiting diatoms, ranging from ecology, evolutionary biology to biotechnology, it is thus essential to increase our understanding of fundamental gene regulatory processes such as transcriptional regulation. To this aim, we explored the functional properties of the 5'-flanking region of the Phaeodatylum tricornutum Lhcf2 gene, encoding a member of the Light Harvesting Complex superfamily and we showed that this region enhances transcription of a GUS reporter gene in an orientation- and distance-independent fashion. This represents the first example of a cis-regulatory sequence with enhancer-like features discovered in diatoms and it is instrumental for the generation of novel genetic tools and diatom exploitation in different areas of study.
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http://dx.doi.org/10.1016/j.margen.2015.06.010DOI Listing
December 2015

The diversity of small non-coding RNAs in the diatom Phaeodactylum tricornutum.

BMC Genomics 2014 Aug 20;15:698. Epub 2014 Aug 20.

Sorbonne Universités, UPMC Univ Paris 06, UMR 7238, Laboratory of Computational and Quantitative Biology, F-75006 Paris, France.

Background: Marine diatoms constitute a major component of eukaryotic phytoplankton and stand at the crossroads of several evolutionary lineages. These microalgae possess peculiar genomic features and novel combinations of genes acquired from bacterial, animal and plant ancestors. Furthermore, they display both DNA methylation and gene silencing activities. Yet, the biogenesis and regulatory function of small RNAs (sRNAs) remain ill defined in diatoms.

Results: Here we report the first comprehensive characterization of the sRNA landscape and its correlation with genomic and epigenomic information in Phaeodactylum tricornutum. The majority of sRNAs is 25 to 30 nt-long and maps to repetitive and silenced Transposable Elements marked by DNA methylation. A subset of this population also targets DNA methylated protein-coding genes, suggesting that gene body methylation might be sRNA-driven in diatoms. Remarkably, 25-30 nt sRNAs display a well-defined and unprecedented 180 nt-long periodic distribution at several highly methylated regions that awaits characterization. While canonical miRNAs are not detectable, other 21-25 nt sRNAs of unknown origin are highly expressed. Besides, non-coding RNAs with well-described function, namely tRNAs and U2 snRNA, constitute a major source of 21-25 nt sRNAs and likely play important roles under stressful environmental conditions.

Conclusions: P. tricornutum has evolved diversified sRNA pathways, likely implicated in the regulation of largely still uncharacterized genetic and epigenetic processes. These results uncover an unexpected complexity of diatom sRNA population and previously unappreciated features, providing new insights into the diversification of sRNA-based processes in eukaryotes.
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http://dx.doi.org/10.1186/1471-2164-15-698DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4247016PMC
August 2014

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

J Plant Physiol 2015 Jan 7;172:42-54. Epub 2014 Jul 7.

Sorbonne Universités, UPMC Univ Paris 06, UMR 7238, Computational and Quantitative Biology, F-75006 Paris, France; CNRS, UMR 7238, Computational and Quantitative Biology, F-75006 Paris, France. Electronic address:

Light 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.
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http://dx.doi.org/10.1016/j.jplph.2014.06.011DOI Listing
January 2015

The molecular life of diatoms.

Mar Genomics 2014 Aug 12;16:1-3. Epub 2014 Jul 12.

Sorbonne Universités, UPMC Univ Paris 06, CNRS UMR 7238, Laboratory of Computational and Quantitative Biology, F-75006 Paris, France.

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http://dx.doi.org/10.1016/j.margen.2014.07.002DOI Listing
August 2014

It's about time: rhythms as a new dimension of molecular marine research.

Mar Genomics 2014 Apr;14:1-2

Sorbonne Universités, UPMC Univ Paris 06, Laboratory of Computational and Quantitative Biology UMR 7238CNRS-UPMC, F-75006 Paris, France. Electronic address:

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http://dx.doi.org/10.1016/j.margen.2014.04.001DOI Listing
April 2014

Genome engineering empowers the diatom Phaeodactylum tricornutum for biotechnology.

Nat Commun 2014 May 29;5:3831. Epub 2014 May 29.

Cellectis S.A., 8 rue de la Croix de Jarry, 75013 Paris, France.

Diatoms, a major group of photosynthetic microalgae, have a high biotechnological potential that has not been fully exploited because of the paucity of available genetic tools. Here we demonstrate targeted and stable modifications of the genome of the marine diatom Phaeodactylum tricornutum, using both meganucleases and TALE nucleases. When nuclease-encoding constructs are co-transformed with a selectable marker, high frequencies of genome modifications are readily attained with 56 and 27% of the colonies exhibiting targeted mutagenesis or targeted gene insertion, respectively. The generation of an enhanced lipid-producing strain (45-fold increase in triacylglycerol accumulation) through the disruption of the UDP-glucose pyrophosphorylase gene exemplifies the power of genome engineering to harness diatoms for biofuel production.
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http://dx.doi.org/10.1038/ncomms4831DOI Listing
May 2014

The Cryptochrome/Photolyase Family in aquatic organisms.

Mar Genomics 2014 Apr 23;14:23-37. Epub 2014 Feb 23.

Sorbonne Universités, UPMC Univ Paris 06, UMR 7238, Computational and Quantitative Biology, F-75006 Paris, France; CNRS, UMR 7238, Computational and Quantitative Biology, F-75006 Paris, France.

The Cryptochrome/Photolyase Family (CPF) represents an ancient group of widely distributed UV-A/blue-light sensitive proteins sharing common structures and chromophores. During the course of evolution, different CPFs acquired distinct functions in DNA repair, light perception and circadian clock regulation. Previous phylogenetic analyses of the CPF have allowed reconstruction of the evolution and distribution of the different CPF super-classes in the tree of life. However, so far only limited information is available from the CPF orthologs in aquatic organisms that evolved in environments harboring great diversity of life forms and showing peculiar light distribution and rhythms. To gain new insights into the evolutionary and functional relationships within the CPF family, we performed a detailed study of CPF members from marine (diatoms, sea urchin and annelid) and freshwater organisms (teleost) that populate diverse habitats and exhibit different life strategies. In particular, we first extended the CPF family phylogeny by including genes from aquatic organisms representative of several branches of the tree of life. Our analysis identifies four major super-classes of CPF proteins and importantly singles out the presence of a plant-like CRY in diatoms and in metazoans. Moreover, we show a dynamic evolution of Cpf genes in eukaryotes with various events of gene duplication coupled to functional diversification and gene loss, which have shaped the complex array of Cpf genes in extant aquatic organisms. Second, we uncover clear rhythmic diurnal expression patterns and light-dependent regulation for the majority of the analyzed Cpf genes in our reference species. Our analyses reconstruct the molecular evolution of the CPF family in eukaryotes and provide a solid foundation for a systematic characterization of novel light activated proteins in aquatic environments.
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http://dx.doi.org/10.1016/j.margen.2014.02.001DOI Listing
April 2014

AUREOCHROME1a-mediated induction of the diatom-specific cyclin dsCYC2 controls the onset of cell division in diatoms (Phaeodactylum tricornutum).

Plant Cell 2013 Jan 4;25(1):215-28. Epub 2013 Jan 4.

Protistology and Aquatic Ecology, Department of Biology, Ghent University, B-9000 Gent, Belgium.

Cell division in photosynthetic organisms is tightly regulated by light. Although the light dependency of the onset of the cell cycle has been well characterized in various phototrophs, little is known about the cellular signaling cascades connecting light perception to cell cycle activation and progression. Here, we demonstrate that diatom-specific cyclin 2 (dsCYC2) in Phaeodactylum tricornutum displays a transcriptional peak within 15 min after light exposure, long before the onset of cell division. The product of dsCYC2 binds to the cyclin-dependent kinase CDKA1 and can complement G1 cyclin-deficient yeast. Consistent with the role of dsCYC2 in controlling a G1-to-S light-dependent cell cycle checkpoint, dsCYC2 silencing decreases the rate of cell division in diatoms exposed to light-dark cycles but not to constant light. Transcriptional induction of dsCYC2 is triggered by blue light in a fluence rate-dependent manner. Consistent with this, dsCYC2 is a transcriptional target of the blue light sensor AUREOCHROME1a, which functions synergistically with the basic leucine zipper (bZIP) transcription factor bZIP10 to induce dsCYC2 transcription. The functional characterization of a cyclin whose transcription is controlled by light and whose activity connects light signaling to cell cycle progression contributes significantly to our understanding of the molecular mechanisms underlying light-dependent cell cycle onset in diatoms.
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http://dx.doi.org/10.1105/tpc.112.106377DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3584536PMC
January 2013

High light acclimation in the secondary plastids containing diatom Phaeodactylum tricornutum is triggered by the redox state of the plastoquinone pool.

Plant Physiol 2013 Feb 3;161(2):853-65. Epub 2012 Dec 3.

Littoral Environnement et Sociétés, Unité Mixte de Recherche 7266, Centre National de la Recherche Scientifique-University of La Rochelle, Institute for Coastal and Environmental Research, 17000 La Rochelle, France.

In diatoms, the process of energy-dependent chlorophyll fluorescence quenching (qE) has an important role in photoprotection. Three components are essential for qE: (1) the light-dependent generation of a transthylakoidal proton gradient; (2) the deepoxidation of the xanthophyll diadinoxanthin (Dd) into diatoxanthin (Dt); and (3) specific nucleus-encoded antenna proteins, called Light Harvesting Complex Protein X (LHCX). We used the model diatom Phaeodactylum tricornutum to investigate the concerted light acclimation response of the qE key components LHCX, proton gradient, and xanthophyll cycle pigments (Dd+Dt) and to identify the intracellular light-responsive trigger. At high-light exposure, the up-regulation of three of the LHCX genes and the de novo synthesis of Dd+Dt led to a pronounced rise of qE. By inhibiting either the conversion of Dd to Dt or the translation of LHCX genes, qE amplification was abolished and the diatom cells suffered from stronger photoinhibition. Artificial modification of the redox state of the plastoquinone (PQ) pool via 3-(3,4-dichlorophenyl)-1,1-dimethylurea and 5-dibromo-6-isopropyl-3-methyl-1,4-benzoquinone resulted in a disturbance of Dd+Dt synthesis in an opposite way. Moreover, we could increase the transcription of two of the four LHCX genes under low-light conditions by reducing the PQ pool using 5-dibromo-6-isopropyl-3-methyl-1,4-benzoquinone. Altogether, our results underline the central role of the redox state of the PQ pool in the light acclimation of diatoms. Additionally, they emphasize strong evidence for the existence of a plastid-to-nucleus retrograde signaling mechanism in an organism with plastids that derived from secondary endosymbiosis.
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http://dx.doi.org/10.1104/pp.112.207811DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3561024PMC
February 2013

Exploring the molecular basis of responses to light in marine diatoms.

J Exp Bot 2012 Feb 10;63(4):1575-91. Epub 2012 Feb 10.

Université Pierre et Marie Curie, Paris 06, Centre National de la Recherche Scientifique, UMR7238, Laboratoire de Génomique des Microorganismes, 75006 Paris, France.

Light is an essential source of energy for life on Earth and is one of the most important signals that organisms use to obtain information from the surrounding environment, on land and in the oceans. Prominent marine microalgae, such as diatoms, display a suite of sophisticated responses (physiological, biochemical, and behavioural) to optimize their photosynthesis and growth under changing light conditions. However, the molecular mechanisms controlling diatom responses to light are still largely unknown. Recent progress in marine diatom genomics and genetics, combined with well-established (eco) physiological and biophysical approaches, now offers novel opportunities to address these issues. This review provides a description of the molecular components identified in diatom genomes that are involved in light perception and acclimation mechanisms. How the initial functional characterizations of specific light regulators provide the basis to investigate the conservation or diversification of light-mediated processes in diatoms is also discussed. Hypotheses on the role of the identified factors in determining the growth, distribution, and adaptation of diatoms in different marine environments are reported.
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http://dx.doi.org/10.1093/jxb/ers005DOI Listing
February 2012

Diatom cell division in an environmental context.

Curr Opin Plant Biol 2010 Dec 20;13(6):623-30. Epub 2010 Oct 20.

Environmental and Evolutionary Genomics Section, Institut de Biologie de l'Ecole Normale Supérieure, Centre National de la Recherche Scientifique UMR8197 INSERM U1024, F-75005 Paris, France.

Studies of cell division in organisms derived from secondary endosymbiosis such as diatoms have revealed that the mechanisms are far from those found in more conventional model eukaryotes. An atypical acentriolar microtuble-organizing centre, centripetal cytokinesis combined with centrifugal cell wall neosynthesis, and the role of sex in relation to cell size restoration make diatoms an exciting system to re-investigate the evolution, differentiation and regulation of cell division. Such studies are further justified considering the ecological relevance of these microalgae in contemporary oceans and the need to understand the mechanisms controlling their growth and distribution in an environmental context. Recent work derived from genome-wide analyses on representative model diatoms reveals that the cell cycle is finely tuned to inputs derived from both endogenous and environmental signals.
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http://dx.doi.org/10.1016/j.pbi.2010.09.014DOI Listing
December 2010

An atypical member of the light-harvesting complex stress-related protein family modulates diatom responses to light.

Proc Natl Acad Sci U S A 2010 Oct 4;107(42):18214-9. Epub 2010 Oct 4.

Physiologie Membranaire et Moléculaire du Chloroplaste, Université Pierre et Marie Curie, Centre National de la Recherche Scientifique UMR7141, 75005 Paris, France.

Diatoms are prominent phytoplanktonic organisms that contribute around 40% of carbon assimilation in the oceans. They grow and perform optimally in variable environments, being able to cope with unpredictable changes in the amount and quality of light. The molecular mechanisms regulating diatom light responses are, however, still obscure. Using knockdown Phaeodactylum tricornutum transgenic lines, we reveal the key function of a member of the light-harvesting complex stress-related (LHCSR) protein family, denoted LHCX1, in modulation of excess light energy dissipation. In contrast to green algae, this gene is already maximally expressed in nonstressful light conditions and encodes a protein required for efficient light responses and growth. LHCX1 also influences natural variability in photoresponse, as evidenced in ecotypes isolated from different latitudes that display different LHCX1 protein levels. We conclude, therefore, that this gene plays a pivotal role in managing light responses in diatoms.
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http://dx.doi.org/10.1073/pnas.1007703107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2964204PMC
October 2010