Rampant asexual reproduction and limited dispersal in a mangrove population of the coral Porites divaricata.

Corals are critical to marine biodiversity. Reproduction and dispersal are key to their resilience, but rarely quantified in nature. Exploiting a unique system-a fully censused, longitudinally characterized, semi-isolated population inhabiting mangroves-we used 2bRAD sequencing to demonstrate that rampant asexual reproduction most likely via parthenogenesis and limited dispersal enable the persistence of a natural population of thin-finger coral (Porites divaricata). Unlike previous studies on coral dispersal, knowledge of colony age and location enabled us to identify plausible parent-offspring relationships within multiple clonal lineages and develop tightly constrained estimates of larval dispersal; the best-fitting model indicates dispersal is largely limited to a few metres from parent colonies. Our results explain why this species is adept at colonizing mangroves but suggest limited genetic diversity in mangrove populations and limited connectivity between mangroves and nearby reefs. As P. divaricata is gonochoristic, and parthenogenesis would be restricted to females (whereas fragmentation, which is presumably common in reef and seagrass habitats, is not), mangrove populations likely exhibit skewed sex ratios. These findings suggest that coral reproductive diversity can lead to distinctly different demographic outcomes in different habitats. Thus, coral conservation will require the protection of the entire coral habitat mosaic, and not just reefs.

Laparoscopic entry techniques.

BACKGROUND: Laparoscopy is a common procedure in many surgical specialties. Complications arising from laparoscopy are often related to initial entry into the abdomen. Life-threatening complications include injury to viscera (e.g. bowel, bladder) or to vasculature (e.g. major abdominal and anterior abdominal wall vessels). No clear consensus has been reached as to the optimal method of laparoscopic entry into the peritoneal cavity. OBJECTIVES: To evaluate the benefits and risks of different laparoscopic entry techniques in gynaecological and non-gynaecological surgery. SEARCH METHODS: We searched the Cochrane Gynaecology and Fertility (CGF) Group trials register, CENTRAL, MEDLINE, Embase, PsycINFO, and trials registers in January 2018. We also checked the references of articles retrieved. SELECTION CRITERIA: We included randomised controlled trials (RCTs) that compared one laparoscopic entry technique versus another. Primary outcomes were major complications including mortality, vascular injury of major vessels and abdominal wall vessels, visceral injury of bladder or bowel, gas embolism, solid organ injury, and failed entry (inability to access the peritoneal cavity). Secondary outcomes were extraperitoneal insufflation, trocar site bleeding, trocar site infection, incisional hernia, omentum injury, and uterine bleeding. DATA COLLECTION AND ANALYSIS: Two review authors independently selected studies, assessed risk of bias, and extracted data. We expressed findings as Peto odds ratios (Peto ORs) with 95% confidence intervals (CIs). We assessed statistical heterogeneity using the I² statistic. We assessed the overall quality of evidence for the main comparisons using GRADE methods. MAIN RESULTS: The review included 57 RCTs including four multi-arm trials, with a total of 9865 participants, and evaluated 25 different laparoscopic entry techniques. Most studies selected low-risk patients, and many studies excluded patients with high body mass index (BMI) and previous abdominal surgery. Researchers did not find evidence of differences in major vascular or visceral complications, as would be anticipated given that event rates were very low and sample sizes were far too small to identify plausible differences in rare but serious adverse events.Open-entry versus closed-entryTen RCTs investigating Veress needle entry reported vascular injury as an outcome. There was a total of 1086 participants and 10 events of vascular injury were reported. Four RCTs looking at open entry technique reported vascular injury as an outcome. There was a total of 376 participants and 0 events of vascular injury were reported. This was not a direct comparison. In the direct comparison of Veress needle and Open-entry technique, there was insufficient evidence to determine whether there was a difference in rates of vascular injury (Peto OR 0.14, 95% CI 0.00 to 6.82; 4 RCTs; n = 915; I² = N/A, very low-quality evidence). Evidence was insufficient to show whether there were differences between groups for visceral injury (Peto OR 0.61, 95% CI 0.06 to 6.08; 4 RCTs; n = 915: I² = 0%; very low-quality evidence), or failed entry (Peto OR 0.45, 95% CI 0.14 to 1.42; 3 RCTs; n = 865; I² = 63%; very low-quality evidence). Two studies reported mortality with no events in either group. No studies reported gas embolism or solid organ injury.Direct trocar versus Veress needle entryTrial results show a reduction in failed entry into the abdomen with the use of a direct trocar in comparison with Veress needle entry (OR 0.24, 95% CI 0.17 to 0.34; 8 RCTs; N = 3185; I² = 45%; moderate-quality evidence). Evidence was insufficient to show whether there were differences between groups in rates of vascular injury (Peto OR 0.59, 95% CI 0.18 to 1.96; 6 RCTs; n = 1603; I² = 75%; very low-quality evidence), visceral injury (Peto OR 2.02, 95% CI 0.21 to 19.42; 5 RCTs; n = 1519; I² = 25%; very low-quality evidence), or solid organ injury (Peto OR 0.58, 95% Cl 0.06 to 5.65; 3 RCTs; n = 1079; I² = 61%; very low-quality evidence). Four studies reported mortality with no events in either group. Two studies reported gas embolism, with no events in either group.Direct vision entry versus Veress needle entryEvidence was insufficient to show whether there were differences between groups in rates of vascular injury (Peto OR 0.39, 95% CI 0.05 to 2.85; 1 RCT; n = 186; very low-quality evidence) or visceral injury (Peto OR 0.15, 95% CI 0.01 to 2.34; 2 RCTs; n = 380; I² = N/A; very low-quality evidence). Trials did not report our other primary outcomes.Direct vision entry versus open entryEvidence was insufficient to show whether there were differences between groups in rates of visceral injury (Peto OR 0.13, 95% CI 0.00 to 6.50; 2 RCTs; n = 392; I² = N/A; very low-quality evidence), solid organ injury (Peto OR 6.16, 95% CI 0.12 to 316.67; 1 RCT; n = 60; very low-quality evidence), or failed entry (Peto OR 0.40, 95% CI 0.04 to 4.09; 1 RCT; n = 60; very low-quality evidence). Two studies reported vascular injury with no events in either arm. Trials did not report our other primary outcomes.Radially expanding (STEP) trocars versus non-expanding trocarsEvidence was insufficient to show whether there were differences between groups in rates of vascular injury (Peto OR 0.24, 95% Cl 0.05 to 1.21; 2 RCTs; n = 331; I² = 0%; very low-quality evidence), visceral injury (Peto OR 0.13, 95% CI 0.00 to 6.37; 2 RCTs; n = 331; very low-quality evidence), or solid organ injury (Peto OR 1.05, 95% CI 0.07 to 16.91; 1 RCT; n = 244; very low-quality evidence). Trials did not report our other primary outcomes.Other studies compared a wide variety of other laparoscopic entry techniques, but all evidence was of very low quality and evidence was insufficient to support the use of one technique over another. AUTHORS' CONCLUSIONS: Overall, evidence was insufficient to support the use of one laparoscopic entry technique over another. Researchers noted an advantage of direct trocar entry over Veress needle entry for failed entry. Most evidence was of very low quality; the main limitations were imprecision (due to small sample sizes and very low event rates) and risk of bias associated with poor reporting of study methods.

Nitrate enhances skeletal muscle fatty acid oxidation via a nitric oxide-cGMP-PPAR-mediated mechanism.

BACKGROUND: Insulin sensitivity in skeletal muscle is associated with metabolic flexibility, including a high capacity to increase fatty acid (FA) oxidation in response to increased lipid supply. Lipid overload, however, can result in incomplete FA oxidation and accumulation of potentially harmful intermediates where mitochondrial tricarboxylic acid cycle capacity cannot keep pace with rates of ß-oxidation. Enhancement of muscle FA oxidation in combination with mitochondrial biogenesis is therefore emerging as a strategy to treat metabolic disease. Dietary inorganic nitrate was recently shown to reverse aspects of the metabolic syndrome in rodents by as yet incompletely defined mechanisms. RESULTS: Herein, we report that nitrate enhances skeletal muscle FA oxidation in rodents in a dose-dependent manner. We show that nitrate induces FA oxidation through a soluble guanylate cyclase (sGC)/cGMP-mediated PPARß/δ- and PPARα-dependent mechanism. Enhanced PPARß/δ and PPARα expression and DNA binding induces expression of FA oxidation enzymes, increasing muscle carnitine and lowering tissue malonyl-CoA concentrations, thereby supporting intra-mitochondrial pathways of FA oxidation and enhancing mitochondrial respiration. At higher doses, nitrate induces mitochondrial biogenesis, further increasing FA oxidation and lowering long-chain FA concentrations. Meanwhile, nitrate did not affect mitochondrial FA oxidation in PPARα(-/-) mice. In C2C12 myotubes, nitrate increased expression of the PPARα targets Cpt1b, Acadl, Hadh and Ucp3, and enhanced oxidative phosphorylation rates with palmitoyl-carnitine; however, these changes in gene expression and respiration were prevented by inhibition of either sGC or protein kinase G. Elevation of cGMP, via the inhibition of phosphodiesterase 5 by sildenafil, also increased expression of Cpt1b, Acadl and Ucp3, as well as CPT1B protein levels, and further enhanced the effect of nitrate supplementation. CONCLUSIONS: Nitrate may therefore be effective in the treatment of metabolic disease by inducing FA oxidation in muscle.

NF-κB is required for cnidocyte development in the sea anemone Nematostella vectensis.

The sea anemone Nematostella vectensis (Nv) is a leading model organism for the phylum Cnidaria, which includes anemones, corals, jellyfishes and hydras. A defining trait across this phylum is the cnidocyte, an ectodermal cell type with a variety of functions including defense, prey capture and environmental sensing. Herein, we show that the Nv-NF-κB transcription factor and its inhibitor Nv-IκB are expressed in a subset of cnidocytes in the body column of juvenile and adult anemones. The size and distribution of the Nv-NF-κB-positive cnidocytes suggest that they are in a subtype known as basitrichous haplonema cnidocytes. Nv-NF-κB is primarily cytoplasmic in cnidocytes in juvenile and adult animals, but is nuclear when first detected in the 30-h post-fertilization embryo. Morpholino-mediated knockdown of Nv-NF-κB expression results in greatly reduced cnidocyte formation in the 5 day-old animal. Taken together, these results indicate that NF-κB plays a key role in the development of the phylum-specific cnidocyte cell type in Nematostella, likely by nuclear Nv-NF-κB-dependent activation of genes required for cnidocyte development.

Production of a reference transcriptome and transcriptomic database (EdwardsiellaBase) for the lined sea anemone, Edwardsiella lineata, a parasitic cnidarian.

BACKGROUND: The lined sea anemone Edwardsiella lineata is an informative model system for evolutionary-developmental studies of parasitism. In this species, it is possible to compare alternate developmental pathways leading from a larva to either a free-living polyp or a vermiform parasite that inhabits the mesoglea of a ctenophore host. Additionally, E. lineata is confamilial with the model cnidarian Nematostella vectensis, providing an opportunity for comparative genomic, molecular and organismal studies. DESCRIPTION: We generated a reference transcriptome for E. lineata via high-throughput sequencing of RNA isolated from five developmental stages (parasite; parasite-to-larva transition; larva; larva-to-adult transition; adult). The transcriptome comprises 90,440 contigs assembled from >15 billion nucleotides of DNA sequence. Using a molecular clock approach, we estimated the divergence between E. lineata and N. vectensis at 215-364 million years ago. Based on gene ontology and metabolic pathway analyses and gene family surveys (bHLH-PAS, deiodinases, Fox genes, LIM homeodomains, minicollagens, nuclear receptors, Sox genes, and Wnts), the transcriptome of E. lineata is comparable in depth and completeness to N. vectensis. Analyses of protein motifs and revealed extensive conservation between the proteins of these two edwardsiid anemones, although we show the NF-κB protein of E. lineata reflects the ancestral structure, while the NF-κB protein of N. vectensis has undergone a split that separates the DNA-binding domain from the inhibitory domain. All contigs have been deposited in a public database (EdwardsiellaBase), where they may be searched according to contig ID, gene ontology, protein family motif (Pfam), enzyme commission number, and BLAST. The alignment of the raw reads to the contigs can also be visualized via JBrowse. CONCLUSIONS: The transcriptomic data and database described here provide a platform for studying the evolutionary developmental genomics of a derived parasitic life cycle. In addition, these data from E. lineata will aid in the interpretation of evolutionary novelties in gene sequence or structure that have been reported for the model cnidarian N. vectensis (e.g., the split NF-κB locus). Finally, we include custom computational tools to facilitate the annotation of a transcriptome based on high-throughput sequencing data obtained from a "non-model system."

A report from the second Nematostella vectensis research conference.

This report summarizes information discussed at the second Nematostella vectensis research conference, which took place on August 27, 2012 in Boston, MA, USA. The startlet sea anemone Nematostella is emerging as one of leading model organisms among cnidarians, in part because of the extensive genome and transcriptome resources that are becoming available for Nematostella, which were the focus of several presentations. In addition, research was presented on the use of Nematostella in developmental, regeneration, signal transduction, host-symbiont, and gene-environment interaction studies.

TEMPERATURE AND SALINITY AFFECT DEVELOPMENT OF THE PARASITIC SEA ANEMONE EDWARDSIELLA LINEATA POTENTIALLY LIMITING ITS IMPACT AS A BIOLOGICAL CONTROL ON THE CTENOPHORE MNEMIOPSIS LEIDYI.

The lined sea anemone, Edwardsiella lineata, parasitizes the ctenophore Mnemiopsis leidyi, which is one of the most destructive marine invasive species in the world. Mnemiopsis leidyi is known to tolerate a wide range of environmental conditions. However, the environmental tolerances of its most prominent parasite have never been characterized. Here we determined the effects of temperature (18, 22, 26, and 30 C) and salinity (6, 15, 24, and 33 ppt) on the survival and development of E. lineata from a vermiform parasite to a free-living polyp. At higher temperatures and lower salinities, E. lineata experienced significantly higher mortality, and it failed to develop into an adult polyp at the highest temperature (30 C) and lowest salinities we tested (6 ppt or 15 ppt). While such temperature and salinity restrictions would not currently prevent E. lineata from infecting M. leidyi in many of the European waters where it has become a destructive invasive species, these environmental limitations may be reducing overlap between host and parasite within the host's native range, a situation that could be exacerbated by climate change.

Evolution of function of a fused metazoan tRNA synthetase.

The origin and evolution of multidomain proteins are driven by diverse processes including fusion/fission, domain shuffling, and alternative splicing. The 20 aminoacyl-tRNA synthetases (AARS) constitute an ancient conserved family of multidomain proteins. The glutamyl-prolyl tRNA synthetase (EPRS) of bilaterian animals is unique among AARSs, containing two functional enzymes catalyzing ligation of glutamate and proline to their cognate transfer RNAs (tRNAs). The ERS and PRS catalytic domains in multiple bilaterian taxa are linked by variable number of helix-turn-helix domains referred to as WHEP-TRS domains. In addition to its canonical aminoacylation activities, human EPRS exhibits a noncanonical function as an inflammation-responsive regulator of translation. Recently, we have shown that the WHEP domains direct this auxiliary function of human EPRS by interacting with an mRNA stem-loop element (interferon-gamma-activated inhibitor of translation [GAIT] element). Here, we show that EPRS is present in the cnidarian Nematostella vectensis, which pushes the origin of the fused protein back to the cnidarian-bilaterian ancestor, 50-75 My before the origin of the Bilateria. Remarkably, the Nematostella EPRS mRNA is alternatively spliced to yield three isoforms with variable number and sequence of WHEP domains and with distinct RNA-binding activities. Whereas one isoform containing a single WHEP domain binds tRNA, a second binds both tRNA and GAIT element RNA. However, the third isoform contains two WHEP domains and like the human ortholog binds specifically to GAIT element RNA. These results suggest that alternative splicing of WHEP domains in the EPRS gene of the cnidarian-bilaterian ancestor gave rise to a novel molecular function of EPRS conserved during metazoan evolution.

Integration of Patient-reported Outcomes Assessment Into Routine Care for Patients Receiving Residential Treatment for Alcohol and/or Substance Use Disorder.

BACKGROUND: More than 3 million individuals receive treatment for alcohol use disorder (AUD) and/or substance use disorder each year, yet there exists no standardized method for measuring patient success in treatment. Quantifying a more comprehensive assessment of treatment outcomes could identify the relative efficacy of different treatment strategies for individuals with AUD/substance use disorders, and help patients to identify, in advance, appropriate treatment options. METHODS: This study developed and embedded patient-reported outcome measures into the routine clinical operations of a residential treatment program. Surveys assessed demographics, drug use history, physical and mental health, and quality of life. Outcomes were assessed among participants at admission (n = 961) and in patients who completed the survey at time of discharge (n = 633). RESULTS: Past 30-day alcohol and/or opioid use at admission were correlated with worse self-reported physical and mental health, sleep, and quality of life, and greater negative affect and craving ( P s < 0.05). Previous history of treatment and/or withdrawal management were associated with worse self-reported physical and mental health, quality of life, and increased craving ( P s < 0.05). Physical and mental health improved across timepoints and was most pronounced when comparing persons receiving treatment for opioid use disorder versus AUD, wherein persons with opioid use disorder had worse physical health at all time points, and greater sleep disturbance and negative affect at discharge ( P s < 0.05). CONCLUSIONS: It is feasible to embed patient outcome monitoring into routine clinic operations, which could be used in the future to tailor treatment plans.

Production of a reference transcriptome and transcriptomic database (PocilloporaBase) for the cauliflower coral, Pocillopora damicornis.

BACKGROUND: Motivated by the precarious state of the world's coral reefs, there is currently a keen interest in coral transcriptomics. By identifying changes in coral gene expression that are triggered by particular environmental stressors, we can begin to characterize coral stress responses at the molecular level, which should lead to the development of more powerful diagnostic tools for evaluating the health of corals in the field. Furthermore, the identification of genetic variants that are more or less resilient in the face of particular stressors will help us to develop more reliable prognoses for particular coral populations. Toward this end, we performed deep mRNA sequencing of the cauliflower coral, Pocillopora damicornis, a geographically widespread Indo-Pacific species that exhibits a great diversity of colony forms and is able to thrive in habitats subject to a wide range of human impacts. Importantly, P. damicornis is particularly amenable to laboratory culture. We collected specimens from three geographically isolated Hawaiian populations subjected to qualitatively different levels of human impact. We isolated RNA from colony fragments ("nubbins") exposed to four environmental stressors (heat, desiccation, peroxide, and hypo-saline conditions) or control conditions. The RNA was pooled and sequenced using the 454 platform. DESCRIPTION: Both the raw reads (n=1, 116, 551) and the assembled contigs (n=70, 786; mean length=836 nucleotides) were deposited in a new publicly available relational database called PocilloporaBase http://www.PocilloporaBase.org. Using BLASTX, 47.2% of the contigs were found to match a sequence in the NCBI database at an E-value threshold of ≤.001; 93.6% of those contigs with matches in the NCBI database appear to be of metazoan origin and 2.3% bacterial origin, while most of the remaining 4.1% match to other eukaryotes, including algae and amoebae. CONCLUSIONS: P. damicornis now joins the handful of coral species for which extensive transcriptomic data are publicly available. Through PocilloporaBase http://www.PocilloporaBase.org, one can obtain assembled contigs and raw reads and query the data according to a wide assortment of attributes including taxonomic origin, PFAM motif, KEGG pathway, and GO annotation.

Two polymorphic residues account for the differences in DNA binding and transcriptional activation by NF-κB proteins encoded by naturally occurring alleles in Nematostella vectensis.

The NF-κB family of transcription factors is activated in response to many environmental and biological stresses, and plays a key role in innate immunity across a broad evolutionary expanse of animals. A simple NF-κB pathway is present in the sea anemone Nematostella vectensis, an important model organism in the phylum Cnidaria. Nematostella has previously been shown to have two naturally occurring NF-κB alleles (Nv-NF-κB-C and Nv-NF-κB-S) that encode proteins with different DNA-binding and transactivation abilities. We show here that polymorphic residues 67 (Cys vs. Ser) and 269 (Ala vs. Glu) play complementary roles in determining the DNA-binding activity of the NF-κB proteins encoded by these two alleles and that residue 67 is primarily responsible for the difference in their transactivation ability. Phylogenetic analysis indicates that Nv-NF-κB-S is the derived allele, consistent with its restricted geographic distribution. These results define polymorphic residues that are important for the DNA-binding and transactivating activities of two naturally occurring variants of Nv-NF-κB. The implications for the appearance of the two Nv-NF-κB alleles in natural populations of sea anemones are discussed.

Distinct Phenotypes Associated with Mangrove and Lagoon Habitats in Two Widespread Caribbean Corals, Porites astreoides and Porites divaricata.

AbstractAs coral reefs experience dramatic declines in coral cover throughout the tropics, there is an urgent need to understand the role that non-reef habitats, such as mangroves, play in the ecological niche of corals. Mangrove habitats present a challenge to reef-dwelling corals because they can differ dramatically from adjacent reef habitats with respect to key environmental parameters, such as light. Because variation in light within reef habitats is known to drive intraspecific differences in coral phenotype, we hypothesized that coral species that can exploit both reef and mangrove habitats will exhibit predictable differences in phenotypes between habitats. To investigate how intraspecific variation, driven by either local adaptation or phenotypic plasticity, might enable particular coral species to exploit these two qualitatively different habitat types, we compared the phenotypes of two widespread Caribbean corals, Porites divaricata and Porites astreoides, in mangrove versus lagoon habitats on Turneffe Atoll, Belize. We document significant differences in colony size, color, structural complexity, and corallite morphology between habitats. In every instance, the phenotypic differences between mangrove prop root and lagoon corals exhibited consistent trends in both P. divaricata and P. astreoides. We believe this study is the first to document intraspecific phenotypic diversity in corals occupying mangrove prop root versus lagoonal patch reef habitats. A difference in the capacity to adopt an alternative phenotype that is well suited to the mangrove habitat may explain why some reef coral species can exploit mangroves, while others cannot.

The evolutionary diversification of LSF and Grainyhead transcription factors preceded the radiation of basal animal lineages.

BACKGROUND: The transcription factors of the LSF/Grainyhead (GRH) family are characterized by the possession of a distinctive DNA-binding domain that bears no clear relationship to other known DNA-binding domains, with the possible exception of the p53 core domain. In triploblastic animals, the LSF and GRH subfamilies have diverged extensively with respect to their biological roles, general expression patterns, and mechanism of DNA binding. For example, Grainyhead (GRH) homologs are expressed primarily in the epidermis, and they appear to play an ancient role in maintaining the epidermal barrier. By contrast, LSF homologs are more widely expressed, and they regulate general cellular functions such as cell cycle progression and survival in addition to cell-lineage specific gene expression. RESULTS: To illuminate the early evolution of this family and reconstruct the functional divergence of LSF and GRH, we compared homologs from 18 phylogenetically diverse taxa, including four basal animals (Nematostella vectensis, Vallicula multiformis, Trichoplax adhaerens, and Amphimedon queenslandica), a choanoflagellate (Monosiga brevicollis) and several fungi. Phylogenetic and bioinformatic analyses of these sequences indicate that (1) the LSF/GRH gene family originated prior to the animal-fungal divergence, and (2) the functional diversification of the LSF and GRH subfamilies occurred prior to the divergence between sponges and eumetazoans. Aspects of the domain architecture of LSF/GRH proteins are well conserved between fungi, choanoflagellates, and metazoans, though within the Metazoa, the LSF and GRH families are clearly distinct. We failed to identify a convincing LSF/GRH homolog in the sequenced genomes of the algae Volvox carteri and Chlamydomonas reinhardtii or the amoebozoan Dictyostelium purpureum. Interestingly, the ancestral GRH locus has become split into two separate loci in the sea anemone Nematostella, with one locus encoding a DNA binding domain and the other locus encoding the dimerization domain. CONCLUSIONS: In metazoans, LSF and GRH proteins play a number of roles that are essential to achieving and maintaining multicellularity. It is now clear that this protein family already existed in the unicellular ancestor of animals, choanoflagellates, and fungi. However, the diversification of distinct LSF and GRH subfamilies appears to be a metazoan invention. Given the conserved role of GRH in maintaining epithelial integrity in vertebrates, insects, and nematodes, it is noteworthy that the evolutionary origin of Grh appears roughly coincident with the evolutionary origin of the epithelium.

Upgrades to StellaBase facilitate medical and genetic studies on the starlet sea anemone, Nematostella vectensis.

The starlet sea anemone, Nematostella vectensis, is a basal metazoan organism that has recently emerged as an important model system in developmental biology and evolutionary genomics. StellaBase, the Nematostella Genomics Database (http://stellabase.org), was developed in 2005 as a resource to support the Nematostella research community. Recently, it has become apparent that Nematostella may be a particularly useful system for studying (i) microevolutionary variation in natural populations, and (ii) the functional evolution of human disease genes. We have developed two new databases that will foster such studies: StellaBase Disease (http://stellabase.org/disease) is a relational database that houses 155 904 invertebrate homologous isoforms of human disease genes from four leading genomic model systems (fly, worm, yeast and Nematostella), including 14 874 predicted genes from the sea anemone itself. StellaBase SNP (http://stellabase.org/SNP) is a relational database that describes the location and underlying type of mutation for 20 063 single nucleotide polymorphisms.

Domain duplication, divergence, and loss events in vertebrate Msx paralogs reveal phylogenomically informed disease markers.

BACKGROUND: Msx originated early in animal evolution and is implicated in human genetic disorders. To reconstruct the functional evolution of Msx and inform the study of human mutations, we analyzed the phylogeny and synteny of 46 metazoan Msx proteins and tracked the duplication, diversification and loss of conserved motifs. RESULTS: Vertebrate Msx sequences sort into distinct Msx1, Msx2 and Msx3 clades. The sister-group relationship between MSX1 and MSX2 reflects their derivation from the 4p/5q chromosomal paralogon, a derivative of the original "MetaHox" cluster. We demonstrate physical linkage between Msx and other MetaHox genes (Hmx, NK1, Emx) in a cnidarian. Seven conserved domains, including two Groucho repression domains (N- and C-terminal), were present in the ancestral Msx. In cnidarians, the Groucho domains are highly similar. In vertebrate Msx1, the N-terminal Groucho domain is conserved, while the C-terminal domain diverged substantially, implying a novel function. In vertebrate Msx2 and Msx3, the C-terminal domain was lost. MSX1 mutations associated with ectodermal dysplasia or orofacial clefting disorders map to conserved domains in a non-random fashion. CONCLUSION: Msx originated from a MetaHox ancestor that also gave rise to Tlx, Demox, NK, and possibly EHGbox, Hox and ParaHox genes. Duplication, divergence or loss of domains played a central role in the functional evolution of Msx. Duplicated domains allow pleiotropically expressed proteins to evolve new functions without disrupting existing interaction networks. Human missense sequence variants reside within evolutionarily conserved domains, likely disrupting protein function. This phylogenomic evaluation of candidate disease markers will inform clinical and functional studies.

Concerted evolution of sea anemone neurotoxin genes is revealed through analysis of the Nematostella vectensis genome.

Gene families, which encode toxins, are found in many poisonous animals, yet there is limited understanding of their evolution at the nucleotide level. The release of the genome draft sequence for the sea anemone Nematostella vectensis enabled a comprehensive study of a gene family whose neurotoxin products affect voltage-gated sodium channels. All gene family members are clustered in a highly repetitive approximately 30-kb genomic region and encode a single toxin, Nv1. These genes exhibit extreme conservation at the nucleotide level which cannot be explained by purifying selection. This conservation greatly differs from the toxin gene families of other animals (e.g., snakes, scorpions, and cone snails), whose evolution was driven by diversifying selection, thereby generating a high degree of genetic diversity. The low nucleotide diversity at the Nv1 genes is reminiscent of that reported for DNA encoding ribosomal RNA (rDNA) and 2 hsp70 genes from Drosophila, which have evolved via concerted evolution. This evolutionary pattern was experimentally demonstrated in yeast rDNA and was shown to involve unequal crossing-over. Through sequence analysis of toxin genes from multiple N. vectensis populations and 2 other anemone species, Anemonia viridis and Actinia equina, we observed that the toxin genes for each sea anemone species are more similar to one another than to those of other species, suggesting they evolved by manner of concerted evolution. Furthermore, in 2 of the species (A. viridis and A. equina) we found genes that evolved under diversifying selection, suggesting that concerted evolution and accelerated evolution may occur simultaneously.

Conserved and novel gene expression between regeneration and asexual fission in Nematostella vectensis.

Due to work in model systems (e.g., flies and mice), the molecular mechanisms of embryogenesis are known in exquisite detail. However, these organisms are incapable of asexual reproduction and possess limited regenerative abilities. Thus, the mechanisms of alternate developmental trajectories and their relation to embryonic mechanisms remain understudied. Because these developmental trajectories are present in a diverse group of animal phyla spanning the metazoan phylogeny, including cnidarians, annelids, and echinoderms, they are likely to have played a major role in animal evolution. The starlet sea anemone Nematostella vectensis, an emerging model system, undergoes larval development, asexual fission, and complete bi-directional regeneration in the field and laboratory. In order to investigate to what extent embryonic patterning mechanisms are utilized during alternate developmental trajectories, we examined expression of developmental regulatory genes during regeneration and fission. When compared to previously reported embryonic expression patterns, we found that all genes displayed some level of expression consistent with embryogenesis. However, five of seven genes investigated also displayed striking differences in gene expression between one or more developmental trajectory. These results demonstrate that alternate developmental trajectories utilize distinct molecular mechanisms upstream of major developmental regulatory genes such as fox, otx, and Hox-like.

Comparative anatomy and histology of developmental and parasitic stages in the life cycle of the lined sea anemone Edwardsiella lineata.

The evolution of parasitism is often accompanied by profound changes to the developmental program. However, relatively few studies have directly examined the developmental evolution of parasitic species from free-living ancestors. The lined sea anemone Edwardsiella lineata is a relatively recently evolved parasite for which closely related free-living outgroups are known, including the starlet sea anemone Nematostella vectensis. The larva of E. lineata parasitizes the ctenophore Mnemiopsis leidyi, and, once embedded in its host, the anemone assumes a novel vermiform body plan. That we might begin to understand how the developmental program of this species has been transformed during the evolution of parasitism, we characterized the gross anatomy, histology, and cnidom of the parasitic stage, post-parasitic larval stage, and adult stage of the E. lineata life cycle. The distinct parasitic stage of the life cycle differs from the post-parasitic larva with respect to overall shape, external ciliation, cnida frequency, and tissue architecture. The parasitic stage and planula both contain holotrichs, a type of cnida not previously reported in Edwardsiidae. The internal morphology of the post-parasitic planula is extremely similar to the adult morphology, with a complete set of mesenterial tissue and musculature despite this stage having little external differentiation. Finally, we observed 2 previously undocumented aspects of asexual reproduction in E. lineata: (1) the parasitic stage undergoes transverse fission via physal pinching, the first report of asexual reproduction in a pre-adult stage in the Edwardsiidae; and (2) the juvenile polyp undergoes transverse fission via polarity reversal, the first time this form of fission has been reported in E. lineata.

The evolutionary origin of the Runx/CBFbeta transcription factors--studies of the most basal metazoans.

BACKGROUND: Members of the Runx family of transcriptional regulators, which bind DNA as heterodimers with CBFbeta, are known to play critical roles in embryonic development in many triploblastic animals such as mammals and insects. They are known to regulate basic developmental processes such as cell fate determination and cellular potency in multiple stem-cell types, including the sensory nerve cell progenitors of ganglia in mammals. RESULTS: In this study, we detect and characterize the hitherto unexplored Runx/CBFbeta genes of cnidarians and sponges, two basal animal lineages that are well known for their extensive regenerative capacity. Comparative structural modeling indicates that the Runx-CBFbeta-DNA complex from most cnidarians and sponges is highly similar to that found in humans, with changes in the residues involved in Runx-CBFbeta dimerization in either of the proteins mirrored by compensatory changes in the binding partner. In situ hybridization studies reveal that Nematostella Runx and CBFbeta are expressed predominantly in small isolated foci at the base of the ectoderm of the tentacles in adult animals, possibly representing neurons or their progenitors. CONCLUSION: These results reveal that Runx and CBFbeta likely functioned together to regulate transcription in the common ancestor of all metazoans, and the structure of the Runx-CBFbeta-DNA complex has remained extremely conserved since the human-sponge divergence. The expression data suggest a hypothesis that these genes may have played a role in nerve cell differentiation or maintenance in the common ancestor of cnidarians and bilaterians.

StellaBase: the Nematostella vectensis Genomics Database.

StellaBase, the Nematostella vectensis Genomics Database, is a web-based resource that will facilitate desktop and bench-top studies of the starlet sea anemone. Nematostella is an emerging model organism that has already proven useful for addressing fundamental questions in developmental evolution and evolutionary genomics. StellaBase allows users to query the assembled Nematostella genome, a confirmed gene library, and a predicted genome using both keyword and homology based search functions. Data provided by these searches will elucidate gene family evolution in early animals. Unique research tools, including a Nematostella genetic stock library, a primer library, a literature repository and a gene expression library will provide support to the burgeoning Nematostella research community. The development of StellaBase accompanies significant upgrades to CnidBase, the Cnidarian Evolutionary Genomics Database. With the completion of the first sequenced cnidarian genome, genome comparison tools have been added to CnidBase. In addition, StellaBase provides a framework for the integration of additional species-specific databases into CnidBase. StellaBase is available at http://www.stellabase.org.