Transcriptome analysis of a cnidarian – dinoflagellate mutualism reveals complex modulation of host gene expression

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 insights into the maintenance and regulation of the symbiosis

NEEMO 20: Science Training, Operations, and Tool Development

The 20th mission of the National Aeronautics and Space Administration (NASA) Extreme Environment Mission Operations (NEEMO) was a highly integrated evaluation of operational protocols and tools designed to enable future exploration beyond low-Earth orbit. NEEMO 20 was conducted from the Aquarius habitat off the coast of Key Largo, FL in July 2015. The habitat and its surroundings provide a convincing analog for space exploration. A crew of six (comprised of astronauts, engineers, and habitat technicians) lived and worked in and around the unique underwater laboratory over a mission duration of 14-days. Incorporated into NEEMO 20 was a diverse Science Team (ST) comprised of geoscientists from the Astromaterials Research and Exploration Science (ARES/XI) Division from the Johnson Space Center (JSC), as well as marine scientists from the Department of Biological Sciences at Florida International University (FIU). This team trained the crew on the science to be conducted, defined sampling techniques and operational procedures, and planned and coordinated the science focused Extra Vehicular Activities (EVAs). The primary science objectives of NEEMO 20 was to study planetary sampling techniques and tools in partial gravity environments under realistic mission communication time delays and operational pressures. To facilitate these objectives two types of science sites were employed 1) geoscience sites with available rocks and regolith for testing sampling procedures and tools and, 2) marine science sites dedicated to specific research focused on assessing the photosynthetic capability of corals and their genetic connectivity between deep and shallow reefs. These marine sites and associated research objectives included deployment of handheld instrumentation, context descriptions, imaging, and sampling; thus acted as a suitable proxy for planetary surface exploration activities. This abstract briefly summarizes the scientific training, scientific operations, and tool development conducted during NEEMO 20 with an emphasis on the primary lessons learned

Genomic signatures in the coral holobiont reveal host adaptations driven by Holocene climate change and reef specific symbionts

Genetic signatures caused by demographic and adaptive processes during past climatic shifts can inform predictions of species’ responses to anthropogenic climate change. To identify these signatures in Acropora tenuis, a reef-building coral threatened by global warming, we first assembled the genome from long reads and then used shallow whole-genome resequencing of 150 colonies from the central inshore Great Barrier Reef to inform population genomic analyses. We identify population structure in the host that reflects a Pleistocene split, whereas photosymbiont differences between reefs most likely reflect contemporary (Holocene) conditions. Signatures of selection in the host were associated with genes linked to diverse processes including osmotic regulation, skeletal development, and the establishment and maintenance of symbiosis. Our results suggest that adaptation to post-glacial climate change in A. tenuis has involved selection on many genes, while differences in symbiont specificity between reefs appear to be unrelated to host population structure

Transcriptional Activation of c3 and hsp70 as Part of the Immune Response of Acropora millepora to Bacterial Challenges

The impact of disease outbreaks on coral physiology represents an increasing concern for the fitness and resilience of reef ecosystems. Predicting the tolerance of corals to disease relies on an understanding of the coral immune response to pathogenic interactions. This study explored the transcriptional response of two putative immune genes (c3 and c-type lectin) and one stress response gene (hsp70) in the reef building coral, Acropora millepora challenged for 48 hours with bacterial strains, Vibrio coralliilyticus and Alteromonas sp. at concentrations of 106 cells ml-1. Coral fragments challenged with V. coralliilyticus appeared healthy while fragments challenged with Alteromonas sp. showed signs of tissue lesions after 48 hr. Coral-associated bacterial community profiles assessed using denaturing gradient gel electrophoresis changed after challenge by both bacterial strains with the Alteromonas sp. treatment demonstrating the greatest community shift. Transcriptional profiles of c3 and hsp70 increased at 24 hours and correlated with disease signs in the Alteromonas sp. treatment. The expression of hsp70 also showed a significant increase in V. coralliilyticus inoculated corals at 24 h suggesting that even in the absence of disease signs, the microbial inoculum activated a stress response in the coral. C-type lectin did not show a response to any of the bacterial treatments. Increase in gene expression of c3 and hsp70 in corals showing signs of disease indicates their potential involvement in immune and stress response to microbial challenges

Regulation of Apoptotic Pathways by Stylophora pistillata (Anthozoa, Pocilloporidae) to Survive Thermal Stress and Bleaching

Elevated seawater temperatures are associated with coral bleaching events and related mortality. Nevertheless, some coral species are able to survive bleaching and recover. The apoptotic responses associated to this ability were studied over 3 years in the coral Stylophora pistillata from the Gulf of Eilat subjected to long term thermal stress. These include caspase activity and the expression profiles of the S. pistillata caspase and Bcl-2 genes (StyCasp and StyBcl-2-like) cloned in this study. In corals exposed to thermal stress (32 or 34°C), caspase activity and the expression levels of the StyBcl-2-like gene increased over time (6–48 h) and declined to basal levels within 72 h of thermal stress. Distinct transcript levels were obtained for the StyCasp gene, with stimulated expression from 6 to 48 h of 34°C thermal stress, coinciding with the onset of bleaching. Increased cell death was detected in situ only between 6 to 48 h of stress and was limited to the gastroderm. The bleached corals survived up to one month at 32°C, and recovered back symbionts when placed at 24°C. These results point to a two-stage response in corals that withstand thermal stress: (i) the onset of apoptosis, accompanied by rapid activation of anti-oxidant/anti-apoptotic mediators that block the progression of apoptosis to other cells and (ii) acclimatization of the coral to the chronic thermal stress alongside the completion of symbiosis breakdown. Accordingly, the coral's ability to rapidly curb apoptosis appears to be the most important trait affecting the coral's thermotolerance and survival

Adaptations to Endosymbiosis in a Cnidarian-Dinoflagellate Association: Differential Gene Expression and Specific Gene Duplications

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

Macroalgal-Associated Dinoflagellates Belonging to the Genus Symbiodinium in Caribbean Reefs

Coral-algal symbiosis has been a subject of great attention during the last two decades in response to global coral reef decline. However, the occurrence and dispersion of free-living dinoflagellates belonging to the genus Symbiodinium are less documented. Here ecological and molecular evidence is presented demonstrating the existence of demersal free-living Symbiodinium populations in Caribbean reefs and the possible role of the stoplight parrotfish (Sparisoma viride) as Symbiodinium spp. dispersers. Communities of free-living Symbiodinium were found within macroalgal beds consisting of Halimeda spp., Lobophora variegata, Amphiroa spp., Caulerpa spp. and Dictyota spp. Viable Symbiodinium spp. cells were isolated and cultured from macroalgal beds and S. viride feces. Further identification of Symbiodinium spp. type was determined by length variation in the Internal Transcribed Spacer 2 (ITS2, nuclear rDNA) and length variation in domain V of the chloroplast large subunit ribosomal DNA (cp23S-rDNA). Determination of free-living Symbiodinium and mechanisms of dispersal is important in understanding the life cycle of Symbiodinium spp

Rapid Evolution of Coral Proteins Responsible for Interaction with the Environment

Christian R. Voolstra is with King Abdullah University of Science and Technology, Shinichi Sunagawa is with the European Molecular Biology Laboratory, Mikhail V. Matz is with UT Austin, Till Bayer is with King Abdullah University of Science and Technology, Manuel Aranda is with King Abdullah University of Science and Technology, Emmanuel Buschiazzo is with University of California Merced, Michael K. DeSalvo is with University of California San Francisco, Erika Lindquist is with the Department of Energy Joint Genome Institute, Alina M. Szmant is with University of North Carolina Wilmington, Mary Alice Coffroth is with State University of New York at Buffalo, Mónica Medina is with University of California Merced.Background -- Corals worldwide are in decline due to climate change effects (e.g., rising seawater temperatures), pollution, and exploitation. The ability of corals to cope with these stressors in the long run depends on the evolvability of the underlying genetic networks and proteins, which remain largely unknown. A genome-wide scan for positively selected genes between related coral species can help to narrow down the search space considerably. Methodology/Principal Findings -- We screened a set of 2,604 putative orthologs from EST-based sequence datasets of the coral species Acropora millepora and Acropora palmata to determine the fraction and identity of proteins that may experience adaptive evolution. 7% of the orthologs show elevated rates of evolution. Taxonomically-restricted (i.e. lineage-specific) genes show a positive selection signature more frequently than genes that are found across many animal phyla. The class of proteins that displayed elevated evolutionary rates was significantly enriched for proteins involved in immunity and defense, reproduction, and sensory perception. We also found elevated rates of evolution in several other functional groups such as management of membrane vesicles, transmembrane transport of ions and organic molecules, cell adhesion, and oxidative stress response. Proteins in these processes might be related to the endosymbiotic relationship corals maintain with dinoflagellates in the genus Symbiodinium. Conclusion/Relevance -- This study provides a birds-eye view of the processes potentially underlying coral adaptation, which will serve as a foundation for future work to elucidate the rates, patterns, and mechanisms of corals' evolutionary response to global climate change.This work was supported by DEB-1054766 to M.V.M. and National Science Foundation grants IOS-0644438 and OCE-0313708 to M.M., and by a Collaborative Travel Fund to C.R.V. made by King Abdullah University of Science and Technology (KAUST). The work conducted by the U.S. Department of Energy Joint Genome Institute is supported by the Office of Science of the U.S. Department of Energy under Contract No. DE-AC02-05CH11231. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Biological Sciences, School o