Ann M. Tarrant

Ocean warming slows coral growth in the central Red Sea.

Red Sea Coral Decline Large, rapid sea surface temperature rises of 1°C or greater typically cause bleaching of corals. Cantin et al. (p. 322) show that smaller temperature increases also have detrimental effects on corals, dramatically reducing their rates of calcification and skeletal extension. Corals in the Red Sea, where water temperatures have risen by 0.4 to 1°C since the mid-1970s, have declined in skeletal extension by about 30%, and decreased in calcification rates by around 18% since 1998. This finding suggests that we may see an end to coral growth in the Red Sea this century. Rising summertime sea surface temperatures are slowing the rate of growth of healthy corals in the Red Sea. Sea surface temperature (SST) across much of the tropics has increased by 0.4° to 1°C since the mid-1970s. A parallel increase in the frequency and extent of coral bleaching and mortality has fueled concern that climate change poses a major threat to the survival of coral reef ecosystems worldwide. Here we show that steadily rising SSTs, not ocean acidification, are already driving dramatic changes in the growth of an important reef-building coral in the central Red Sea. Three-dimensional computed tomography analyses of the massive coral Diploastrea heliopora reveal that skeletal growth of apparently healthy colonies has declined by 30% since 1998. The same corals responded to a short-lived warm event in 1941/1942, but recovered within 3 years as the ocean cooled. Combining our data with climate model simulations by the Intergovernmental Panel on Climate Change, we predict that should the current warming trend continue, this coral could cease growing altogether by 2070.

Estrone and estradiol-17β concentration in tissue of the scleractinian coral, Montipora verrucosa

Spawnings of scleractinian corals are affected by light, temperature, and other environmental cues, but no studies elucidate physiological mechanisms that regulate coral gametogenesis. We hypothesized that estrogens may act as bioregulators of coral reproduction. Estrone (E1) and estradiol-17 beta (E2) concentrations were measured in homogenates of tissue and skeleton from M. verrucosa. Tissue samples were collected monthly throughout the year, and more frequently in July and August around spawning. Steroids were extracted with diethyl ether, purified via celite chromatography and assayed with radioimmunoassay. Non-specific binding in coral tissue varied with sample weight and was elevated relative to standards. Monthly mean E1 ranged from 20-70 ng E1 g ash-free dry weight (AFDW)-1, with highest values in April. Smaller asynchronous peaks occurred in early July, prior to spawning. Monthly mean E2 ranged from 8-25 ng E2 g AFDW-1, with highest values in February and March. Peaks in E2 preceded peaks in E1, indicating metabolism of a pool of estrogen. E1 was positively correlated with protein concentration, which is consistent with a bioregulatory role of estrogens. Estrogen peaks in spring and prior to the July spawn corroborate our hypothesis that estrogens regulate coral gametogenesis and spawning.

Heat shock protein expression during stress and diapause in the marine copepod Calanus finmarchicus

Calanoid copepods, such as Calanus finmarchicus, are a key component of marine food webs. C. finmarchicus undergo a facultative diapause during juvenile development, which profoundly affects their seasonal distribution and availability to their predators. The current ignorance of how copepod diapause is regulated limits understanding of copepod population dynamics, distribution, and ecosystem interactions. Heat shock proteins (Hsps) are a superfamily of molecular chaperones characteristically upregulated in response to stress conditions and frequently associated with diapause in other taxa. In this study, 8 heat shock proteins were identified in C. finmarchicus C5 copepodids (Hsp21, Hsp22, p26, Hsp90, and 4 forms of Hsp70), and expression of these transcripts was characterized in response to handling stress and in association with diapause. Hsp21, Hsp22, and Hsp70A (cytosolic subfamily) were induced by handling stress. Expression of Hsp70A was also elevated in shallow active copepodids relative to deep diapausing copepodids, which may reflect induction of this gene by varied stressors in active animals. In contrast, expression of Hsp22 was elevated in deep diapausing animals; Hsp22 may play a role both in short-term stress responses and in protecting proteins from degradation during diapause. Expression of most of the Hsps examined did not vary in response to diapause, perhaps because the diapause of C. finmarchicus is not associated with the extreme environmental conditions (e.g., freezing and desiccation) experienced by many other taxa, such as overwintering insects or Artemia cysts.

Light entrained rhythmic gene expression in the sea anemone Nematostella vectensis : the evolution of the animal circadian clock

Background Circadian rhythms in behavior and physiology are the observable phenotypes from cycles in expression of, interactions between, and degradation of the underlying molecular components. In bilaterian animals, the core molecular components include Timeless-Timeout, photoreceptive cryptochromes, and several members of the basic-loop-helix-Per-ARNT-Sim (bHLH-PAS) family. While many of core circadian genes are conserved throughout the Bilateria, their specific roles vary among species. Here, we identify and experimentally study the rhythmic gene expression of conserved circadian clock members in a sea anemone in order to characterize this gene network in a member of the phylum Cnidaria and to infer critical components of the clockwork used in the last common ancestor of cnidarians and bilaterians. Methodology/Principal Findings We identified homologs of circadian regulatory genes in the sea anemone Nematostella vectensis, including a gene most similar to Timeout, three cryptochromes, and several key bHLH-PAS transcription factors. We then maintained N. vectensis either in complete darkness or in a 12 hour light: 12 hour dark cycle in three different light treatments (blue only, full spectrum, blue-depleted). Gene expression varied in response to light cycle and light treatment, with a particularly strong pattern observed for NvClock. The cryptochromes more closely related to the light-sensitive clade of cryptochromes were upregulated in light treatments that included blue wavelengths. With co-immunoprecipitation, we determined that heterodimerization between CLOCK and CYCLE is conserved within N. vectensis. Additionally, we identified E-box motifs, DNA sequences recognized by the CLOCK:CYCLE heterodimer, upstream of genes showing rhythmic expression. Conclusions/Significance This study reveals conserved molecular and functional components of the circadian clock that were in place at the divergence of the Cnidaria and Bilateria, suggesting the animal circadian clockwork is more ancient than previous data suggest. Characterizing circadian regulation in a cnidarian provides insight into the early origins of animal circadian rhythms and molecular regulation of environmentally cued behaviors.

Nuclear receptor complement of the cnidarian Nematostella vectensis: phylogenetic relationships and developmental expression patterns

BackgroundNuclear receptors are a superfamily of metazoan transcription factors that regulate diverse developmental and physiological processes. Sequenced genomes from an increasing number of bilaterians have provided a more complete picture of duplication and loss of nuclear receptors in protostomes and deuterostomes but have left open the question of which nuclear receptors were present in the cnidarian-bilaterian ancestor. In addition, nuclear receptor expression and function are largely uncharacterized within cnidarians, preventing determination of conserved and novel nuclear receptor functions in the context of animal evolution.ResultsHere we report the first complete set of nuclear receptors from a cnidarian, the starlet sea anemone Nematostella vectensis. Genomic searches using conserved DNA- and ligand-binding domains revealed seventeen nuclear receptors in N. vectensis. Phylogenetic analyses support N. vectensis orthologs of bilaterian nuclear receptors in four nuclear receptor subfamilies within nuclear receptor family 2 (COUP-TF, TLL, HNF4, TR2/4) and one putative ortholog of GCNF (nuclear receptor family 6). Other N. vectensis genes grouped well with nuclear receptor family 2 but represented lineage-specific duplications somewhere within the cnidarian lineage and were not clear orthologs of bilaterian genes. Three nuclear receptors were not well-supported within any particular nuclear receptor family. The seventeen nuclear receptors exhibited distinct developmental expression patterns, with expression of several nuclear receptors limited to a subset of developmental stages.ConclusionN. vectensis contains a diverse complement of nuclear receptors including orthologs of several bilaterian nuclear receptors. Novel nuclear receptors in N. vectensis may be ancient genes lost from triploblastic lineages or may represent cnidarian-specific radiations. Nuclear receptors exhibited distinct developmental expression patterns, which are consistent with diverse regulatory roles for these genes. Understanding the evolutionary relationships and developmental expression of the N. vectensis nuclear receptor complement provides insight into the evolution of the nuclear receptor superfamily and a foundation for mechanistic characterization of cnidarian nuclear receptor function.

Endocrine-like Signaling in Cnidarians: Current Understanding and Implications for Ecophysiology.

Abstract The vertebrate endocrine system is well-characterized, with many reports of disruption by environmental chemicals. In contrast, cnidarians are less compartmentalized, physiological regulation is poorly understood, and the potential for disruption is unknown. Endocrine-like activity has not been systematically studied in cnidarians, but several classical vertebrate hormones (e.g., steroids, iodinated organic compounds, neuropeptides, and indoleamines) have been identified in cnidarian tissues. Investigators have made progress in identifying putative bioregulatory molecules in cnidarians, and testing the effects of these individual compounds. Less progress has been made in elucidating signaling pathways. For example, putative gonadotropin-releasing hormone and sex steroids have been identified in cnidarian tissues, but it is unknown whether these compounds are components of a larger signal cascade comparable to the vertebrate hypothalamic-pituitary-gonadal axis. Further, while sex steroids and iodinated organic compounds may help to regulate cnidarian physiology, the mechanisms of action are unknown. Homologs to the vertebrate steroid and thyroid receptors have not been identified in cnidarians, so more research is needed to understand the mechanisms of endocrine-like signaling in cnidarians. Elucidation of cnidarian regulatory pathways will provide insight into evolution of hormonal signaling. These studies will also improve understanding of how cnidarians respond to environmental cues and will provide a basis to investigate disruption of physiological processes by physical and chemical stressors.

Metabolism of estrogens and androgens by scleractinian corals.

Estrogens and androgens are steroids that act as reproductive hormones in vertebrates. These compounds have also been detected in reef-building corals and other invertebrates, where they are hypothesized to act as bioregulatory molecules. Experiments were conducted using labeled steroid substrates to evaluate metabolism of estrogens and androgens by coral homogenates. GC-MS analysis of 13C-labeled steroids showed that Montipora capitata coral homogenates or fragments could convert estradiol to estrone and testosterone to androstenedione and androstanedione, evidence that M. capitata contains 17beta-hydroxysteroid dehydrogenase and 5alpha-reductase. When homogenates from three coral species and symbiotic dinoflagellates (zooxanthellae) were incubated with tritiated steroid substrates, metabolites separated by thin-layer chromatography confirmed that 17beta-hydroxysteroid dehydrogenase activity occurred in all species tested. NADP+ was the preferred cofactor in dehydrogenation reactions with coral homogenates. Reduction of estrone and androstenedione occurred at lower rates and aromatization of androgens was not observed. It is unclear whether estrogens detected previously in coral tissues are produced endogenously or sequestered in coral tissue from dietary or environmental sources. Previous studies have demonstrated that corals can take up estrogens from the water column overlying coral reefs. Considered in total, these observations suggest corals could alter the concentration or form of steroids available to reef organisms.

Estrogen responses in killifish (Fundulus heteroclitus) from polluted and unpolluted environments are site- and gene-specific

Epidemiological, ecological, and laboratory-based studies support the hypothesis that endocrine disrupting chemicals (EDCs) in the environment are responsible for developmental and reproductive abnormalities. We have previously described a killifish population resident in a highly polluted Superfund site (New Bedford Harbor, NBH) that shows evidence of exposure to an estrogenic environment and endocrine disruption. Here, we compare NBH with a local reference population (Scorton Creek, SC) for developmental patterns and direct effects of exogenous estradiol on the estrogenic markers, brain cytochrome P450 aromatase (CYP19A2 or AroB), hepatic vitellogenin (Vtg), and hepatic estrogen receptor alpha (ER alpha). In contrast to our previous observation of elevated ER alpha in NBH embryos, developmental levels of AroB and Vtg mRNAs did not differ between the two sites, demonstrating that not all estrogen-responsive genes are upregulated in NBH embryos. A dose-response experiment showed that NBH larvae are less responsive (lower maximum induction, as measured by ER alpha) and less sensitive (higher EC(50) for induction, as measured by AroB) to estradiol than SC larvae, changes that would be adaptive in an estrogenic environment. In contrast, induction of Vtg mRNA is similar in the two populations, indicating that the adaptive mechanism is target gene-specific. Based on the lower basal levels of ER alpha mRNA in several tissues from adult NBH fish vs SC fish (Greytak and Callard, 2007), we predicted estrogen hyporesponsiveness; however, induction of ER alpha by estradiol exposure in reproductively inactive males did not differ between the two sites. Moreover, AroB was more responsive and Vtg induction was greater (2d) or similar (5d) in NBH as compared to SC males. Worth noting is the high inter-individual variability in estrogen responses of gene targets, especially in NBH killifish, which may indicate evolving preadaptive or adaptive mechanisms. In conclusion, although multi-generational exposure to a highly polluted environment is associated with changes in basal levels of ER alpha mRNA, this is not a simple predictor of estrogen responsiveness. We hypothesize that adaptation of killifish to the estrogenic and polluted environment may be occurring through diverse mechanisms that are gene-, tissue type- and life-stage-specific.

The Active Form of Human Aryl Hydrocarbon Receptor (AHR) Repressor Lacks Exon 8, and Its Pro185 and Ala185 Variants Repress both AHR and Hypoxia-Inducible Factor

ABSTRACT The aryl hydrocarbon receptor (AHR) repressor (AHRR) inhibits AHR-mediated transcription and has been associated with reproductive dysfunction and tumorigenesis in humans. Previous studies have characterized the repressor function of AHRRs from mice and fish, but the human AHRR ortholog (AHRR715) appeared to be nonfunctional in vitro. Here, we report a novel human AHRR cDNA (AHRRΔ8) that lacks exon 8 of AHRR715. AHRRΔ8 was the predominant AHRR form expressed in human tissues and cell lines. AHRRΔ8 effectively repressed AHR-dependent transactivation, whereas AHRR715 was much less active. Similarly, AHRRΔ8, but not AHRR715, formed a complex with AHR nuclear translocator (ARNT). Repression of AHR by AHRRΔ8 was not relieved by overexpression of ARNT or AHR coactivators, suggesting that competition for these cofactors is not the mechanism of repression. AHRRΔ8 interacted weakly with AHR but did not inhibit its nuclear translocation. In a survey of transcription factor specificity, AHRRΔ8 did not repress the nuclear receptor pregnane X receptor or estrogen receptor α but did repress hypoxia-inducible factor (HIF)-dependent signaling. AHRRΔ8-Pro185 and -Ala185 variants, which have been linked to human reproductive disorders, both were capable of repressing AHR or HIF. Together, these results identify AHRRΔ8 as the active form of human AHRR and reveal novel aspects of its function and specificity as a repressor.

Ecdysteroid receptor from the American lobster Homarus americanus: EcR/RXR isoform cloning and ligand-binding properties.

In arthropods, ecdysteroids regulate molting by activating a heterodimer formed by the ecdysone receptor (EcR) and retinoid X receptor (RXR). While this mechanism is similar in insects and crustaceans, variation in receptor splicing, dimerization and ligand affinity adds specificity to molting processes. This study reports the EcR and RXR sequences from American lobster, a commercially and ecologically important crustacean. We cloned two EcR splice variants, both of which specifically bind ponasterone A, and two RXR variants, both of which enhance binding of ponasterone A to the EcR. Lobster EcR has high affinity for ponasterone A and muristerone and moderately high affinity for the insecticide tebufenozide. Bisphenol A, diethyl phthalate, and two polychlorinated biphenyls (PCB 29 and PCB 30), environmental chemicals shown to interfere with crustacean molting, showed little or no affinity for lobster EcR. These studies establish the molecular basis for investigation of lobster ecdysteroid signaling and signal disruption by environmental chemicals.