Proteomics quantifies protein expression changes in a model cnidarian colonised by a thermally tolerant but suboptimal symbiont.
ABSTRACT: The acquisition of thermally tolerant algal symbionts by corals has been proposed as a natural or assisted mechanism of increasing coral reef resilience to anthropogenic climate change, but the cell-level processes determining the performance of new symbiotic associations are poorly understood. We used liquid chromatography-mass spectrometry to investigate the effects of an experimentally induced symbiosis on the host proteome of the model sea anemone Exaiptasia pallida. Aposymbiotic specimens were colonised by either the homologous dinoflagellate symbiont (Breviolum minutum) or a thermally tolerant, ecologically invasive heterologous symbiont (Durusdinium trenchii). Anemones containing D. trenchii exhibited minimal expression of Niemann-Pick C2 proteins, which have predicted biochemical roles in sterol transport and cell recognition, and glutamine synthetases, which are thought to be involved in nitrogen assimilation and recycling between partners. D. trenchii-colonised anemones had higher expression of methionine-synthesising betaine-homocysteine S-methyltransferases and proteins with predicted oxidative stress response functions. Multiple lysosome-associated proteins were less abundant in both symbiotic treatments compared with the aposymbiotic treatment. The differentially abundant proteins are predicted to represent pathways that may be involved in nutrient transport or resource allocation between partners. These results provide targets for specific experiments to elucidate the mechanisms underpinning compensatory physiology in the coral-dinoflagellate symbiosis.
Project description:The acquisition of thermally tolerant algal symbionts by corals has been proposed as a natural or assisted mechanism of increasing coral reef resilience to anthropogenic climate change, but the cell-level processes determining the performance of new symbiotic associations are poorly understood. We used liquid chromatography-mass spectrometry to investigate the effects of an experimentally-induced symbiosis on the host proteome of the model sea anemone Exaiptasia pallida. Aposymbiotic specimens were colonised by either the homologous dinoflagellate symbiont (Breviolum minutum) or a thermally tolerant, ecologically invasive heterologous symbiont (Durusdinium trenchii). Anemones containing D. trenchii exhibited minimal expression of Niemann-Pick C2 proteins, which have predicted biochemical roles in sterol transport and cell recognition, and glutamine synthetases, which are thought to be involved in nitrogen assimilation and recycling between partners. D. trenchii-colonised anemones had higher expression of methionine-synthesizing betaine–homocysteine S-methyltransferases and proteins with predicted oxidative stress response functions. Multiple lysosome-associated proteins were less abundant in both symbiotic treatments compared with the aposymbiotic treatment. The differentially abundant proteins are predicted to represent pathways that may be involved in nutrient transport or resource allocation between partners. These results provide targets for specific experiments to elucidate the mechanisms underpinning compensatory physiology in the coral–dinoflagellate symbiosis.
Project description:Reef corals and sea anemones form symbioses with unicellular symbiotic dinoflagellates. The molecular circumventions that underlie the successful intracellular colonization of hosts by symbionts are still largely unknown. We conducted proteomic analyses to determine molecular differences of Exaiptasia pallida anemones colonized by physiologically different symbiont species, in comparison with symbiont-free (aposymbiotic) anemones. We compared one homologous species, Symbiodinium linucheae, that is natively associated with the clonal Exaiptasia strain (CC7) to another heterologous species, Durusdinium trenchii, a thermally tolerant species that colonizes numerous coral species. This approach allowed the discovery of a core set of host genes that are differentially regulated as a function of symbiosis regardless of symbiont species. The findings revealed that symbiont colonization at higher densities requires circumvention of the host cellular immunological response, enhancement of ammonium regulation, and suppression of phagocytosis after a host cell in colonized. Furthermore, the heterologous symbionts failed to duplicate the same level of homologous colonization within the host, evidenced by substantially lower symbiont densities. This reduced colonization of D. trenchii correlated with its inability to circumvent key host systems including autophagy-suppressing modulators, cytoskeletal alteration, and isomerase activity. The larger capability of host molecular circumvention by homologous symbionts could be the result of a longer evolutionary history of host/symbiont interactions, which translates into a more finely tuned symbiosis. These findings are of great importance within the context of the response of reef corals to climate change since it has been suggested that coral may acclimatize to ocean warming by changing their dominant symbiont species.
Project description:The relationship between corals and dinoflagellates of the genus Symbiodinium is fundamental to the functioning of coral ecosystems. It has been suggested that reef corals may adapt to climate change by changing their dominant symbiont type to a more thermally tolerant one, although the capacity for such a shift is potentially hindered by the compatibility of different host-symbiont pairings. Here we combined transcriptomic and metabolomic analyses to characterize the molecular, cellular, and physiological processes that underlie this compatibility, with a particular focus on Symbiodinium trenchii, an opportunistic, thermally tolerant symbiont that flourishes in coral tissues after bleaching events. Symbiont-free individuals of the sea anemone Exaiptasia pallida (commonly referred to as Aiptasia), an established model system for the study of the cnidarian-dinoflagellate symbiosis, were colonized with the "normal" (homologous) symbiont Symbiodinium minutum and the heterologous S. trenchii Analysis of the host gene and metabolite expression profiles revealed that heterologous symbionts induced an expression pattern intermediate between the typical symbiotic state and the aposymbiotic state. Furthermore, integrated pathway analysis revealed that increased catabolism of fixed carbon stores, metabolic signaling, and immune processes occurred in response to the heterologous symbiont type. Our data suggest that both nutritional provisioning and the immune response induced by the foreign "invader" are important factors in determining the capacity of corals to adapt to climate change through the establishment of novel symbioses.
Project description:Trophic endosymbiosis between anthozoans and photosynthetic dinoflagellates forms the key foundation of reef ecosystems. Dysfunction and collapse of symbiosis lead to bleaching (symbiont expulsion), which is responsible for the severe worldwide decline of coral reefs. Molecular signals are central to the stability of this partnership and are therefore closely related to coral health. To decipher inter-partner signaling, we developed genomic resources (cDNA library and microarrays) from the symbiotic sea anemone Anemonia viridis. Here we describe differential expression between symbiotic (also called zooxanthellate anemones) or aposymbiotic (also called bleached) A. viridis specimens, using microarray hybridizations and qPCR experiments. We mapped, for the first time, transcript abundance separately in the epidermal cell layer and the gastrodermal cells that host photosynthetic symbionts. Transcriptomic profiles showed large inter-individual variability, indicating that aposymbiosis could be induced by different pathways. We defined a restricted subset of 39 common genes that are characteristic of the symbiotic or aposymbiotic states. We demonstrated that transcription of many genes belonging to this set is specifically enhanced in the symbiotic cells (gastroderm). A model is proposed where the aposymbiotic and therefore heterotrophic state triggers vesicular trafficking, whereas the symbiotic and therefore autotrophic state favors metabolic exchanges between host and symbiont. Several genetic pathways were investigated in more detail: i) a key vitamin K-dependant process involved in the dinoflagellate-cnidarian recognition; ii) two cnidarian tissue-specific carbonic anhydrases involved in the carbon transfer from the environment to the intracellular symbionts; iii) host collagen synthesis, mostly supported by the symbiotic tissue. Further, we identified specific gene duplications and showed that the cnidarian-specific isoform was also up-regulated both in the symbiotic state and in the gastroderm. Our results thus offer new insight into the inter-partner signaling required for the physiological mechanisms of the symbiosis that is crucial for coral health.
Project description:The ability of corals and other cnidarians to survive climate change depends partly on the composition of their endosymbiont communities. The dinoflagellate family Symbiodiniaceae is genetically and physiologically diverse, and one proposed mechanism for cnidarians to acclimate to rising temperatures is to acquire more thermally tolerant symbionts. However, cnidarian-dinoflagellate associations vary in their degree of specificity, which may limit their capacity to alter symbiont communities. Here, we inoculated symbiont-free polyps of the sea anemone Exaiptasia pallida (commonly referred to as 'Aiptasia'), a model system for the cnidarian-dinoflagellate symbiosis, with simultaneous or sequential mixtures of thermally tolerant and thermally sensitive species of Symbiodiniaceae. We then monitored symbiont success (relative proportional abundance) at normal and elevated temperatures across two to four weeks. All anemones showed signs of bleaching at high temperature. During simultaneous inoculations, the native, thermally sensitive Breviolum minutum colonized polyps most successfully regardless of temperature when paired against the non-native but more thermally tolerant Symbiodinium microadriaticum or Durusdinium trenchii. Furthermore, anemones initially colonized with B. minutum and subsequently exposed to S. microadriaticum failed to acquire the new symbiont. These results highlight how partner specificity may place strong limitations on the ability of certain cnidarians to acquire more thermally tolerant symbionts, and hence their adaptive potential under climate change.
Project description:The endosymbiotic relationship between cnidarians and photosynthetic dinoflagellate algae provides the foundation of coral reef ecosystems. This essential interaction is globally threatened by anthropogenic disturbance. As such, it is important to understand the molecular mechanisms underpinning the cnidarian-algal association. Here we investigated phosphorylation-mediated protein signalling as a mechanism of regulation of the cnidarian-algal interaction, and we report on the generation of the first phosphoproteome for the coral model system Aiptasia. Mass spectrometry-based phosphoproteomics using data-independent acquisition allowed consistent quantification of over 3,000 phosphopeptides totalling more than 1,600 phosphoproteins across aposymbiotic (symbiont-free) and symbiotic anemones. Comparison of the symbiotic states showed distinct phosphoproteomic profiles attributable to the differential phosphorylation of 539 proteins that cover a broad range of functions, from receptors to structural and signal transduction proteins. A subsequent pathway enrichment analysis identified the processes of "protein digestion and absorption," "carbohydrate metabolism," and "protein folding, sorting and degradation," and highlighted differential phosphorylation of the "phospholipase D signalling pathway" and "protein processing in the endoplasmic reticulum." Targeted phosphorylation of the phospholipase D signalling pathway suggests control of glutamate vesicle trafficking across symbiotic compartments, and phosphorylation of the endoplasmic reticulum machinery suggests recycling of symbiosome-associated proteins. Our study shows for the first time that changes in the phosphorylation status of proteins between aposymbiotic and symbiotic Aiptasia anemones may play a role in the regulation of the cnidarian-algal symbiosis. This is the first phosphoproteomic study of a cnidarian-algal symbiotic association as well as the first application of quantification by data-independent acquisition in the coral field.
Project description:The symbiotic relationship between cnidarians and dinoflagellates is the cornerstone of coral reef ecosystems. Although research has focused on the molecular mechanisms underlying this symbiosis, the role of epigenetic mechanisms, that is, the study of heritable changes that do not involve changes in the DNA sequence, is unknown. To assess the role of DNA methylation in the cnidarian-dinoflagellate symbiosis, we analyzed genome-wide CpG methylation, histone associations, and transcriptomic states of symbiotic and aposymbiotic anemones in the model system Aiptasia. We found that methylated genes are marked by histone 3 lysine 36 trimethylation (H3K36me3) and show significant reduction of spurious transcription and transcriptional noise, revealing a role of DNA methylation in the maintenance of transcriptional homeostasis. Changes in DNA methylation and expression show enrichment for symbiosis-related processes, such as immunity, apoptosis, phagocytosis recognition, and phagosome formation, and reveal intricate interactions between the underlying pathways. Our results demonstrate that DNA methylation provides an epigenetic mechanism of transcriptional homeostasis that responds to symbiosis.
Project description:To clarify the establishment process of coral-algal symbiotic relationships, coral transcriptome changes during increasing algal symbiont densities were examined in juvenile corals following inoculation with the algae Symbiodinium goreaui (clade C) and S. trenchii (clade D), and comparison of their transcriptomes with aposymbiotic corals by RNA-sequencing. Since Symbiodinium clades C and D showed very different rates of density increase, comparisons were made of early onsets of both symbionts, revealing that the host behaved differently for each. RNA-sequencing showed that the number of differentially-expressed genes in corals colonized by clade D increased ca. two-fold from 10 to 20 days, whereas corals with clade C showed unremarkable changes consistent with a slow rate of density increase. The data revealed dynamic metabolic changes in symbiotic corals. In addition, the endocytosis pathway was also upregulated, while lysosomal digestive enzymes and the immune system tended to be downregulated as the density of clade D algae increased. The present dataset provides an enormous number of candidate symbiosis-related molecules that exhibit the detailed process by which coral-algal endosymbiosis is established.
Project description:Background: Cnidarian – dinoflagellate intracellular symbioses are one of the most important mutualisms in the marine environment. They form the trophic and structural foundation of coral reef ecosystems, and have played a key role in the evolutionary radiation and biodiversity of cnidarian species. Despite the prevalence of these symbioses, we still know very little about the molecular modulators that initiate, regulate, and maintain the interaction between these two different biological entities. In this study, we conducted a comparative host anemone transcriptome analysis using a cDNA microarray platform to identify genes involved in cnidarian – algal symbiosis. Results: We detected statistically significant differences in host gene expression profiles between sea anemones (Anthopleura elegantissima) in a symbiotic and non-symbiotic state. The group of genes, whose expression is altered, is diverse, suggesting that the molecular regulation of the symbiosis is governed by changes in multiple cellular processes. In the context of cnidarian – dinoflagellate symbioses, we discuss pivotal host gene expression changes involved in lipid metabolism, cell adhesion, cell proliferation, apoptosis, and oxidative stress. Conclusion: Our data do not support the existence of symbiosis-specific genes involved in controlling and regulating the symbiosis. Instead, it appears that the symbiosis is maintained by altering expression of existing genes involved in vital cellular processes. Specifically, the finding of key genes involved in cell cycle progression and apoptosis have led us to hypothesize that a suppression of apoptosis, together with a deregulation of the host cell cycle, create a platform that might be necessary for symbiont and/or symbiont-containing host cell survival. This first comprehensive molecular examination of the cnidarian – dinoflagellate associations provides critical insight into the maintenance and regulation of the symbiosis. Keywords: comparative genomic hybridization Overall design: We applied a multiple dye-swap experimental design for the two conditions, aposymbiotic and symbiotic anemone groups, compared in our experiment. For this experiment, there were no reference samples. Six biological replicates per condition were used as recommended for this type of two-comparison experimental design. There were not technical replicates on the array.
Project description:Corals rely on a symbiosis with dinoflagellate algae (Symbiodinium spp.) to thrive in nutrient poor tropical oceans. However, the coral-algal symbiosis can break down during bleaching events, potentially leading to coral death. While genome-wide expression studies have shown the genes associated with the breakdown of this partnership, the full conglomerate of genes responsible for the establishment and maintenance of a healthy symbiosis remains unknown. Results from previous studies suggested little transcriptomic change associated with the establishment of symbiosis. In order to elucidate the transcriptomic response of the coral host in the presence of its associated symbiont, we utilized a comparative framework. Post-metamorphic aposymbiotic coral polyps of Orbicella faveolata were compared to symbiotic coral polyps 9 days after metamorphosis and the subsequent differential gene expression between control and treatment was quantified using cDNA microarray technology. Coral polyps exhibited differential expression of genes associated with nutrient metabolism and development, providing insight into pathways turned as a result of symbiosis driving early polyp growth. Furthermore, genes associated with lysosomal fusion were also upregulated, suggesting host regulation of symbiont densities soon after infection. Overall design: RNA from 3 control and 3 infected polyp samples were hybridized in a 3-replicate pooled reference (6 total hyb's)