Preliminary hazard assessment of a new nature-inspired antifouling (NIAF) agent.

A recently synthesized aminated 3,4-dioxygenated xanthone (Xantifoul2) was found to have promising antifouling (AF) effects against the settlement of the macrofouler Mytilus galloprovincialis larvae. Preliminary assessment indicated that Xantifoul2 has reduced ecotoxicological impacts: e.g., being non-toxic to the marine crustacea Artemia salina (<10 % mortality at 50 µM) and showing low bioconcentration factor in marine organisms. In order to meet the EU Biocidal Product Regulation, a preliminary hazard assessment of this new nature-inspired antifouling (NIAF) agent was conducted in this work. Xantifoul2 did not affect the swimming ability of the planktonic crustacean Daphnia magna, the growth of the diatom Phaeodactylum tricornutum, and the cellular respiration of luminescent Gram-negative bacteria Vibrio fischeri, supporting the low toxicity towards several non-target marine species. Regarding human cytotoxicity, Xantifoul2 did not affect the cell viability of retinal human cells (hTERT-RPE-1) and lipidomic studies revealed depletion of lipids involved in cell death, membrane modeling, lipid storage, and oxidative stress only at a high concentration (10 µM). Accelerated degradation studies in water were conducted under simulated sunlight to allow the understanding of putative transformation products (TPs) that could be generated in the aquatic ecosystems. Both Xantifoul2 and photolytic-treated Xantifoul2 in the aqueous matrix were therefore evaluated on several nuclear receptors (NRs). The results of this preliminary hazard assessment of Xantifoul2, combined with the high degradation rates in water, provide strong evidence of the safety of this AF agent under the evaluated conditions, and provide the support for future validation studies before this compound can be introduced in the market.

An historical "wreck": A transcriptome assembly of the naval shipworm, Teredo navalis Linnaeus, 1978.

Historically famous for their negative impact on human-built marine wood structures, mollusc shipworms play a central ecological role in marine ecosystems. Their association with bacterial symbionts, providing cellulolytic and nitrogen-fixing activities, underscores their exceptional wood-eating and wood-boring behaviours, improving energy transfer and the recycling of essential nutrients locked in the wood cellulose. Importantly, from a molecular standpoint, a minute of omic resources are available from this lineage of Bivalvia. Here, we produced and assembled a transcriptome from the globally distributed naval shipworm, Teredo navalis (family Teredinidae). The transcriptome was obtained by sequencing the total RNA from five equidistant segments of the whole body of a T. navalis specimen. The quality of the produced assembly was accessed with several statistics, revealing a highly contiguous (1194 N50) and complete (over 90% BUSCO scores for Eukaryote and Metazoan databases) transcriptome, with nearly 38,000 predicted ORF, more than half being functionally annotated. Our findings pave the way to investigate the unique evolutionary biology of these highly modified bivalves and lay the foundation for an adequate gene annotation of a full genome sequence of the species.

A whole-body transcriptome assembly of the annelid worm Hediste diversicolor.

The Annelida phylum is composed of a myriad of species exhibiting key phenotypic adaptations. They occupy key ecological niches in a variety of marine, freshwater and terrestrial ecosystems. Importantly, the increment of omic resources is rapidly modifying the taxonomic landscape and knowledge of species belonging to this phylum. Here, we comprehensively characterised and annotated a transcriptome of the common ragworm, Hediste diversicolor (OF Müller). This species belongs to the family Nereididae and inhabits estuarine and lagoon areas on the Atlantic coasts of Europe and North America. Ecologically, H. diversicolor plays an important role in benthic food webs. Given its commercial value, H. diversicolor is a promising candidate for aquaculture development and production in farming facilities, under a circular economy framework. We used Illumina next-generation sequencing technology, to produce a total of 105 million (M) paired-end (PE) raw reads and generate the first whole-body transcriptome assembly of H. diversicolor species. This high-quality transcriptome contains 69,335 transcripts with an N50 transcript length of 2313 bp and achieved a BUSCO gene completeness of 97.7% and 96% in Eukaryota and Metazoa lineage-specific profile libraries. Our findings offer a valuable resource for multiple biological applications using this species.

Extensive gene loss parallels kidney aglomerulism in Syngnathidae.

The eccentric seahorses, seadragons, pipehorses and pipefishes (Syngnathidae) have an aglomerular kidney1. Here, we show that nephron genes2 conserved in Bilateria are secondarily eroded/deleted in Syngnathidae genomes. A transcriptome enrichment analysis suggests the predominance of excretion processes in the Syngnathidae kidney. In a lineage where crypsis and idleness are tightly associated, we propose that aglomerulism evolved as an energy-saving strategy.

Decay of Skin-Specific Gene Modules in Pangolins.

The mammalian skin exhibits a rich spectrum of evolutionary adaptations. The pilosebaceous unit, composed of the hair shaft, follicle, and the sebaceous gland, is the most striking synapomorphy. The evolutionary diversification of mammals across different ecological niches was paralleled by the appearance of an ample variety of skin modifications. Pangolins, order Pholidota, exhibit keratin-derived scales, one of the most iconic skin appendages. This formidable armor is intended to serve as a deterrent against predators. Surprisingly, while pangolins have hair on their abdomens, the occurrence of sebaceous and sweat glands is contentious. Here, we explore various molecular modules of skin physiology in four pangolin genomes, including that of sebum production. We show that genes driving wax monoester formation, Awat1/2, show patterns of inactivation in the stem pangolin branch, while the triacylglycerol synthesis gene Dgat2l6 seems independently eroded in the African and Asian clades. In contrast, Elovl3 implicated in the formation of specific neutral lipids required for skin barrier function is intact and expressed in the pangolin skin. An extended comparative analysis shows that genes involved in skin pathogen defense and structural integrity of keratinocyte layers also show inactivating mutations: associated with both ancestral and independent pseudogenization events. Finally, we deduce that the suggested absence of sweat glands is not paralleled by the inactivation of the ATP-binding cassette transporter Abcc11, as previously described in Cetacea. Our findings reveal the sophisticated and complex history of gene retention and loss as key mechanisms in the evolution of the highly modified mammalian skin phenotypes.

A Multiplex Molecular Cell-Based Sensor to Detect Ligands of PPARs: An Optimized Tool for Drug Discovery in Cyanobacteria.

Cyanobacteria produce a wealth of secondary metabolites. Since these organisms attach fatty acids into molecules in unprecedented ways, cyanobacteria can serve as a novel source for bioactive compounds acting as ligands for Peroxisome Proliferator-Activated Receptors (PPAR). PPARs (PPARα, PPARß/δ and PPARγ) are ligand-activated nuclear receptors, involved in the regulation of various metabolic and cellular processes, thus serving as potential drug targets for a variety of pathologies. Yet, given that PPARs' agonists can have pan-, dual- or isoform-specific action, some controversy has been raised over currently approved drugs and their side effects, highlighting the need for novel molecules. Here, we expand and validate a cell-based PPAR transactivation activity biosensor, and test it in a screening campaign to guide drug discovery. Biosensor upgrades included the use of different reporter genes to increase signal intensity and stability, a different promoter to modulate reporter gene expression, and multiplexing to improve efficiency. Sensor's limit of detection (LOD) ranged from 0.36-0.89 nM in uniplex and 0.89-1.35 nM in multiplex mode. In triplex mode, the sensor's feature screening, a total of 848 fractions of 96 cyanobacteria extracts were screened. Hits were confirmed in multiplex mode and in uniplex mode, yielding one strain detected to have action on PPARα and three strains to have dual action on PPARα and -ß.

Using zebrafish embryo bioassays to identify chemicals modulating the regulation of the epigenome: a case study with simvastatin.

Contaminants of emerging concern have been increasingly associated with the modulation of the epigenome, leading to potentially inherited and persistent impacts on apical endpoints. Here, we address the performance of the OECD Test No. 236 FET (fish embryo acute toxicity) in the identification of chemicals able to modulate the epigenome. Using zebrafish (Danio rerio) embryos, acute and chronic exposures were performed with the pharmaceutical, simvastatin (SIM), a widely prescribed hypocholesterolemic drug reported to induce inter and transgenerational effects. In the present study, the epigenetic effects of environmentally relevant concentrations of SIM (from 8 ng/L to 2000 ng/L) were addressed following (1) an acute embryo assay based on OECD Test No. 236 FET, (2) a chronic partial life-cycle exposure using adult zebrafish (90 days), and (3) F1 embryos obtained from parental exposed animals. Simvastatin induced significant effects in gene expression of key epigenetic biomarkers (DNA methylation and histone acetylation/deacetylation) in the gonads of exposed adult zebrafish and in 80 hpf zebrafish embryos (acute and chronic parental intergenerational exposure), albeit with distinct effect profiles between biological samples. In the chronic exposure, SIM impacted particularly DNA methyltransferase genes in males and female gonads, whereas in F1 embryos SIM affected mostly genes associated with histone acetylation/deacetylation. In the embryo acute direct exposure, SIM modulated the expression of both genes involved in DNA methylation and histone deacetylase. These findings further support the use of epigenetic biomarkers in zebrafish embryos in a high throughput approach to identify and prioritize epigenome-modulating chemicals.

Antifouling Marine Coatings with a Potentially Safer and Sustainable Synthetic Polyphenolic Derivative.

The development of harmless substances to replace biocide-based coatings used to prevent or manage marine biofouling and its unwanted consequences is urgent. The formation of biofilms on submerged marine surfaces is one of the first steps in the marine biofouling process, which facilitates the further settlement of macrofoulers. Anti-biofilm properties of a synthetic polyphenolic compound, with previously described anti-settlement activity against macrofoulers, were explored in this work. In solution this new compound was able to prevent biofilm formation and reduce a pre-formed biofilm produced by the marine bacterium, Pseudoalteromonas tunicata. Then, this compound was applied to a marine coating and the formation of P. tunicata biofilms was assessed under hydrodynamic conditions to mimic the marine environment. For this purpose, polyurethane (PU)-based coating formulations containing 1 and 2 wt.% of the compound were prepared based on a prior developed methodology. The most effective formulation in reducing the biofilm cell number, biovolume, and thickness was the PU-based coating containing an aziridine-based crosslinker and 2 wt.% of the compound. To assess the marine ecotoxicity impact of this compound, its potential to disrupt endocrine processes was evaluated through the modulation of two nuclear receptors (NRs), peroxisome proliferator-activated receptor γ (PPARγ), and pregnane X receptor (PXR). Transcriptional activation of the selected NRs upon exposure to the polyphenolic compound (10 µM) was not observed, thus highlighting the eco-friendliness towards the addressed NRs of this new dual-acting anti-macro- and anti-microfouling agent towards the addressed NRs.

The Preservation of PPARγ Genome Duplicates in Some Teleost Lineages: Insights into Lipid Metabolism and Xenobiotic Exploitation.

Three peroxisome proliferator-activated receptor paralogues (PPARα, -ß and -γ) are currently recognized in vertebrate genomes. PPARγ is known to modulate nutrition, adipogenesis and immunity in vertebrates. Natural ligands of PPARγ have been proposed; however, the receptor also binds synthetic ligands such as endocrine disruptors. Two paralogues of PPARα and PPARß have been documented in teleost species, a consequence of the 3R WGD. Recently, two PPARγ paralogue genes were also identified in Astyanax mexicanus. We aimed to determine whether the presence of two PPARγ paralogues is prevalent in other teleost genomes, through genomic and phylogenetic analysis. Our results showed that besides Characiformes, two PPARγ paralogous genes were also identified in other teleost taxa, coinciding with the teleost-specific, whole-genome duplication and with the retention of both genes prior to the separation of the Clupeocephala. To functionally characterize these genes, we used the European sardine (Sardina pilchardus) as a model. PPARγA and PPARγB display a different tissue distribution, despite the similarity of their functional profiles: they are unresponsive to tested fatty acids and other human PPARγ ligands yet yield a transcriptional response in the presence of tributyltin (TBT). This observation puts forward the relevance of comparative analysis to decipher alternative binding architectures and broadens the disruptive potential of man-made chemicals for aquatic species.

A genome assembly of the Atlantic chub mackerel (Scomber colias): a valuable teleost fishing resource.

The Atlantic chub mackerel, Scomber colias (Gmelin, 1789), is a medium-sized pelagic fish with substantial importance in the fisheries of the Atlantic Ocean and the Mediterranean Sea. Over the past decade, this species has gained special relevance, being one of the main targets of pelagic fisheries in the NE Atlantic. Here, we sequenced and annotated the first high-quality draft genome assembly of S. colias, produced with PacBio HiFi long reads and Illumina paired-end short reads. The estimated genome size is 814 Mbp, distributed into 2,028 scaffolds and 2,093 contigs with an N50 length of 4.19 and 3.34 Mbp, respectively. We annotated 27,675 protein-coding genes and the BUSCO analyses indicated high completeness, with 97.3% of the single-copy orthologs in the Actinopterygii library profile. The present genome assembly represents a valuable resource to address the biology and management of this relevant fishery. Finally, this genome assembly ranks fourth in high-quality genome assemblies within the order Scombriformes and first in the genus Scomber.

Functional or Vestigial? The Genomics of the Pineal Gland in Xenarthra.

Vestigial organs are historical echoes of past phenotypes. Determining whether a specific organ constitutes a functional or vestigial structure can be a challenging task, given that distinct levels of atrophy may arise between and within lineages. The mammalian pineal gland, an endocrine organ involved in melatonin biorhythmicity, represents a classic example, often yielding contradicting anatomical observations. In Xenarthra (sloths, anteaters, and armadillos), a peculiar mammalian order, the presence of a distinct pineal organ was clearly observed in some species (i.e., Linnaeus's two-toed sloth), but undetected in other closely related species (i.e., brown-throated sloth). In the nine-banded armadillo, contradicting evidence supports either functional or vestigial scenarios. Thus, to untangle the physiological status of the pineal gland in Xenarthra, we used a genomic approach to investigate the evolution of the gene hub responsible for melatonin synthesis and signaling. We show that both synthesis and signaling compartments are eroded and were probably lost independently among Xenarthra orders. Additionally, by expanding our analysis to 157 mammal genomes, we offer a comprehensive view showing that species with very distinctive habitats and lifestyles have convergently evolved a similar phenotype: Cetacea, Pholidota, Dermoptera, Sirenia, and Xenarthra. Our findings suggest that the recurrent inactivation of melatonin genes correlates with pineal atrophy and endorses the use of genomic analyses to ascertain the physiological status of suspected vestigial structures.

An ancestral nuclear receptor couple, PPAR-RXR, is exploited by organotins.

Environmental chemicals have been reported to greatly disturb the endocrine and metabolic systems of multiple animal species. A recent example involves the exploitation of the nuclear receptor (NR) heterodimeric pair composed by PPAR/RXR (peroxisome proliferator-activated receptor/retinoid X receptor), which shows lipid perturbation in mammalian species. While gene orthologues of both of these receptors have been described outside vertebrates, no functional characterization of PPAR has been carried in protostome lineages. We provide the first functional analysis of PPAR in Patella sp. (Mollusca), using model obesogens such as tributyltin (TBT), triphenyltin (TPT), and proposed natural ligands (fatty acid molecules). To gain further insights, we used site-directed mutagenesis to PPAR and replaced the tyrosine 277 by a cysteine (the human homologous amino acid and TBT anchor residue) and an alanine. Additionally, we explored the alterations in the fatty acid profiles after an exposure to the model obesogen TBT, in vivo. Our results show that TBT and TPT behave as an antagonist of Patella sp. PPAR/RXR and that the tyrosine 277 is important, but not essential in the response to TBT. Overall, these results suggest a relation between the response of the mollusc PPAR-RXR to TBT and the lipid profile alterations reported at environmentally relevant concentrations. Our findings highlight the importance of comparative analysis between protostome and deuterostome lineages to decipher the differential impact of environmental chemicals.

A drastic shift in the energetic landscape of toothed whale sperm cells.

Mammalian spermatozoa are a notable example of metabolic compartmentalization.1 Energy in the form of ATP production, vital for motility, capacitation, and fertilization, is subcellularly separated in sperm cells. While glycolysis provides a local, rapid, and low-yielding input of ATP along the flagellum fibrous sheath, oxidative phosphorylation (OXPHOS), far more efficient over a longer time frame, is concentrated in the midpiece mitochondria.2 The relative weight of glycolysis and OXPHOS pathways in sperm function is variable among species and sensitive to oxygen and substrate availability.3-5 Besides partitioning energy production, sperm cell energetics display an additional singularity: the occurrence of sperm-specific gene duplicates and alternative spliced variants, with conserved function but structurally bound to the flagellar fibrous sheath.6,7 The wider selective forces driving the compartmentalization and adaptability of this energy system in mammalian species remain largely unknown, much like the impact of ecosystem resource availability (e.g., carbohydrates, fatty acids, and proteins) and dietary adaptations in reproductive physiology traits.8 Here, we investigated the Cetacea, an iconic group of fully aquatic and carnivorous marine mammals, evolutionarily related to extant terrestrial herbivores.9 In this lineage, episodes of profound trait remodeling have been accompanied by clear genomic signatures.10-14 We show that toothed whales exhibit impaired sperm glycolysis, due to gene and exon erosion, and demonstrate that dolphin spermatozoa motility depends on endogenous fatty acid ß-oxidation, but not carbohydrates. Such unique energetic rewiring substantiates the observation of large mitochondria in toothed whale spermatozoa and emphasizes the radical physiological reorganization imposed by the transition to a carbohydrate-depleted marine environment.

Epigenetic biomarkers as tools for chemical hazard assessment: Gene expression profiling using the model Danio rerio.

Recent reports raise the concern that exposure to several environmental chemicals may induce persistent changes that go beyond the exposed organisms, being transferred to subsequent generations even in the absence of the original chemical insult. These changes in subsequent non-exposed generations have been related to epigenetic changes. Although highly relevant for hazard and risk assessment, biomarkers of epigenetic modifications that can be associated with adversity, are still not integrated into hazard assessment frameworks. Here, in order to validate new biomarkers of epigenetic modifications in a popular animal model, zebrafish embryos were exposed to different concentrations of Bisphenol A (0.01, 0.1, 1 and 10 mg/L) and Valproic Acid (0.8, 4, 20 and 100 mg/L), two chemicals reported to alter the modulation of the epigenome. Morphological abnormalities and epigenetic changes were assessed at 80 hours-post fertilization, including DNA global methylation and gene expression of both DNA and histone epigenetic modifications. Gene expression changes were detected at concentrations below those inducing morphological abnormalities. These results further support the importance of combining epigenetic biomarkers with apical endpoints to improve guidelines for chemical testing and hazard assessment, and favour the integration of new biomarkers of epigenetic modifications into the standardized OECD test guideline 236 with zebrafish embryos.

From Extrapolation to Precision Chemical Hazard Assessment: The Ecdysone Receptor Case Study.

Hazard assessment strategies are often supported by extrapolation of damage probabilities, regarding chemical action and species susceptibilities. Yet, growing evidence suggests that an adequate sampling of physiological responses across a representative taxonomic scope is of paramount importance. This is particularly relevant for Nuclear Receptors (NR), a family of transcription factors, often triggered by ligands and thus, commonly exploited by environmental chemicals. Within NRs, the ligand-induced Ecdysone Receptor (EcR) provides a remarkable example. Long regarded as arthropod specific, this receptor has been extensively targeted by pesticides, seemingly innocuous to non-target organisms. Yet, current evidence clearly suggests a wider presence of EcR orthologues across metazoan lineages, with unknown physiological consequences. Here, we address the state-of-the-art regarding the phylogenetic distribution and functional characterization of metazoan EcRs and provide a critical analysis of the potential disruption of such EcRs by environmental chemical exposure. Using EcR as a case study, hazard assessment strategies are also discussed in view of the development of a novel "precision hazard assessment paradigm.

Convergent Cortistatin losses parallel modifications in circadian rhythmicity and energy homeostasis in Cetacea and other mammalian lineages.

The ancestors of Cetacea underwent profound morpho-physiological alterations. By displaying an exclusive aquatic existence, cetaceans evolved unique patterns of locomotor activity, vigilant behaviour, thermoregulation and circadian rhythmicity. Deciphering the molecular landscape governing many of these adaptations is key to understand the evolution of phenotypes. Here, we investigate Cortistatin (CORT), a neuropeptide displaying an important role in mammalian biorhythm regulation. This neuropeptide is a known neuroendocrine factor, stimulating slow-wave sleep, but also involved in the regulation of energy metabolism and hypomotility inducement. We assessed the functional status of CORT in 359 mammalian genomes (25 orders), including 30 species of Cetacea. Our findings indicate that cetaceans and other mammals with atypical biorhythms, thermal constraints and/or energy metabolism, have accumulated deleterious mutations in CORT. In light of the pleiotropic action of this neuropeptide, we suggest that this inactivation contributed to a plethora of phenotypic adjustments to accommodate adaptive solutions to specific ecological niches.

Transcriptomic data on the transgenerational exposure of the keystone amphipod Gammarus locusta to simvastatin.

The use of transcriptomics data brings new insights and works as a powerful tool to explore the molecular mode of action (MoA) of transgenerational inheritance effects of contaminants of emerging concern. Therefore, in this dataset, we present the transcriptomic data of the transgenerational effects of environmentally relevant simvastatin levels, one of the most prescribed human pharmaceuticals, in the keystone amphipod species Gammarus locusta. In summary, G. locusta juveniles were maintained under simvastatin exposure up to adulthood (exposed group - F0E) and the offspring of F0E were transferred to control water for the three subsequent generations (transgenerational group - F1T, F2T and F3T). To gain insights into the biological functions and canonical pathways transgenerationally disrupted by simvastatin, a G. locusta de novo transcriptome assembly was produced and the transcriptomic profiles of three individual G. locusta females, per group, over the four generations (F0 to F3) - solvent control groups (F0.C, F1.C, F2.C and F3.C), F0 320 ng/L simvastatin exposed group (F0.320E) and F1 to F3 320 transgenerational group (F1.320T; F2.320T and F3.320T) - were analyzed. Briefly, Illumina HiSeq™ 2500 platform was used to perform RNA sequencing, and due to the unavailability of G. locusta genome, the RNA-seq datasets were assembled de novo using Trinity and annotated with Trinotate software. After assembly and post-processing steps, 106093 transcripts with N50 of 2371 bp and mean sequence length of 1343.98 bp was produced. BUSCO analyses showed a transcriptome with gene completeness of 97.5 % Arthropoda library profile. The Bowtie2, RSEM and edgeR tools were used for the differential gene expression (DEGs) analyses that allowed the identification of a high quantity of genes differentially expressed in all generations. Finally, to identify the main metabolic pathways affected by the transgenerational effects of SIM across all generations, the DGEs genes were blasted onto KEGG pathways database using the KAAS webserver. The data furnished in this article allows a better molecular understanding of the transgenerational effects produced by simvastatin in the keystone amphipod G. locusta and has major implications for hazard and risk assessment of pharmaceuticals and other emerging contaminants. This article is related to the research article entitled "Transgenerational inheritance of chemical-induced signature: a case study with simvastatin [1].

Cartilaginous fishes offer unique insights into the evolution of the nuclear receptor gene repertoire in gnathostomes.

Nuclear receptors (NRs) are key transcription factors that originated in the common ancestor of metazoans. The vast majority of NRs are triggered by binding to either endogenous (e.g. retinoic acid) or exogenous (e.g. xenobiotics) ligands, and their evolution and expansion is tightly linked to the function of endocrine systems. Importantly, they represent classic targets of physiological exploitation by endocrine disrupting chemicals. The NR gene repertoire in different lineages has been shaped by gene loss, duplication and mutation, denoting a dynamic evolutionary route. As the earliest diverging class of gnathostomes (jawed vertebrates), cartilaginous fishes offer an exceptional opportunity to address the early diversification of NR gene families and the evolution of the endocrine system in jawed vertebrates. Here we provide an exhaustive analysis into the NR gene composition in five elasmobranch (sharks and rays) and two holocephalan (chimaeras) species. For this purpose, we generated also a low coverage draft genome assembly of the chimaera small-eyed rabbitfish, Hydrolagus affinis. We show that cartilaginous fish retain an archetypal NR gene repertoire, similar to that of mammals and coincident with the two rounds of whole genome duplication that occurred in the gnathostome ancestor. Furthermore, novel gene members of the non-canonical NR0B receptors were found in the genomes of this lineage. Our findings provide an essential view into the early diversification of NRs in gnathostomes, paving the way for functional studies.

PseudoChecker: an integrated online platform for gene inactivation inference.

The rapid expansion of high-quality genome assemblies, exemplified by ongoing initiatives such as the Genome-10K and i5k, demands novel automated methods to approach comparative genomics. Of these, the study of inactivating mutations in the coding region of genes, or pseudogenization, as a source of evolutionary novelty is mostly overlooked. Thus, to address such evolutionary/genomic events, a systematic, accurate and computationally automated approach is required. Here, we present PseudoChecker, the first integrated online platform for gene inactivation inference. Unlike the few existing methods, our comparative genomics-based approach displays full automation, a built-in graphical user interface and a novel index, PseudoIndex, for an empirical evaluation of the gene coding status. As a multi-platform online service, PseudoChecker simplifies access and usability, allowing a fast identification of disruptive mutations. An analysis of 30 genes previously reported to be eroded in mammals, and 30 viable genes from the same lineages, demonstrated that PseudoChecker was able to correctly infer 97% of loss events and 95% of functional genes, confirming its reliability. PseudoChecker is freely available, without login required, at http://pseudochecker.ciimar.up.pt.

Of Retinoids and Organotins: The Evolution of the Retinoid X Receptor in Metazoa.

Nuclear receptors (NRs) are transcription factors accomplishing a multiplicity of functions, essential for organismal homeostasis. Among their numerous members, the retinoid X receptor (RXR) is a central player of the endocrine system, with a singular ability to operate as a homodimer or a heterodimer with other NRs. Additionally, RXR has been found to be a critical actor in various processes of endocrine disruption resulting from the exposure to a known class of xenobiotics termed organotins (e.g., tributyltin (TBT)), including imposex in gastropod molluscs and lipid perturbation across different metazoan lineages. Thus, given its prominent physiological and endocrine role, RXR is present in the genomes of most extant metazoan species examined to date. Here, we expand on the phylogenetic distribution of RXR across the metazoan tree of life by exploring multiple next-generation sequencing projects of protostome lineages. By addressing amino acid residue conservation in combination with cell-based functional assays, we show that RXR induction by 9-cis retinoic acid (9cisRA) and TBT is conserved in more phyla than previously described. Yet, our results highlight distinct activation efficacies and alternative modes of RXR exploitation by the organotin TBT, emphasizing the need for broader species sampling to clarify the mechanistic activation of RXR.