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Benthic surveys are a key component of monitoring and conservation efforts for coral reefs worldwide. While traditional image-based surveys rely on manual annotation of photographs to characterise benthic composition, automatic image annotation based on computer vision is becoming increasingly common. However, accurate classification of some benthic groups from reflectance images presents a challenge to local ecologists and computers alike. Most coral reef organisms produce one or a combination of fluorescent pigments, such as Green Fluorescent Protein (GFP)-like proteins found in corals, chlorophyll-a found in all photosynthetic organisms, and phycobiliproteins found in red macroalgae, crustose coralline algae (CCA) and cyanobacteria. Building on the potential of these pigments as a target for automatic image annotation, we developed a novel imaging method based on off-the-shelf components to improve classification of coral and other biotic substrates using a multi-excitation fluorescence (MEF) imaging system. We used RGB cameras to image the fluorescence emission of coral and algal pigments stimulated by narrow-waveband blue and green light, and then combined the information into three-channel pseudocolour images. Using a set of a priori rules defined by the relative pixel intensity produced in different channels, the method achieved successful classification of organisms into three categories based on the dominant fluorescent pigment expressed, facilitating discrimination of traditionally problematic groups. This work provides a conceptual foundation for future technological developments that will improve the cost, accuracy and speed of coral reef surveys.

Pigments homologous to the Green Fluorescent Protein (GFP) have been proposed to fine-tune the internal light microclimate of corals, facilitating photoacclimation of photosynthetic coral symbionts (Symbiodiniaceae) to life in different reef habitats and environmental conditions. However, direct measurements of the in vivo light conditions inside the coral tissue supporting this conclusion are lacking. Here, we quantified the intra-tissue spectral light environment of corals expressing GFP-like proteins from widely different light regimes. We focus on (1) photoconvertible red fluorescent proteins (pcRFPs), thought to enhance photosynthesis in mesophotic habitats via wavelength conversion, and (2) chromoproteins (CPs), which provide photoprotection to the symbionts in shallow water via light absorption. Optical microsensor measurements indicated that both pigment groups strongly alter the coral tissue light environment. Estimates derived from light spectra measured in pcRFP-containing corals showed that fluorescence emission can contribute to >50% of orange-red light available to the photosynthetic symbionts at mesophotic depths. We further show that upregulation of pink CPs in shallow-water corals during bleaching leads to a reduction of orange light by 10-20% compared to low-CP tissue. Thus, screening by CPs has an important role in mitigating the light-enhancing effect of coral tissue scattering during bleaching. Our results provide the first experimental quantification of the importance of GFP-like proteins in fine-tuning the light microclimate of corals during photoacclimation.

Glycogen and starch are the major carbon and energy reserve polysaccharides in nature, providing living organisms with a survival advantage. The evolution of the enzymatic machinery responsible for the biosynthesis and degradation of such polysaccharides, led the development of mechanisms to control the assembly and disassembly rate, to store and recover glucose according to cell energy demands. The tetrameric enzyme ADP-glucose pyrophosphorylase (AGPase) catalyzes and regulates the initial step in the biosynthesis of both α-polyglucans. AGPase displays cooperativity and allosteric regulation by sensing metabolites from the cell energy flux. The understanding of the allosteric signal transduction mechanisms in AGPase arises as a long-standing challenge. In this work, we disclose the cryoEM structures of the paradigmatic homotetrameric AGPase from Escherichia coli (EcAGPase), in complex with either positive or negative physiological allosteric regulators, fructose-1,6-bisphosphate (FBP) and AMP respectively, both at 3.0 Å resolution. Strikingly, the structures reveal that FBP binds deeply into the allosteric cleft and overlaps the AMP site. As a consequence, FBP promotes a concerted conformational switch of a regulatory loop, RL2, from a “locked” to a “free” state, modulating ATP binding and activating the enzyme. This notion is strongly supported by our complementary biophysical and bioinformatics evidence, and a careful analysis of vast enzyme kinetics data on single-point mutants of EcAGPase. The cryoEM structures uncover the residue interaction networks (RIN) between the allosteric and the catalytic components of the enzyme, providing unique details on how the signaling information is transmitted across the tetramer, from which cooperativity emerges. Altogether, the conformational states visualized by cryoEM reveal the regulatory mechanism of EcAGPase, laying the foundations to understand the allosteric control of bacterial glycogen biosynthesis at the molecular level of detail., Graphical abstract Image 1, Highlights • CryoEM structures of AGPase from E. coli were solved in complex with FBP and AMP. • FBP binds deeply into the allosteric cleft and overlaps the AMP site. • Allosteric regulators promote a conformational switch of the RL2 loop. • FBP modulates ATP binding and activates the enzyme. • A mechanism of allosteric regulation for bacterial AGPases is proposed.

Climate change-induced global warming threatens the survival of key ecosystems including shallow water coral reefs. Elevated temperatures can disrupt the normal physiological functioning of photosynthetic organisms by altering the fluidity and permeability of chloroplast membranes that is defined and regulated by their lipid composition. Since the habitat-forming reef corals rely on the obligatory symbiosis with dinoflagellates of the family Symbiodiniaceae, their heat stress response can be expected to be strongly influenced by the symbiont's lipid metabolism. However, in contrast to the steady increase in the knowledge of the functioning of coral symbionts at the genomic and transcriptomic level, the understanding of their membrane lipid composition and regulation in response to temperature stress is lagging behind. We have utilised mass spectrometry-based lipidomic analyses to identify the key polar lipids that form the biological membranes of reef coral symbionts, comparing the thermotolerant species Durusdinium trenchii with the thermosensitive taxon Cladocopium C3, both hosted by Acropora valida. Our results indicate that the superior thermotolerance D. trenchii inside the host corals could be achieved through (1) the amount and saturation of sulfoquinovosyldiacylglycerols, in particular through putative photosystem II interactions, (2) the increased digalactosyldiacylglycerol to monogalactosyldiacylglycerol ratio with the potential to stabilise thylakoid membranes and integrated proteins, and (3) the chaperone-like function of lyso-lipids. Thereby, our study provides novel insights into the heat tolerance of coral symbionts, contributing to the understanding of the potential of coral reef ecosystems to respond and adjust to heat stress events that are becoming more frequent due to climate change. Finally, our identification of multiple mechanisms of heat tolerance in Symbiodiniaceae furthers the knowledge of the general stress physiology of photosynthetic organisms.

Boron isotopic and elemental analysis of coral aragonite can give important insights into the calcification strategies employed in coral skeletal construction. Traditional methods of analysis have limited spatial (and thus temporal) resolution, hindering attempts to unravel skeletal heterogeneity. Laser ablation mass spectrometry allows a much more refined view, and here we employ these techniques to explore boron isotope and co-varying elemental ratios in the tropical coral Siderastrea siderea. We generate two-dimensional maps of the carbonate parameters within the calcification medium that deposited the skeleton, which reveal large heterogeneities in carbonate chemistry across the macro-structure of a coral polyp. These differences have the potential to bias proxy interpretations, and indicate that different processes facilitated precipitation of different parts of the coral skeleton: the low-density columella being precipitated from a fluid with a carbonate composition closer to seawater, compared to the high-density inter-polyp walls where aragonite saturation was ~ 5 times that of external seawater. Therefore, the skeleton does not precipitate from a spatially homogeneous fluid and its different parts may thus have varying sensitivity to environmental stress. This offers new insights into the mechanisms behind the response of the S. siderea skeletal phenotype to ocean acidification.

1. The increasing illumination of our world by artificial light at night (ALAN) has created a new field of global change research with impacts now being demonstrated across taxa, biological ranks and spatial scales. Following advances in terrestrial ecology, marine ALAN has become a rapidly growing research area attracting scientists from across all biomes. Technological limitations, complexities of researching many coastal and marine ecosystems and the interdisciplinary nature of ALAN research present numerous challenges. 2. Drawing on expertise from optical oceanographers, modellers, community ecologists, experimental and molecular biologists, we share practical advice and solutions that have proven useful for marine ALAN research. Discussing lessons learnt early on can help in the effective and efficient development of a field. 3. The guide follows a sensory ecology approach to marine light pollution and consolidates physics, ecology and biology. First, we introduce marine lightscapes highlighting how these differ from terrestrial ones and provide an overview of biological adaptations to them. Second, we discuss study design and technology to best quantify ALAN exposure of and impacts on marine and coastal organisms including molecular tools and approaches to scale-up marine ALAN research. 4. We conclude that the growing field of marine ALAN research presents opportunities not only for improving our understanding of this globally widespread stressor, but also for advancing fundamental marine photobiology, chronobiology and night-time ecology. Interdisciplinary research will be essential to gain insights into natural marine lightscapes shaping the ecology and evolution coastal and marine ecosystems.

The phosphatidyl-myo-inositol mannosyltransferase A (PimA) is an essential peripheral membrane glycosyltransferase that initiates the biosynthetic pathway of phosphatidyl-myo-inositol mannosides (PIMs), key structural elements and virulence factors of Mycobacterium tuberculosis. PimA undergoes functionally important conformational changes, including (i) α-helix-to-β-strand and β-strand-to-α-helix transitions and (ii) an “open-to-closed” motion between the two Rossmann-fold domains, a conformational change that is necessary to generate a catalytically competent active site. In previous work, we established that GDP-Man and GDP stabilize the enzyme and facilitate the switch to a more compact active state. To determine the structural contribution of the mannose ring in such an activation mechanism, we analyzed a series of chemical derivatives, including mannose phosphate (Man-P) and mannose pyrophosphate-ribose (Man-PP-RIB), and additional GDP derivatives, such as pyrophosphate ribose (PP-RIB) and GMP, by the combined use of X-ray crystallography, limited proteolysis, circular dichroism, isothermal titration calorimetry, and small angle X-ray scattering methods. Although the β-phosphate is present, we found that the mannose ring, covalently attached to neither phosphate (Man-P) nor PP-RIB (Man-PP-RIB), does promote the switch to the active compact form of the enzyme. Therefore, the nucleotide moiety of GDP-Man, and not the sugar ring, facilitates the “open-to-closed” motion, with the β-phosphate group providing the high-affinity binding to PimA. Altogether, the experimental data contribute to a better understanding of the structural determinants involved in the “open-to-closed” motion not only observed in PimA but also visualized and/or predicted in other glycosyltransfeases. In addition, the experimental data might prove to be useful for the discovery and/or development of PimA and/or glycosyltransferase inhibitors., This work was supported by grants from the European Commission, New Medicine for Tuberculosis, Contract LSHP-CT-2005-018923, and More Medicines for Tuberculosis, Contract HEALTH-F3-2011-260872 (M.E.G.); MARIE CURIE Reintegration Contract 844905 (B.T.); MINECO/FEDER EU Contracts SAF2010-19096, BIO2013-49022-C2-2-R, and BFU2016-77427-C2-2-R; Severo Ochoa Excellence Accreditation SEV-2016-0644 (M.E.G.); MINECO/FEDER EU Contract PID2019-104090RB-100 (P.M.); and “Juan de la Cierva Postdoctoral program” Contract IJCI-2014-19206 (B.T.).

Summary Coral bleaching, caused by the loss of brownish-colored dinoflagellate photosymbionts from the host tissue of reef-building corals, is a major threat to reef survival. Occasionally, bleached corals become exceptionally colorful rather than white. These colors derive from photoprotective green fluorescent protein (GFP)-like pigments produced by the coral host. There is currently no consensus regarding what causes colorful bleaching events and what the consequences for the corals are. Here, we document that colorful bleaching events are a recurring phenomenon in reef regions around the globe. Our analysis of temperature conditions associated with colorful bleaching events suggests that corals develop extreme coloration within 2 to 3 weeks after exposure to mild or temporary heat stress. We demonstrate that the increase of light fluxes in symbiont-depleted tissue promoted by reflection of the incident light from the coral skeleton induces strong expression of the photoprotective coral host pigments. We describe an optical feedback loop involving both partners of the association, discussing that the mitigation of light stress offered by host pigments could facilitate recolonization of bleached tissue by symbionts. Our data indicate that colorful bleaching has the potential to identify local environmental factors, such as nutrient stress, that can exacerbate the impact of elevated temperatures on corals, to indicate the severity of heat stress experienced by corals and to gauge their post-stress recovery potential. Video Abstract Download : Download video (160MB)

Despite a very strong El Nino Southern Oscillation in 2015/2016, no coral mortality associated with bleaching was observed at the northern Galapagos (Ecuador) Islands of Darwin and Wolf. From March 2016 to March 2018, coral cover and health as well as water chemistry (NO3− and PO43−) and temperature were recorded. A marked heat anomaly reached 30 °C at Wolf in February 2016, but peak temperatures were attenuated after 2 d by a 4 °C drop. Temperature patterns at three depths (10, 15, and 20 m) and a subsequent and persistent phytoplankton bloom suggest topographically driven upwelling as the source of colder water and dissolved inorganic nutrients—both of which helped corals endure the heating episode. Consequently, no mortality and only partial bleaching were recorded in March 2016. Partially bleached corals contained numerous healthy zooxanthellae in deeper tissue layers. A continuous temperature record from 2012 to 2014 suggests that such upwelling events are common, inducing temperature fluctuations of up to 6 °C within 24 h during the observation period. Events at Wolf in 2016 suggest local upwelling reduced coral stress by relieving heat and by delivering nutrients required by corals to retain their regular temperature tolerance.

The evolution of metabolic pathways is a major force behind natural selection. In the spotlight of such process lies the structural evolution of the enzymatic machinery responsible for the central energy metabolism. Specifically, glycogen metabolism has emerged to allow organisms to save available environmental surplus of carbon and energy, using dedicated glucose polymers as a storage compartment that can be mobilized at future demand. The origins of such adaptive advantage rely on the acquisition of an enzymatic system for the biosynthesis and degradation of glycogen, along with mechanisms to balance the assembly and disassembly rate of this polysaccharide, in order to store and recover glucose according to cell energy needs. The first step in the classical bacterial glycogen biosynthetic pathway is carried out by the adenosine 5′-diphosphate (ADP)-glucose pyrophosphorylase. This allosteric enzyme synthesizes ADP-glucose and acts as a point of regulation. The second step is carried out by the glycogen synthase, an enzyme that generates linear α-(1→4)-linked glucose chains, whereas the third step catalyzed by the branching enzyme produces α-(1→6)-linked glucan branches in the polymer. Two enzymes facilitate glycogen degradation: glycogen phosphorylase, which functions as an α-(1→4)-depolymerizing enzyme, and the debranching enzyme that catalyzes the removal of α-(1→6)-linked ramifications. In this work, we rationalize the structural basis of glycogen metabolism in bacteria to the light of the current knowledge. We describe and discuss the remarkable progress made in the understanding of the molecular mechanisms of substrate recognition and product release, allosteric regulation and catalysis of all those enzymes.

High levels of phytoplankton visible in satellite imagery are correlated with an increased uptake of carbon compounds by corals. This suggests that corals rely less on carbon production by photosynthetic symbionts when other resources are plentiful, and that the changes in the acquisition mode of carbon can be inferred by remote-sensing techniques.

Bis-(3'-5')-cyclic diguanylic acid (c-di-GMP) belongs to the class of cyclic dinucleotides, key carriers of cellular information in prokaryotic and eukaryotic signal transduction pathways. In bacteria, the intracellular levels of c-di-GMP and their complex physiological outputs are dynamically regulated by environmental and internal stimuli, which control the antagonistic activities of diguanylate cyclases (DGCs) and c-di-GMP specific phosphodiesterases (PDEs). Allostery is one of the major modulators of the c-di-GMP-dependent response. Both the c-di-GMP molecule and the proteins interacting with this second messenger are characterized by an extraordinary structural plasticity, which has to be taken into account when defining and possibly predicting c-di-GMP-related processes. Here, we report a structure-function relationship study on the catalytic portion of the PA0575 protein from Pseudomonas aeruginosa, bearing both putative DGC and PDE domains. The kinetic and structural studies indicate that the GGDEF-EAL portion is a GTP-dependent PDE. Moreover, the crystal structure confirms the high degree of conformational flexibility of this module. We combined structural analysis and protein engineering studies to propose the possible molecular mechanism guiding the nucleotide-dependent allosteric control of catalysis; we propose that the role exerted by GTP via the GGDEF domain is to allow the two EAL domains to form a dimer, the species competent to enter PDE catalysis.

The binary designation Symbiodinium thermophilum was invalid due to the absence of an illustration as required by Article 44.2 of the ICN. Herein, it is validated. This species is the most common symbiont in reef corals in the southern Persian/Arabian Gulf, the world's hottest body of water sustaining reef coral growth.

In response to high salinity, algal endosymbionts of corals produce floridoside, an osmolyte with antioxidant potential., The endosymbiosis between Symbiodinium dinoflagellates and stony corals provides the foundation of coral reef ecosystems. The survival of these ecosystems is under threat at a global scale, and better knowledge is needed to conceive strategies for mitigating future reef loss. Environmental disturbance imposing temperature, salinity, and nutrient stress can lead to the loss of the Symbiodinium partner, causing so-called coral bleaching. Some of the most thermotolerant coral-Symbiodinium associations occur in the Persian/Arabian Gulf and the Red Sea, which also represent the most saline coral habitats. We studied whether Symbiodinium alter their metabolite content in response to high-salinity environments. We found that Symbiodinium cells exposed to high salinity produced high levels of the osmolyte 2-O-glycerol-α-d-galactopyranoside (floridoside), both in vitro and in their coral host animals, thereby increasing their capacity and, putatively, the capacity of the holobiont to cope with the effects of osmotic stress in extreme environments. Given that floridoside has been previously shown to also act as an antioxidant, this osmolyte may serve a dual function: first, to serve as a compatible organic osmolyte accumulated by Symbiodinium in response to elevated salinities and, second, to counter reactive oxygen species produced as a consequence of potential salinity and heat stress.

Photoconvertible fluorescent proteins (pcRFPs) are a group of fluorophores that undergo an irreversible green-to-red shift in emission colour upon irradiation with near-ultraviolet (near-UV) light. Despite their wide application in biotechnology, the high-level expression of pcRFPs in mesophotic and depth-generalist coral species currently lacks a biological explanation. Additionally, reduced penetration of near-UV wavelengths in water poses the question whether light-driven photoconversion is relevant in the mesophotic zone, or whether a different mechanism is involved in the post-translational pigment modification in vivo. Here, we show in a long-term mesocosm experiment that photoconversion in vivo is entirely dependent on near-UV wavelengths. However, a near-UV intensity equivalent to the mesophotic underwater light field at 80 m depth is sufficient to drive the process in vitro, suggesting that photoconversion can occur near the lower distribution limits of these corals. Furthermore, live coral colonies showed evidence of efficient Förster Resonance Energy Transfer (FRET). Our simulated mesophotic light field maintained the pcRFP pool in a partially photoconverted state in vivo, maximising intra-tetrameric FRET and creating a long-range wavelength conversion system with higher quantum yield than other native RFPs. We hypothesise that efficient conversion of blue wavelengths, abundant at depth, into orange-red light could constitute an adaptation of corals to life in light-limited environments.

Enrichment of reef environments with dissolved inorganic nutrients is considered a major threat to the survival of corals living in symbiosis with dinoflagellates (Symbiodinium sp.). We argue, however, that the direct negative effects on the symbiosis are not necessarily caused by the nutrient enrichment itself but by the phosphorus starvation of the algal symbionts that can be caused by skewed nitrogen (N) to phosphorus (P) ratios. We exposed corals to imbalanced N:P ratios in long-term experiments and found that the undersupply of phosphate severely disturbed the symbiosis, indicated by the loss of coral biomass, malfunctioning of algal photosynthesis and bleaching of the corals. In contrast, the corals tolerated an undersupply with nitrogen at high phosphate concentrations without negative effects on symbiont photosynthesis, suggesting a better adaptation to nitrogen limitation. Transmission electron microscopy analysis revealed that the signatures of ultrastructural biomarkers represent versatile tools for the classification of nutrient stress in symbiotic algae. Notably, high N:P ratios in the water were clearly identified by the accumulation of uric acid crystals., Highlights • Undersupply with dissolved inorganic phosphate causes coral bleaching. • Ultrastructural biomarkers in algal symbionts identify nutrient stress in reef corals. • Uric acid crystals in zooxanthellae identify high N:P ratios in the water column. • Nitrate enrichment of the water causes phosphate deficiency in Symbiodinium. • Coral symbionts tolerate nitrogen limitation better than phosphorus limitation.

Anthropogenic nutrient enrichment is often associated with coral reef decline. Consequently, there is a large consent that increased nutrient influxes in reef waters have negative longterm consequences for corals. However, the mechanisms by which dissolved inorganic nutrients can disturb corals and their symbiotic algae are subject to controversial debate. Herein, we discuss recent studies that demonstrate how nutrient enrichment affects the heat and light stress tolerance of corals and their bleaching susceptibility. We integrate direct and indirect effects of nutrient enrichment on corals in a model that explains why healthy coral reefs can exist over a rather broad range of natural nutrient environments at the lower end of the concentration scale and that anthropogenic nutrient enrichment can disturb the finely balanced processes via multiple pathways. We conceptualise that corals can suffer from secondary negative nutrient effects due to the alteration of their natural nutrient environment by increased phytoplankton loads. In this context, we suggest that phytoplankton represents a likely vector that can translate nutrients effects, induced for instance by coastal run-off, into nutrient stress on coral reefs in considerable distance to the site of primary nutrient enrichment. The presented synthesis of the literature suggests that the effects of nutrient enrichment and eutrophication beyond certain thresholds are negative for the physiological performance of the coral individual and for ecosystem functioning. Hence, the immediate implementation of knowledge-based nutrient management strategies is crucial for coral reef survival.

Fatal infestations of land-based Acropora cultures with so-called Acropora-eating flatworms (AEFWs) are a global phenomenon. We evaluate the hypothesis that AEFWs represent a risk to coral reefs by studying the biology and the invasive potential of an AEFW strain from the UK. Molecular analyses identified this strain as Amakusaplana acroporae, a new species described from two US aquaria and one natural location in Australia. Our molecular data together with life history strategies described here suggest that this species accounts for most reported cases of AEFW infestations. We show that local parasitic activity impairs the light-acclimation capacity of the whole host colony. A. acroporae acquires excellent camouflage by harbouring photosynthetically competent, host-derived zooxanthellae and pigments of the green-fluorescent protein family. It shows a preference for Acropora valida but accepts a broad host range. Parasite survival in isolation (5–7 d) potentially allows for an invasion when introduced as non-native species in coral reefs.

Green fluorescent protein (GFP)-like pigments are responsible for the vivid colouration of many reef-building corals and have been proposed to act as photoprotectants. Their role remains controversial because the functional mechanism has not been elucidated. We provide direct evidence to support a photoprotective role of the non-fluorescent chromoproteins (CPs) that form a biochemically and photophysically distinct group of GFP-like proteins. Based on observations of Acropora nobilis from the Great Barrier Reef, we explored the photoprotective role of CPs by analysing five coral species under controlled conditions. In vitro and in hospite analyses of chlorophyll excitation demonstrate that screening by CPs leads to a reduction in chlorophyll excitation corresponding to the spectral properties of the specific CPs present in the coral tissues. Between 562 and 586 nm, the CPs maximal absorption range, there was an up to 50 % reduction of chlorophyll excitation. The screening was consistent for established and regenerating tissue and amongst symbiont clades A, C and D. Moreover, among two differently pigmented morphs of Acropora valida grown under identical light conditions and hosting subclade type C3 symbionts, high CP expression correlated with reduced photodamage under acute light stress.

International audience; Phosphorus (P), an essential structural and functional component of all living organisms, is considered to be the ultimate limiting nutrient in marine ecosystems. To optimize its acquisition, marine species such as protozoa, sponges, foraminifera, clams, and reef corals, among others, have entered symbiotic relationships with algae, which recycle waste products of the animal host and transform dissolved inorganic nutrients into organic molecules, making them bioavailable to their host. Such associations provide a competitive edge in an environment where ambient nutrient availability is low. The aim of this review is to summarize the current knowledge on the P sources available to reef organisms with algal symbionts, to discuss the means by which P is taken up and stored within the symbiosis, and to assess the effects of eutrophication-induced phosphate enrichment on the algal and host physiology. Finally, we give an overview of knowledge gaps and open questions that should be addressed to better understand the role of phosphorus in reef symbioses functioning. © 2016 by the Ecological Society of America.

A severe bleaching event affected coral communities off the coast of Abu Dhabi, UAE in August/September, 2012. In Saadiyat and Ras Ghanada reefs ~ 40% of the corals showed signs of bleaching. In contrast, only 15% of the corals were affected on Delma reef. Bleaching threshold temperatures for these sites were established using remotely sensed sea surface temperature (SST) data recorded by MODIS-Aqua. The calculated threshold temperatures varied between locations (34.48 °C, 34.55 °C, 35.05 °C), resulting in site-specific deviations in the numbers of days during which these thresholds were exceeded. Hence, the less severe bleaching of Delma reef might be explained by the lower relative heat stress experienced by this coral community. However, the dominance of Porites spp. that is associated with the long-term exposure of Delma reef to elevated temperatures, as well as the more pristine setting may have additionally contributed to the higher coral bleaching threshold for this site.

Mass coral bleaching, resulting from the breakdown of coral-algal symbiosis has been identified as the most severe threat to coral reef survival on a global scale. Regionally, nutrient enrichment of reef waters is often associated with a significant loss of coral cover and diversity. Recently, increased dissolved inorganic nitrogen concentrations have been linked to a reduction of the temperature threshold of coral bleaching, a phenomenon for which no mechanistic explanation is available. Here we show that increased levels of dissolved inorganic nitrogen in combination with limited phosphate concentrations result in an increased susceptibility of corals to temperature-and light-induced bleaching. Mass spectrometric analyses of the algal lipidome revealed a marked accumulation of sulpholipids under these conditions. Together with increased phosphatase activities, this change indicates that the imbalanced supply of dissolved inorganic nitrogen results in phosphate starvation of the symbiotic algae. Based on these findings we introduce a conceptual model that links unfavourable ratios of dissolved inorganic nutrients in the water column with established mechanisms of coral bleaching. Notably, this model improves the understanding of the detrimental effects of coastal nutrient enrichment on coral reefs, which is urgently required to support knowledge-based management strategies to mitigate the effects of climate change. © 2013 Macmillan Publishers Limited. All rights reserved.

Molecular biological methods including genomic and proteomic approaches hold a specific promise to provide new insights into the stress physiology of corals. However, to fully exploit the power of these techniques, aquarium setups are required that allow biological assays under tightly controlled laboratory conditions. Here, details are provided about the successful development of a closed coral mesocosm at the National Oceanography Centre, Southampton. The system can be operated without access to natural seawater and allows long-term observations and experimental studies of reef corals. The individual experimental tanks allow the corals to be exposed, for example, to different light and/or temperature conditions without the need to disconnect them from the system.

Aquarium systems allow technically sophisticated experiments that promise new opportunities to answer urgent questions about reef coral biology, for instance assessing the responses to decreasing environmental pH and/or increased temperatures. Over recent years, long-term culture and (predominantly asexual) propagation of corals has become possible in such systems. At present however, only limited data are available that clarify whether or not responses of the coral holobiont are dominated by the acclimatization to life in captivity or continue to reflect, for example, taxonomic differences seen in nature. We studied the diversity of the symbiotic algae in corals and sea anemones after long-term aquaculture by analysis of their small subunit (SSU) ribosomal DNA gene. A field sample ofAcropora clathratafrom the Arabian Gulf which was used as a control contained clade C zooxanthellae. The aquarium corals also harboured clade C symbionts, but sequencing of the SSU DNA suggested that the analysed animals host different subclades. A prevalence of clade C is also found among corals from the Indo-Pacific region, the origin of most of the aquarium samples. An individual of the temperate sea anemoneAnemonia sulcata (viridis)contained clade A symbionts, similar to those found in nature, even after nearly 10 years of co-culture with sea anemones(Entacmaea quadricolor)and corals hosting clade C symbionts. The results indicate that the specific host–symbiont association occurring in nature appears to persist over >2 years timescales in captivity, with no mixing of symbionts between hosts maintained in the same aquarium or apparent selection of stress-tolerant symbiont strains such as clade D.

We examined the physiological responses to bleaching of two shallow water anemones, Anemonia sulcata var. smaragdina and A. rustica, common to the Mediterranean Sea. The tentacle coloration of A. sulcata var. smaragdina is caused by proteins homologous to the green fluorescent protein (GFP) from Aequorea victoria. Minor amounts of the transcripts for the same GFP-like proteins were also detected in tentacles of the non-fluorescent Anemonia rustica. During a bleaching event observed in the Mediterranean Sea, representatives of both species lost 90% of their zooxanthellae, indicating that the green fluorescent host pigmentation had no influence on the degree of bleaching of the two sea anemone species. Interestingly, the content of fluorescent proteins was also significantly reduced in bleached individuals of A. sulcata. The content of mycosporine-like aminoacids (MAAs) was essentially unaltered in bleached specimens. Bleached anemones were characterized by increased levels of superoxide dismutase activity, whereas the catalase activity in these animals was significantly reduced. The latter results indicate that oxidative stress might have been involved in the observed bleaching of Anemonia spp.

Pigments homologous to the green fluorescent protein (GFP) contribute up to approximately 14% of the soluble protein content of many anthozoans. Maintenance of such high tissue levels poses a severe energetic penalty to the animals if protein turnover is fast. To address this as yet unexplored issue, we established that the irreversible green-to-red conversion of the GFP-like pigments from the reef corals Montastrea cavernosa (mcavRFP) and Lobophyllia hemprichii (EosFP) is driven by violet-blue radiation in vivo and in situ. In the absence of photoconverting light, we subsequently tracked degradation of the red-converted forms of the two proteins in coral tissue using in vivo spectroscopy and immunochemical detection of the post-translational peptide backbone modification. The pigments displayed surprisingly slow decay rates, characterized by half-lives of approximately 20 days. The slow turnover of GFP-like proteins implies that the associated energetic costs for being colorful are comparatively low. Moreover, high in vivo stability makes GFP-like proteins suitable for functions requiring high pigment concentrations, such as photoprotection.

The tolerance responses of plants to many abiotic stresses are conjectured to be controlled by complex gene networks. In the frame of the AtGenExpress project a comprehensive Arabidopsis thaliana genome transcript expression study was performed using the Affymetrix ATH1 microarray in order to understand these regulatory networks in detail. In contrast to earlier studies, we subjected, side-by-side and in a high-resolution kinetic series, Arabidopsis plants, of identical genotype grown under identical conditions, to different environmental stresses comprising heat, cold, drought, salt, high osmolarity, UV-B light and wounding. Furthermore, the harvesting of tissue and RNA isolation were performed in parallel at the same location using identical experimental protocols. Here we describe the technical performance of the experiments. We also present a general overview of environmental abiotic stress-induced gene expression patterns and the results of a model bioinformatics analysis of gene expression in response to UV-B light, drought and cold stress. Our results suggest that the initial transcriptional stress reaction of Arabidopsis might comprise a set of core environmental stress response genes which, by adjustment of the energy balance, could have a crucial function in various stress responses. In addition, there are indications that systemic signals generated by the tissue exposed to stress play a major role in the coordination and execution of stress responses. In summary, the information reported provides a prime reference point and source for the subsequent exploitation of this important resource for research into plant abiotic stress.

For a variety of coral species, we have studied the molecular origin of their coloration to assess the contributions of host and symbiont pigments. For the corals Catalaphyllia jardinei and an orange-emitting color morph of Lobophyllia hemprichii, the pigments belong to a particular class of green fluorescent protein-like proteins that change their color from green to red upon irradiation with ∼400 nm light. The optical absorption and emission properties of these proteins were characterized in detail. Their spectra were found to be similar to those of phycoerythrin from cyanobacterial symbionts. To unambiguously determine the molecular origin of the coloration, we performed immunochemical studies using double diffusion in gel analysis on tissue extracts, including also a third coral species, Montastrea cavernosa, which allowed us to attribute the red fluorescent coloration to green-to-red photoconvertible fluorescent proteins. The red fluorescent proteins are localized mainly in the ectodermal tissue and contribute up to 7.0% of the total soluble cellular proteins in these species. Distinct spatial distributions of green and cyan fluorescent proteins were observed for the tissues of M. cavernosa. This observation may suggest that differently colored green fluorescent protein-like proteins have different, specific functions. In addition to green fluorescent protein-like proteins, the pigments of zooxanthellae have a strong effect on the visual appearance of the latter species.

Intracellular release of calcium ions belongs to the earliest events in cellular stress perception. The molecular mechanisms integrating signals from different environmental cues and translating them into an optimized response are largely unknown. We report here the functional characterization of CIPK1, a protein kinase interacting strongly with the calcium sensors CBL1 and CBL9. Comparison of the expression patterns indicates that the three proteins execute their functions in the same tissues. Physical interaction of CIPK1 with CBL1 and CBL9 targets the kinase to the plasma membrane. We show that, similarly to loss of CBL9 function, mutation of either CBL1 or CIPK1 renders plants hypersensitive to osmotic stress. Remarkably, in contrast to the cbl1 mutant and similarly to the cbl9 mutant, loss of CIPK1 function impairs abscisic acid (ABA) responsiveness. We therefore suggest that, by alternative complex formation with either CBL1 or CBL9, the kinase CIPK1 represents a convergence point for ABA-dependent and ABA-independent stress responses. Based on our genetic, physiological and protein-protein interaction data, we propose a general model for information processing in calcium-regulated signalling networks.

MADS-box genes in plants are putative transcription factors involved in regulating numerous developmental processes, such as meristem and organ identity in inflorescences and in flowers. Recent reports indicate that they are involved in other processes than flower development such as the establishment of developing embryos, seed coat and ultimately in root and fruit development. We have identified seven tomato MADS-box genes that are highly expressed during the first steps of tomato fruit development. According to comparisons of their deduced amino acid sequences, they were classified into two groups: (1) already identified tomato MADS-box genes previously defined as flower identity genes (TAG1, TDR4 and TDR6) and (2) new tomato MADS-box genes (TAGL1, TAGL2, TAGL11 and TAGL12). With the exception of TAGL12, which is expressed near uniformly in every tissue, the other genes show an induction during the tomato fruit development phase I (anthesis) and phase II, when active cell division occurs. In situ hybridization analyses show a specific expression pattern for each gene within the fruit and embryo sac tissues suggesting an important role in the establishment of tissue identity. Yeast two-hybrid analyses indicate that some of these proteins could potentially form dimers suggesting they could act together to accomplish their proposed role.

Coral reefs are in rapid decline on a global scale due to human activities and a changing climate. Shallow water reefs depend on the obligatory symbiosis between the habitat forming coral host and its algal symbiont from the genus Symbiodinium (zooxanthellae). This association is highly sensitive to thermal perturbations and temperatures as little as 1°C above the average summer maxima can cause the breakdown of this symbiosis, termed coral bleaching. Predicting the capacity of corals to survive the expected increase in seawater temperatures depends strongly on our understanding of the thermal tolerance of the symbiotic algae. Here we use molecular phylogenetic analysis of four genetic markers to describe Symbiodinium thermophilum, sp. nov. from the Persian/Arabian Gulf, a thermally tolerant coral symbiont. Phylogenetic inference using the non-coding region of the chloroplast psbA gene resolves S. thermophilum as a monophyletic lineage with large genetic distances from any other ITS2 C3 type found outside the Gulf. Through the characterisation of Symbiodinium associations of 6 species (5 genera) of Gulf corals, we demonstrate that S. thermophilum is the prevalent symbiont all year round in the world's hottest sea, the southern Persian/Arabian Gulf.

Predicted increases in seawater temperatures accelerate coral reef decline due to mortality by heat-driven coral bleaching. Alteration of the natural nutrient environment of reef corals reduces tolerance of corals to heat and light stress and thus will exacerbate impacts of global warming on reefs. Still, many reefs demonstrate remarkable regeneration from past stress events. This paper investigates the effects of sea surface temperature (SST) and water column productivity on recovery of coral reefs. In 71 Indo-Pacific sites, coral cover changes over the past 1-3 decades correlated negative-exponentially with mean SST, chlorophyll a, and SST rise. At six monitoring sites (Persian/Arabian Gulf, Red Sea, northern and southern Galápagos, Easter Island, Panama), over half of all corals were

The genomic framework that enables corals to adjust to unfavourable conditions is crucial for coral reef survival in a rapidly changing climate. We have explored the striking intraspecific variability in the expression of coral pigments from the green fluorescent protein (GFP) family to elucidate the genomic basis for the plasticity of stress responses among reef corals. We show that multicopy genes can greatly increase the dynamic range over which corals can modulate transcript levels in response to the light environment. Using the red fluorescent protein amilFP597 in the coral Acropora millepora as a model, we demonstrate that its expression increases with light intensity, but both the minimal and maximal gene transcript levels vary markedly among colour morphs. The pigment concentration in the tissue of different morphs is strongly correlated with the number of gene copies with a particular promoter type. These findings indicate that colour polymorphism in reef corals can be caused by the environmentally regulated expression of multicopy genes. High‐level expression of amilFP597 is correlated with reduced photodamage of zooxanthellae under acute light stress, supporting a photoprotective function of this pigment. The cluster of light‐regulated pigment genes can enable corals to invest either in expensive high‐level pigmentation, offering benefits under light stress, or to rely on low tissue pigment concentrations and use the conserved resources for other purposes, which is preferable in less light‐exposed environments. The genomic framework described here allows corals to pursue different strategies to succeed in habitats with highly variable light stress levels. In summary, our results suggest that the intraspecific plasticity of reef corals’ stress responses is larger than previously thought.

The symbiotic association of corals and unicellular algae of the genus Symbiodinium in the southern Persian/Arabian Gulf (PAG) display an exceptional heat tolerance, enduring summer peak temperatures of up to 36?°C. As yet, it is not clear whether this resilience is related to the presence of specific symbiont types that are exclusively found in this region. Therefore, we used molecular markers to identify the symbiotic algae of three Porites species along >1000?km of coastline in the PAG and the Gulf of Oman and found that a recently described species, Symbiodinium thermophilum, is integral to coral survival in the southern PAG, the world’s hottest sea. Despite the geographic isolation of the PAG, we discovered that representatives of the S. thermophilum group can also be found in the adjacent Gulf of Oman providing a potential source of thermotolerant symbionts that might facilitate the adaptation of Indian Ocean populations to the higher water temperatures expected for the future. However, corals from the PAG associated with S. thermophilum show strong local adaptation not only to high temperatures but also to the exceptionally high salinity of their habitat. We show that their superior heat tolerance can be lost when these corals are exposed to reduced salinity levels common for oceanic environments elsewhere. Consequently, the salinity prevailing in most reefs outside the PAG might represent a distribution barrier for extreme temperature-tolerant coral/Symbiodinium associations from the PAG.

Both polyethylene glycols (PEG) of M w 8,000 and that of 10 000 stabilize the native compact state of human albumin showing negative preferential interaction with the protein. Interaction between these polymers and the protein is thermodynamically unfavorable, and becomes even more unfavorable for denatured protein whose surface areas are larger than those of native ones. PEG of low M w 1000 and 4000 did not show steric exclusion, interacting favorably with hydrophobic side chains made available when the protein was unfolded and leading to a stabilization of the unfolded state, which is manifested as a lowering of the thermal transition temperature. Perturbation of the absorption spectrum of albumin by PEGs confirms that at high temperature the polymers preferentially interact with the denatured state of albumin, but is excluded from the native state at low temperature. This observation is consistent with the fact that PEG is hydrophobic in nature and may interact favorably with the hydrophobic side chain exposed upon unfolding. The lower activation energy for thermal unfolding in the presence of PEG 1000 is in favour of preferential interaction of this polymer with human albumin. PEG of low M w favours the ionization of the tyrosine residues of albumin. It is apparent that pKa decreased with the increase in M w of synthetic polymer. Such variation might be a consequence of the change in dielectric constant at the domain of the protein by PEG binding.

Corals in the Arabian/Persian Gulf endure summer temperatures of up to 36 °C, making them ideal subjects to study the mechanisms underlying thermal tolerance. Unexpectedly, we found the “generalist” Symbiodinium clade C3 to be the prevalent symbiont among seven coral species from Abu Dhabi (UAE) waters. Moreover, C3 represented the only dominant symbiont type in Porites spp. from this region. The “thermotolerant” symbionts D1a and C15 were not encountered, indicating that the association with these symbionts cannot be the sole reason for the heat tolerance of Gulf corals. The association of Porites lobata with specific symbiont types (C3 vs. C15) in samples from habitats with very different temperature regimes (Abu Dhabi vs. Fiji) remained unaffected by laboratory culture. During temperature stress experiments specimens from both locations strongly downregulated green fluorescent protein (GFP)-like pigments. However, the Abu Dhabi samples were less prone to bleaching and showed lower mortality.

Fluorescent proteins (FPs) of the green fluorescent protein (GFP)-like protein family are not only responsible for the spectacular colors of some reef corals, but they can be also found in several other marine invertebrate taxa. The chromophore of these proteins resides in the center of an 11-stranded ?-barrel and is formed by autocatalytic reactions in the presence of oxygen. In some proteins, photochemical reactions are involved in the maturation of the chromophore. The variability of the chromophore structures and the interactions with the surrounding protein scaffold are responsible for the diverse emission colors, ranging from cyan to green and yellow to red. FPs develop their fluorescence also upon expression in recombinant cells which allows their use as genetically encoded markers. Application examples include the labeling of cells, cellular compartment or proteins, studies of gene activity, and sensor studies. Recently, photoactivatable FPs enabled live-cell imaging with a resolution beyond the diffraction barrier of optical microscopy. Nevertheless, before their potential as advanced markers can be fully exploited, FPs often need to undergo extensive protein engineering to alter some detrimental properties such as the tendency to form stable multimers.