Duplication and Losses of Opsin Genes in Lophotrochozoan Evolution.

Opsins are G-coupled receptors playing a key role in metazoan visual processes. While many studies enriched our understanding of opsin diversity in several animal clades, the opsin evolution in Lophotrochozoa, one of the major metazoan groups, remains poorly understood. Using recently developed phylogenetic approaches, we investigated the opsin evolution in 74 lophotrochozoan genomes. We found that the common ancestor of Lophotrochozoa possessed at least seven opsin paralog groups that underwent divergent evolutionary history in the different phyla. Furthermore, we showed for the first time opsin-related molecules in Bilateria that we named pseudopsins, which may prove critical in uncovering opsin evolution.

Rethinking Opsins.

Opsins, the protein moieties of animal visual photo-pigments, have emerged as moonlighting proteins with diverse, light-dependent and -independent physiological functions. This raises the need to revise some basic assumptions concerning opsin expression, structure, classification, and evolution.

Ciliary photoreceptors in sea urchin larvae indicate pan-deuterostome cell type conservation.

BACKGROUND: The evolutionary history of cell types provides insights into how morphological and functional complexity arose during animal evolution. Photoreceptor cell types are particularly broadly distributed throughout Bilateria; however, their evolutionary relationship is so far unresolved. Previous studies indicate that ciliary photoreceptors are homologous at least within chordates, and here, we present evidence that a related form of this cell type is also present in echinoderm larvae. RESULTS: Larvae of the purple sea urchin Strongylocentrotus purpuratus have photoreceptors that are positioned bilaterally in the oral/anterior apical neurogenic ectoderm. Here, we show that these photoreceptors express the transcription factor Rx, which is commonly expressed in ciliary photoreceptors, together with an atypical opsin of the GO family, opsin3.2, which localizes in particular to the cilia on the cell surface of photoreceptors. We show that these ciliary photoreceptors express the neuronal marker synaptotagmin and are located in proximity to pigment cells. Furthermore, we systematically identified additional transcription factors expressed in these larval photoreceptors and found that a majority are orthologous to transcription factors expressed in vertebrate ciliary photoreceptors, including Otx, Six3, Tbx2/3, and Rx. Based on the developmental expression of rx, these photoreceptors derive from the anterior apical neurogenic ectoderm. However, genes typically involved in eye development in bilateria, including pax6, six1/2, eya, and dac, are not expressed in sea urchin larval photoreceptors but are instead co-expressed in the hydropore canal. CONCLUSIONS: Based on transcription factor expression, location, and developmental origin, we conclude that the sea urchin larval photoreceptors constitute a cell type that is likely homologous to the ciliary photoreceptors present in chordates.

Genomic data do not support comb jellies as the sister group to all other animals.

Understanding how complex traits, such as epithelia, nervous systems, muscles, or guts, originated depends on a well-supported hypothesis about the phylogenetic relationships among major animal lineages. Traditionally, sponges (Porifera) have been interpreted as the sister group to the remaining animals, a hypothesis consistent with the conventional view that the last common animal ancestor was relatively simple and more complex body plans arose later in evolution. However, this premise has recently been challenged by analyses of the genomes of comb jellies (Ctenophora), which, instead, found ctenophores as the sister group to the remaining animals (the "Ctenophora-sister" hypothesis). Because ctenophores are morphologically complex predators with true epithelia, nervous systems, muscles, and guts, this scenario implies these traits were either present in the last common ancestor of all animals and were lost secondarily in sponges and placozoans (Trichoplax) or, alternatively, evolved convergently in comb jellies. Here, we analyze representative datasets from recent studies supporting Ctenophora-sister, including genome-scale alignments of concatenated protein sequences, as well as a genomic gene content dataset. We found no support for Ctenophora-sister and conclude it is an artifact resulting from inadequate methodology, especially the use of simplistic evolutionary models and inappropriate choice of species to root the metazoan tree. Our results reinforce a traditional scenario for the evolution of complexity in animals, and indicate that inferences about the evolution of Metazoa based on the Ctenophora-sister hypothesis are not supported by the currently available data.

Metazoan opsin evolution reveals a simple route to animal vision.

All known visual pigments in Neuralia (Cnidaria, Ctenophora, and Bilateria) are composed of an opsin (a seven-transmembrane G protein-coupled receptor), and a light-sensitive chromophore, generally retinal. Accordingly, opsins play a key role in vision. There is no agreement on the relationships of the neuralian opsin subfamilies, and clarifying their phylogeny is key to elucidating the origin of this protein family and of vision. We used improved methods and data to resolve the opsin phylogeny and explain the evolution of animal vision. We found that the Placozoa have opsins, and that the opsins share a common ancestor with the melatonin receptors. Further to this, we found that all known neuralian opsins can be classified into the same three subfamilies into which the bilaterian opsins are classified: the ciliary (C), rhabdomeric (R), and go-coupled plus retinochrome, retinal G protein-coupled receptor (Go/RGR) opsins. Our results entail a simple scenario of opsin evolution. The first opsin originated from the duplication of the common ancestor of the melatonin and opsin genes in a eumetazoan (Placozoa plus Neuralia) ancestor, and an inference of its amino acid sequence suggests that this protein might not have been light-sensitive. Two more gene duplications in the ancestral neuralian lineage resulted in the origin of the R, C, and Go/RGR opsins. Accordingly, the first animal with at least a C, an R, and a Go/RGR opsin was a neuralian progenitor.

The origin, evolution, and molecular diversity of the chemokine system.

Chemokine signalling performs key functions in cell migration via chemoattraction, such as attracting leukocytes to the site of infection during host defence. The system consists of a ligand, the chemokine, usually secreted outside the cell, and a chemokine receptor on the surface of a target cell that recognises the ligand. Several noncanonical components interact with the system. These include a variety of molecules that usually share some degree of sequence similarity with canonical components and, in some cases, are known to bind to canonical components and/or to modulate cell migration. Whereas canonical components have been described in vertebrate lineages, the distribution of the noncanonical components is less clear. Uncertainty over the relationships between canonical and noncanonical components hampers our understanding of the evolution of the system. We used phylogenetic methods, including gene-tree to species-tree reconciliation, to untangle the relationships between canonical and noncanonical components, identify gene duplication events, and clarify the origin of the system. We found that unrelated ligand groups independently evolved chemokine-like functions. We found noncanonical ligands outside vertebrates, such as TAFA "chemokines" found in urochordates. In contrast, all receptor groups are vertebrate-specific and all-except ACKR1-originated from a common ancestor in early vertebrates. Both ligand and receptor copy numbers expanded through gene duplication events at the base of jawed vertebrates, with subsequent waves of innovation occurring in bony fish and mammals.

Hagfish genome elucidates vertebrate whole-genome duplication events and their evolutionary consequences.

Polyploidy or whole-genome duplication (WGD) is a major event that drastically reshapes genome architecture and is often assumed to be causally associated with organismal innovations and radiations. The 2R hypothesis suggests that two WGD events (1R and 2R) occurred during early vertebrate evolution. However, the timing of the 2R event relative to the divergence of gnathostomes (jawed vertebrates) and cyclostomes (jawless hagfishes and lampreys) is unresolved and whether these WGD events underlie vertebrate phenotypic diversification remains elusive. Here we present the genome of the inshore hagfish, Eptatretus burgeri. Through comparative analysis with lamprey and gnathostome genomes, we reconstruct the early events in cyclostome genome evolution, leveraging insights into the ancestral vertebrate genome. Genome-wide synteny and phylogenetic analyses support a scenario in which 1R occurred in the vertebrate stem-lineage during the early Cambrian, and 2R occurred in the gnathostome stem-lineage, maximally in the late Cambrian-earliest Ordovician, after its divergence from cyclostomes. We find that the genome of stem-cyclostomes experienced an additional independent genome triplication. Functional genomic and morphospace analyses demonstrate that WGD events generally contribute to developmental evolution with similar changes in the regulatory genome of both vertebrate groups. However, appreciable morphological diversification occurred only in the gnathostome but not in the cyclostome lineage, calling into question the general expectation that WGDs lead to leaps of bodyplan complexity.

The monoaminergic system is a bilaterian innovation.

Monoamines like serotonin, dopamine, and adrenaline/noradrenaline (epinephrine/norepinephrine) act as neuromodulators in the nervous system. They play a role in complex behaviours, cognitive functions such as learning and memory formation, as well as fundamental homeostatic processes such as sleep and feeding. However, the evolutionary origin of the genes required for monoaminergic modulation is uncertain. Using a phylogenomic approach, in this study, we show that most of the genes involved in monoamine production, modulation, and reception originated in the bilaterian stem group. This suggests that the monoaminergic system is a bilaterian novelty and that its evolution may have contributed to the Cambrian diversification.

Shallow Whole-Genome Sequencing of Aedes japonicus and Aedes koreicus from Italy and an Updated Picture of Their Evolution Based on Mitogenomics and Barcoding.

Aedes japonicus and Aedes koreicus are two invasive mosquitoes native to East Asia that are quickly establishing in temperate regions of Europe. Both species are vectors of arboviruses, but we currently lack a clear understanding of their evolution. Here, we present new short-read, shallow genome sequencing of A. japonicus and A. koreicus individuals from northern Italy, which we used for downstream phylogenetic and barcode analyses. We explored associated microbial DNA and found high occurrences of Delftia bacteria in both samples, but neither Asaia nor Wolbachia. We then assembled complete mitogenomes and used these data to infer divergence times estimating the split of A. japonicus from A. koreicus in the Oligocene, which was more recent than that previously reported using mitochondrial markers. We recover a younger age for most other nodes within Aedini and other Culicidae. COI barcoding and phylogenetic analyses indicate that A. japonicus yaeyamensis, A. japonicus amamiensis, and the two A. koreicus sampled from Europe should be considered as separate species within a monophyletic species complex. Our studies further clarify the evolution of A. japonicus and A. koreicus, and indicate the need to obtain whole-genome data from putative species in order to disentangle their complex patterns of evolution.

BRAFV600E-mutated serrated colorectal neoplasia drives transcriptional activation of cholesterol metabolism.

BRAF mutations occur early in serrated colorectal cancers, but their long-term influence on tissue homeostasis is poorly characterized. We investigated the impact of short-term (3 days) and long-term (6 months) expression of BrafV600E in the intestinal tissue of an inducible mouse model. We show that BrafV600E perturbs the homeostasis of intestinal epithelial cells, with impaired differentiation of enterocytes emerging after prolonged expression of the oncogene. Moreover, BrafV600E leads to a persistent transcriptional reprogramming with enrichment of numerous gene signatures indicative of proliferation and tumorigenesis, and signatures suggestive of metabolic rewiring. We focused on the top-ranking cholesterol biosynthesis signature and confirmed its increased expression in human serrated lesions. Functionally, the cholesterol lowering drug atorvastatin prevents the establishment of intestinal crypt hyperplasia in BrafV600E-mutant mice. Overall, our work unveils the long-term impact of BrafV600E expression in intestinal tissue and suggests that colorectal cancers with mutations in BRAF might be prevented by statins.

Resolving phylogenetic signal from noise when divergence is rapid: a new look at the old problem of echinoderm class relationships.

Resolving evolutionary relationships in groups that underwent fast radiation in deep time is a problem for molecular phylogeny, as the scant phylogenetic signal that characterises short internal branches is generally swamped by more recent substitutions. We implement an approach, that maps how the support for rival phylogenies changes when analysing subsets of sites with either faster and more heterogeneous rates or slower and more homogeneous rates, to address a long-standing problem in deuterostome phylogeny - the interrelationships of the eleutherozoan echinoderm classes. We show that miRNA genes are phylogenetically uninformative as to the relationships of asteroids, echinoids and ophiuroids, consistent with a rapid radiation of these groups as suggested by their fossil record. Using three nuclear rRNAs and seven nuclear housekeeping genes, we map the support for the three possible phylogenetic arrangements of asteroids, ophiuroids and echinoids when moving between subsets of the data with very similar or very different rates of evolution. Only one of the three possible topologies (asteroids (ophiuroids+echinoids)) strengthens when the most rate-homogeneous subset of data are analysed. The other two possible pairings become stronger in a less reliable data subset, which includes the fastest and thus homoplasy-rich data in our alignment. Thus, while superficial analysis of our concatenated alignment identifies asteroids and ophiuroids as sister taxa, more thorough analyses suggest that ophiuroids may be more closely related to echinoids. Divergence of these echinoderm groups, using a relaxed molecular clock, is estimated to have occurred within ≈ 5 million years. Our results illustrate that the analytic approach of phylogenetic signal dissection can be a powerful tool to investigate rapid radiations in deep geologic time.

The temporal dynamics of the sea urchin regulome.

In this work, we used Nanostring N-counter technology, to evaluate the mRNA expression level of more than 330 regulatory genes over 34 time points covering the first three days of development of the sea urchin larvae. The hierarchical clustering of the mRNAs expression levels has identified groups corresponding to the major developmental landmarks (e.g. maternal to zygotic transition and gastrulation). Furthermore, comparison with previous experiments indicates high reproducibility of mRNA level temporal dynamics across batches. Finally, we generated an online tool to visualise gene expression during sea urchin larval development. The site can be accessed at and https://www621.lamp.le.ac.uk/nanostring_app/nanostring/.

Homologous gene regulatory networks control development of apical organs and brains in Bilateria.

Apical organs are relatively simple larval nervous systems. The extent to which apical organs are evolutionarily related to the more complex nervous systems of other animals remains unclear. To identify common developmental mechanisms, we analyzed the gene regulatory network (GRN) controlling the development of the apical organ in sea urchins. We characterized the developmental expression of 30 transcription factors and identified key regulatory functions for FoxQ2, Hbn, Delta/Notch signaling, and SoxC in the patterning of the apical organ and the specification of neurons. Almost the entire set of apical transcription factors is expressed in the nervous system of worms, flies, zebrafish, frogs, and mice. Furthermore, a regulatory module controlling the axial patterning of the vertebrate brain is expressed in the ectoderm of sea urchin embryos. We conclude that GRNs controlling the formation of bilaterian nervous systems share a common origin and that the apical GRN likely resembles an ancestral regulatory program.

Phylogenomics: Is less more when using large-scale datasets?

Phylogenetic studies have traditionally placed the simple Xenoacoelomorph worms as the sister group of all other animals with bilateral body symmetry. A new study shows that misidentification of orthologous genes might have been the source of at least some support for this placement.

Resolving tricky nodes in the tree of life through amino acid recoding.

Genomic data allowed a detailed resolution of the Tree of Life, but "tricky nodes" such as the root of the animals remain unresolved. Genome-scale datasets are heterogeneous as genes and species are exposed to different pressures, and this can negatively impacts phylogenetic accuracy. We use simulated genomic-scale datasets and show that recoding amino acid data improves accuracy when the model does not account for the compositional heterogeneity of the amino acid alignment. We apply our findings to three datasets addressing the root of the animal tree, where the debate centers on whether sponges (Porifera) or comb jellies (Ctenophora) represent the sister of all other animals. We show that results from empirical data follow predictions from simulations and suggest that, at the least in phylogenies inferred from amino acid sequences, a placement of the ctenophores as sister to all the other animals is best explained as a tree reconstruction artifact.

Phylogenomics of Opsin Genes in Diptera Reveals Lineage-Specific Events and Contrasting Evolutionary Dynamics in Anopheles and Drosophila.

Diptera is one of the biggest insect orders and displays a large diversity of visual adaptations. Similarly to other animals, the dipteran visual process is mediated by opsin genes. Although the diversity and function of these genes are well studied in key model species, a comprehensive comparative genomic study across the dipteran phylogeny is missing. Here we mined the genomes of 61 dipteran species, reconstructed the evolutionary affinities of 528 opsin genes, and determined the selective pressure acting in different species. We found that opsins underwent several lineage-specific events, including an independent expansion of Long Wave Sensitive opsins in flies and mosquitoes, and numerous family-specific duplications and losses. Both the Drosophila and the Anopheles complement are derived in comparison with the ancestral dipteran state. Molecular evolutionary studies suggest that gene turnover rate, overall mutation rate, and site-specific selective pressure are higher in Anopheles than in Drosophila. Overall, our findings indicate an extremely variable pattern of opsin evolution in dipterans, showcasing how two similarly aged radiations, Anopheles and Drosophila, are characterized by contrasting dynamics in the evolution of this gene family. These results provide a foundation for future studies on the dipteran visual system.

Profiling cellular diversity in sponges informs animal cell type and nervous system evolution.

The evolutionary origin of metazoan cell types such as neurons and muscles is not known. Using whole-body single-cell RNA sequencing in a sponge, an animal without nervous system and musculature, we identified 18 distinct cell types. These include nitric oxide­sensitive contractile pinacocytes, amoeboid phagocytes, and secretory neuroid cells that reside in close contact with digestive choanocytes that express scaffolding and receptor proteins. Visualizing neuroid cells by correlative x-ray and electron microscopy revealed secretory vesicles and cellular projections enwrapping choanocyte microvilli and cilia. Our data show a communication system that is organized around sponge digestive chambers, using conserved modules that became incorporated into the pre- and postsynapse in the nervous systems of other animals.

Sensory Neuroscience: A Taste for Light and the Origin of Animal Vision.

Opsins are genes underpinning vision in animals. A new study shows that they are also involved in taste perception in fruit flies, significantly expanding their scope of action. This has important implications for our understanding of the evolution of vision.

A Novel Approach to Investigate the Effect of Tree Reconstruction Artifacts in Single-Gene Analysis Clarifies Opsin Evolution in Nonbilaterian Metazoans.

Our ability to correctly reconstruct a phylogenetic tree is strongly affected by both systematic errors and the amount of phylogenetic signal in the data. Current approaches to tackle tree reconstruction artifacts, such as the use of parameter-rich models, do not translate readily to single-gene alignments. This, coupled with the limited amount of phylogenetic information contained in single-gene alignments, makes gene trees particularly difficult to reconstruct. Opsin phylogeny illustrates this problem clearly. Opsins are G-protein coupled receptors utilized in photoreceptive processes across Metazoa and their protein sequences are roughly 300 amino acids long. A number of incongruent opsin phylogenies have been published and opsin evolution remains poorly understood. Here, we present a novel approach, the canary sequence approach, to investigate and potentially circumvent errors in single-gene phylogenies. First, we demonstrate our approach using two well-understood cases of long-branch attraction in single-gene data sets, and simulations. After that, we apply our approach to a large collection of well-characterized opsins to clarify the relationships of the three main opsin subfamilies.