Detection of evolutionary conserved and accelerated genomic regions related to adaptation to thermal niches in Anolis lizards.

Understanding the genetic basis for adapting to thermal environments is important due to serious effects of global warming on ectothermic species. Various genes associated with thermal adaptation in lizards have been identified mainly focusing on changes in gene expression or the detection of positively selected genes using coding regions. Only a few comprehensive genome-wide analyses have included noncoding regions. This study aimed to identify evolutionarily conserved and accelerated genomic regions using whole genomes of eight Anolis lizard species that have repeatedly adapted to similar thermal environments in multiple lineages. Evolutionarily conserved genomic regions were extracted as regions with overall sequence conservation (regions with fewer base substitutions) across all lineages compared with the neutral model. Genomic regions that underwent accelerated evolution in the lineage of interest were identified as those with more base substitutions in the target branch than in the entire background branch. Conserved elements across all branches were relatively abundant in "intergenic" genomic regions among noncoding regions. Accelerated regions (ARs) of each lineage contained a significantly greater proportion of noncoding RNA genes than the entire multiple alignment. Common genes containing ARs within 5 kb of their vicinity in lineages with similar thermal habitats were identified. Many genes associated with circadian rhythms and behavior were found in hot-open and cool-shaded habitat lineages. These genes might play a role in contributing to thermal adaptation and assist future studies examining the function of genes involved in thermal adaptation via genome editing.

Transcriptomic signatures associated with underlying rapid changes in the early phase brain of bi-directional sex change in Trimma okinawae.

Teleost fish exhibit remarkable sexual plasticity and divergent developmental systems, including sequential hermaphroditism. One of the more fascinating models of sexual plasticity is socially controlled sex change, which is often observed in coral reef fish. The Okinawa rubble goby, Trimma okinawae, is a bi-directional sex-changing fish. It can rapidly change sex in either direction based on social circumstances. Although behavioural and neuroendocrine sex change occurs immediately and is believed to trigger gonadal changes, the underlying mechanisms remain poorly understood. In this study, we conducted a de novo transcriptome analysis of the T. okinawae brain and identified genes that are differentially expressed between the sexes and genes that were immediately controlled by social stimulation implicating sex change. Several genes showed concordant expression shifts regardless of the sex change direction and were associated with histone modification in nerve cells. These genes are known to function in the neuroendocrine control of reproduction in nerve cells. Overall, we identified genes associated with the initiation of sex change, which provides insight into the regulation of sex change and sexual plasticity.

Genetic differentiation and local adaptation of the Japanese honeybee, Apis cerana japonica.

We examine the population genetic structure and divergence among the regional populations of the Japanese honeybee, Apis cerana japonica, by re-sequencing the genomes of 105 individuals from the three main Japanese islands with diverse climates. The genetic structure results indicated that these individuals are distinct from the mainland Chinese A. cerana samples. Furthermore, population structure analyses have identified three genetically distinct geographic regions in Japan: Northern (Tohoku-Kanto-Chubu districts), Central (Chugoku district), and Southern (Kyushu district). In some districts, "possible non-native" individuals, likely introduced from other regions in recent years, were discovered. Then, genome-wide scans were conducted to detect candidate genes for adaptation by two different approaches. We performed a population branch statistics (PBS) analysis to identify candidate genes for population-specific divergence. A latent factor mixed model (LFMM) was used to identify genes associated with climatic variables along a geographic gradient. The PBSmax analysis identified 25 candidate genes for population-specific divergence whereas the LFMM analysis identified 73 candidate genes for adaptation to climatic variables along a geographic gradient. However, no common genes were identified by both methods.

Environmental pH signals the release of monosaccharides from cell wall in coral symbiotic alga.

Reef-building corals thrive in oligotrophic environments due to their possession of endosymbiotic algae. Confined to the low pH interior of the symbiosome within the cell, the algal symbiont provides the coral host with photosynthetically fixed carbon. However, it remains unknown how carbon is released from the algal symbiont for uptake by the host. Here we show, using cultured symbiotic dinoflagellate, Breviolum sp., that decreases in pH directly accelerates the release of monosaccharides, that is, glucose and galactose, into the ambient environment. Under low pH conditions, the cell surface structures were deformed and genes related to cellulase were significantly upregulated in Breviolum. Importantly, the release of monosaccharides was suppressed by the cellulase inhibitor, glucopyranoside, linking the release of carbon to degradation of the agal cell wall. Our results suggest that the low pH signals the cellulase-mediated release of monosaccharides from the algal cell wall as an environmental response in coral reef ecosystems.

Coral reefs are known as 'treasure troves of biodiversity' because of the enormous variety of different fish, crustaceans and other marine life they support. Colonies of marine animals, known as corals, which are anchored to rocks on the sea bed, form the main structures of a coral reef. Many corals rely on partnerships with microscopic algae known as dinoflagellates for most of their energy needs. The dinoflagellates use sunlight to make sugars and other carbohydrates and they give some of these to the coral. In exchange, the coral provides a home for the dinoflagellates inside its body. The algae live inside special compartments within coral cells known as symbiosomes. These compartments have a lower pH (that is, they are more acidic) than the rest of the coral cell. Previous studies have shown that the algae release sugars into the symbiosome but it remains unclear what triggers this release and whether it only occurs when the algae are in a partnership. Ishii et al. studied a type of dinoflagellate known as Breviolum sp. that had been grown in sea water-like liquid in a laboratory. The experiments found that the alga released two sugar molecules known as glucose and galactose into its surroundings even in the absence of a host coral. Increasing the acidity of the liquid caused the alga to release more sugars and resulted in changes to some of the structures on the surface of its cells. The alga also produced an enzyme, called cellulase, to degrade the wall that normally surrounds the cell of an alga. Treating the alga with a drug that inhibits the activity of cellulase also suppressed the release of sugars from the cells. These findings suggest that when dinoflagellates enter acidic environments, like the guts of marine animals or symbiosomes inside coral cells, the decrease in pH can activate the algal cellulase enzyme, which in turn triggers the release of sugars for the coral. This research will provide a new viewpoint to those interested in how partnerships between animals and algae are sustained in marine environments. It also highlights the importance of the alga cell wall in establishing partnerships with corals. Further work will seek to clarify the precise biological mechanisms involved.

On the origin of appetite: GLWamide in jellyfish represents an ancestral satiety neuropeptide.

Food intake is regulated by internal state. This function is mediated by hormones and neuropeptides, which are best characterized in popular model species. However, the evolutionary origins of such feeding-regulating neuropeptides are poorly understood. We used the jellyfish Cladonema to address this question. Our combined transcriptomic, behavioral, and anatomical approaches identified GLWamide as a feeding-suppressing peptide that selectively inhibits tentacle contraction in this jellyfish. In the fruit fly Drosophila, myoinhibitory peptide (MIP) is a related satiety peptide. Surprisingly, we found that GLWamide and MIP were fully interchangeable in these evolutionarily distant species for feeding suppression. Our results suggest that the satiety signaling systems of diverse animals share an ancient origin.

Differentially Expressed Genes Associated with Body Size Changes and Transposable Element Insertions between Caenorhabditis elegans and Its Sister Species, Caenorhabditis inopinata.

Why the recently discovered nematode Caenorhabditis inopinata differs so greatly from its sibling species Caenorhabditis elegans remains unknown. A previous study showed that C. inopinata has more transposable elements (TEs), sequences that replicate and move autonomously throughout the genome, potentially altering the expression of neighboring genes. In this study, we focused on how the body size of this species has evolved and whether TEs could affect the expression of genes related to species-specific traits such as body size. First, we compared gene expression levels between C. inopinata and C. elegans in the L4 larval and young adult stages-when growth rates differ most prominently between these species-to identify candidate genes contributing to their differences. The results showed that the expression levels of collagen genes were consistently higher in C. inopinata than in C. elegans and that some genes related to cell size were differentially expressed between the species. Then, we examined whether genes with TE insertions are differentially expressed between species. Indeed, the genes featuring C. inopinata-specific TE insertions had higher expression levels in C. inopinata than in C. elegans. These upregulated genes included those related to body size, suggesting that these genes could be candidates for artificial TE insertion to examine the role of TEs in the body size evolution of C. inopinata.

Draft genome of six Cuban Anolis lizards and insights into genetic changes during their diversification.

BACKGROUND: Detecting genomic variants and their accumulation processes during species diversification and adaptive radiation is important for understanding the molecular and genetic basis of evolution. Anolis lizards in the West Indies are good models for studying evolutionary mechanisms because of the repeated evolution of their morphology and the ecology. We performed de novo genome assembly of six Cuban Anolis lizards with different ecomorphs and thermal habitats (Anolis isolepis, Anolis allisoni, Anolis porcatus, Anolis allogus, Anolis homolechis, and Anolis sagrei). We carried out a comparative analysis of these genome assemblies to investigate the genetic changes that occurred during their diversification. RESULTS: We reconstructed novel draft genomes with relatively long scaffolds and high gene completeness, with the scaffold N50 ranging from 5.56 to 39.79 Mb and vertebrate Benchmarking Universal Single-Copy Orthologs completeness ranging from 77.5% to 86.9%. Comparing the repeat element compositions and landscapes revealed differences in the accumulation process between Cuban trunk-crown and trunk-ground species and separate expansions of several families of LINE in each Cuban trunk-ground species. Duplicated gene analysis suggested that the proportional differences in duplicated gene numbers among Cuban Anolis lizards may be associated with differences in their habitat ranges. Additionally, Pairwise Sequentially Markovian Coalescent analysis suggested that the effective population sizes of each species may have been affected by Cuba's geohistory. CONCLUSIONS: We provide draft genomes of six Cuban Anolis lizards and detected species and lineage-specific transposon accumulation and gene copy number changes that may be involved in adaptive evolution. The change processes in the past effective population size was also estimated, and the factors involved were inferred. These results provide new insights into the genetic basis of Anolis lizard diversification and are expected to serve as a stepping stone for the further elucidation of their diversification mechanisms.

Humanized substitutions of Vmat1 in mice alter amygdala-dependent behaviors associated with the evolution of anxiety.

The human vesicular monoamine transporter 1 (VMAT1) harbors unique substitutions (Asn136Thr/Ile) that affect monoamine uptake into synaptic vesicles. These substitutions are absent in all known mammals, suggesting their contributions to distinct aspects of human behavior modulated by monoaminergic transmissions, such as emotion and cognition. To directly test the impact of these human-specific mutations, we introduced the humanized residues into mouse Vmat1 via CRISPR/Cas9-mediated genome editing and examined changes at the behavioral, neurophysiological, and molecular levels. Behavioral tests revealed reduced anxiety-related traits of Vmat1 Ile mice, consistent with human studies, and electrophysiological recordings showed altered oscillatory activity in the amygdala under anxiogenic conditions. Transcriptome analyses further identified changes in gene expressions in the amygdala involved in neurodevelopment and emotional regulation, which may corroborate the observed phenotypes. This knock-in mouse model hence provides compelling evidence that the mutations affecting monoaminergic signaling and amygdala circuits have contributed to the evolution of human socio-emotional behaviors.

Genes associated with hot defensive bee ball in the Japanese honeybee, Apis cerana japonica.

BACKGROUND: The Japanese honeybee, Apis cerana japonica, shows a specific defensive behavior, known as a "hot defensive bee ball," used against the giant hornet, Vespa mandarinia. Hundreds of honeybee workers surround a hornet and make a "bee ball" during this behavior. They maintain the ball for around 30 min, and its core temperature can reach 46. Although various studies have been conducted on the characteristics of this behavior, its molecular mechanism has yet to be elucidated. Here, we performed a comprehensive transcriptomic analysis to detect candidate genes related to balling behavior. RESULTS: The expression levels of differentially expressed genes (DEGs) in the brain, flight muscle, and fat body were evaluated during ball formation and incubation at 46 °C. The DEGs detected during ball formation, but not in response to heat, were considered important for ball formation. The expression of genes related to rhodopsin signaling were increased in all tissues during ball formation. DEGs detected in one or two tissues during ball formation were also identified. CONCLUSIONS: Given that rhodopsin is involved in temperature sensing in Drosophila, the rhodopsin-related DEGs in A. cerana japonica may be involved in temperature sensing specifically during ball formation.

Gene expression alterations from reversible to irreversible stages during coral metamorphosis.

For corals, metamorphosis from planktonic larvae to sedentary polyps is an important life event, as it determines the environment in which they live for a lifetime. Although previous studies on the reef-building coral Acropora have clarified a critical time point during metamorphosis when cells are committed to their fates, as defined by an inability to revert back to their previous states as swimming larvae (here referred to as the "point of no return"), the molecular mechanisms of this commitment to a fate remain unclear. To address this issue, we analyzed the transcriptomic changes before and after the point of no return by inducing metamorphosis of Acropora tenuis with Hym-248, a metamorphosis-inducing neuropeptide. Gene Ontology and pathway enrichment analysis of the 5893 differentially expressed genes revealed that G protein-coupled receptors (GPCRs) were enriched, including GABA receptor and Frizzled gene subfamilies, which showed characteristic temporal expression patterns. The GPCRs were then classified by comparison with those of Homo sapiens, Nematostella vectensis and Platynereis dumerilii. Classification of the differentially expressed genes into modules based on expression patterns showed that some modules with large fluctuations after the point of no return were biased toward functions such as protein metabolism and transport. This result suggests that in precommitted larvae, different types of GPCR genes function to ensure a proper environment, whereas in committed larvae, intracellular protein transport and proteolysis may cause a loss of the reversibility of metamorphosis as a result of cell differentiation.

Deep learning increases the availability of organism photographs taken by citizens in citizen science programs.

Citizen science programs using organism photographs have become popular, but there are two problems related to photographs. One problem is the low quality of photographs. It is laborious to identify species in photographs taken outdoors because they are out of focus, partially invisible, or under different lighting conditions. The other is difficulty for non-experts to identify species. Organisms usually have interspecific similarity and intraspecific variation, which hinder species identification by non-experts. Deep learning solves these problems and increases the availability of organism photographs. We trained a deep convolutional neural network, Xception, to identify bee species using various quality of bee photographs that were taken by citizens. These bees belonged to two honey bee species and 10 bumble bee species with interspecific similarity and intraspecific variation. We investigated the accuracy of species identification by biologists and deep learning. The accuracy of species identification by Xception (83.4%) was much higher than that of biologists (53.7%). When we grouped bee photographs by different colors resulting from intraspecific variation in addition to species, the accuracy of species identification by Xception increased to 84.7%. The collaboration with deep learning and experts will increase the reliability of species identification and their use for scientific researches.

Genetic basis of orange spot formation in the guppy (Poecilia reticulata).

BACKGROUND: To understand the evolutionary significance of female mate choice for colorful male ornamentation, the underlying regulatory mechanisms of such ornamentation must be understood for examining how the ornaments are associated with "male qualities" that increase the fitness or sexual attractiveness of offspring. In the guppy (Poecilia reticulata), an established model system for research on sexual selection, females prefer males possessing larger and more highly saturated orange spots as potential mates. Although previous studies have identified some chromosome regions and genes associated with orange spot formation, the regulation and involvement of these genetic elements in orange spot formation have not been elucidated. In this study, the expression patterns of genes specific to orange spots and certain color developmental stages were investigated using RNA-seq to reveal the genetic basis of orange spot formation. RESULTS: Comparing the gene expression levels of male guppy skin with orange spots (orange skin) with those without any color spots (dull skin) from the same individuals identified 1102 differentially expressed genes (DEGs), including 630 upregulated genes and 472 downregulated genes in the orange skin. Additionally, the gene expression levels of the whole trunk skin were compared among the three developmental stages and 2247 genes were identified as DEGs according to color development. These analyses indicated that secondary differentiation of xanthophores may affect orange spot formation. CONCLUSIONS: The results suggested that orange spots might be formed by secondary differentiation, rather than de novo generation, of xanthophores, which is induced by Csf1 and thyroid hormone signaling pathways. Furthermore, we suggested candidate genes associated with the areas and saturation levels of orange spots, which are both believed to be important for female mate choice and independently regulated. This study provides insights into the genetic and cellular regulatory mechanisms underlying orange spot formation, which would help to elucidate how these processes are evolutionarily maintained as ornamental traits relevant to sexual selection.

The effect of the doublesex gene in body colour masculinization of the damselfly Ischnura senegalensis.

Odonata species display a remarkable diversity of colour patterns, including intrasexual polymorphisms. In the damselfly (Ischnura senegalensis), the expression of a sex-determining transcription factor, the doublesex (Isdsx) gene is reportedly associated with female colour polymorphism (CP) (gynomorph for female-specific colour and andromorph for male-mimicking colour). Here, the function of Isdsx in thoracic coloration was investigated by electroporation-mediated RNA interference (RNAi). RNAi of the Isdsx common region in males and andromorphic females reduced melanization and thus changed the colour pattern into that of gynomorphic females, while the gynomorphic colour pattern was not affected. By contrast, RNAi against the Isdsx long isoform produced no changes, suggesting that the Isdsx short isoform is important for body colour masculinization in both males and andromorphic females. When examining the expression levels of five genes with differences between sexes and female morphs, two melanin-suppressing genes, black and ebony, were expressed at higher levels in the Isdsx RNAi body area than a control area. Therefore, the Isdsx short isoform may induce thoracic colour differentiation by suppressing black and ebony, thereby generating female CP in I. senegalensis. These findings contribute to the understanding of the molecular and evolutionary mechanisms underlying female CP in Odonata.

Detection of genes positively selected in Cuban Anolis lizards that naturally inhabit hot and open areas and currently thrive in urban areas.

Species of Anolis lizards of the West Indies that naturally inhabit hot and open areas also tend to thrive in urban areas. In this study, transcriptome was sequenced for nine species of Cuban Anolis lizards that are closely related to each other, but inhabit different thermal microhabitats. Using PAML and HyPhy software, we attempted to identify genes and amino acid sites under positive selection in the common ancestral branch of A. porcatus and A. allisoni, and the branch of A. sagrei, which inhabit hot and open areas, and thrive in urban areas. Although there were no genes where positive selection was commonly detected on both of the tested branches, positive selection was detected in genes involved in the stress response (e.g., DNA damage and oxidative stress) and cardiac function, which could be related to adaptive evolution of tolerance to heat or ultraviolet radiation, on both branches. These findings suggest that adaptive evolution of the response to stress caused by heat or ultraviolet radiation might have occurred in ancestors of Anolis species inhabiting hot and open areas and might be related to the current thriving in urban areas of them.

Identification of a reptile lyssavirus in Anolis allogus provided novel insights into lyssavirus evolution.

Lyssaviruses (genus Lyssavirus) are negative-strand RNA viruses belonging to the family Rhabdoviridae. Although a lyssa-like virus (frog lyssa-like virus 1 [FLLV-1]), which is distantly related to lyssaviruses, was recently identified in frogs, a large phylogenetic gap exists between those viruses, and thus the evolution of lyssaviruses is unclear. In this study, we detected a lyssa-like virus from publicly available RNA-seq data obtained using the brain and skin of Anolis allogus (Spanish flag anole), which was designated anole lyssa-like virus 1 (ALLV-1), and determined its complete coding sequence. Via mapping analysis, we demonstrated that ALLV-1 was actively replicating in the original brain and skin samples. Phylogenetic analyses revealed that ALLV-1 is more closely related to lyssaviruses than FLLV-1. Overall, the topology of the tree is compatible with that of hosts, suggesting the long-term co-divergence of lyssa-like and lyssaviruses and vertebrates. The ψ region, which is a long 3' untranslated region of unknown origin present in the G mRNA of lyssaviruses (approximately 400-700 nucleotides), is also present in the genome of ALLV-1, but it is much shorter (approximately 180 nucleotides) than those of lyssaviruses. Interestingly, FLLV-1 lacks the ψ region, suggesting that the ψ region was acquired after the divergence of the FLLV-1 and ALLV-1/lyssavirus lineages. To the best of our knowledge, this is the first report to identify a lyssa-like virus in reptiles, and thus, our findings provide novel insights into the evolution of lyssaviruses.

Estimating possible bumblebee range shifts in response to climate and land cover changes.

Wild bee decline has been reported worldwide. Some bumblebee species (Bombus spp.) have declined in Europe and North America, and their ranges have shrunk due to climate and land cover changes. In countries with limited historical and current occurrence data, it is often difficult to investigate bumblebee range shifts. Here we estimated the past/present distributions of six major bumblebee species in Japan with species distribution modeling using current occurrence data and past/present climate and land cover data. The differences identified between estimated past and present distributions indicate possible range shifts. The estimated ranges of B. diversus, B. hypocrita, B. ignitus, B. honshuensis, and B. beaticola shrank over the past 26 years, but that of B. ardens expanded. The lower altitudinal limits of the estimated ranges became higher as temperature increased. When focusing on the effects of land cover change, the estimated range of B. diversus slightly shrank due to an increase in forest area. Such increase in forest area may result from the abandonment of agricultural lands and the extension of the rotation time of planted coniferous forests and secondary forests. Managing old planted coniferous forests and secondary forests will be key to bumblebee conservation for adaptation to climate change.

Contradictory effects of leaf rolls in a leaf-mining weevil.

Leaf rolls by herbivorous insects evolved in various lepidopteran groups, aphids, and some attelabid weevil species. Leaf rolls are known to have a positive effect on the survival of immature insects, protecting them from natural enemies such as birds, ants, predatory wasps, and parasitoids as well as environmental stress. On the other hand, leaf rolls are considered to have a negative effect on immature survival, attracting natural enemies because of their noticeability and subsequent learning or specialization. In this study, we directly tested the effects of leaf rolls using an attelabid species by comparing the fate of immature insects between artificial leaf rolls and unrolled leaves. The results showed the following positive effects of leaf rolls: avoidance of parasitism by eulophid wasps and avoidance of egg predation by unknown predators. On the other hand, a negative effect of leaf rolls was also detected, specifically and increase in mortality via leaf roll herbivory. This study indicated that leaf shelters are not only protective refuges but are also sometimes risky hiding places, although total survival rates increased in leaf shelters.

Genetic factors for short life span associated with evolution of the loss of flight ability.

Acquisition or loss of flying ability is evolutionarily linked with maximum life span (MLS) in mammals and birds. Although ecological factors, such as extrinsic mortality, may lead to either shortened or extended life spans through natural selection, MLS is influenced by complex molecular and metabolic processes, and the genetic changes associated with flying ability that have led to either a longer or shorter MLS are unknown. Here, we examine the parallel evolution of flight in mammals and birds and investigate positively selected genes at branches where either the acquisition (in little brown bats and large flying foxes) or loss (in Adélie penguins, emperor penguins, common ostriches, emus, great spotted kiwis, little spotted kiwis, okarito brown kiwis, greater rheas, lesser rheas, and cassowaries) of flight abilities occurred. Although we found no shared genes under selection among all the branches of interest, 7 genes were found to be positively selected in 2 of the branches. Among the 7 genes, only IGF2BP2 is known to affect both life span and energy expenditure. The positively selected mutations detected in IGF2BP2 likely affected the functionality of the encoded protein. IGF2BP2, which has been reported to simultaneously prolong life span and increase energy expenditure, could be responsible for the evolution of shortened MLS associated with the loss of flying ability.

Genetic segregation for male body coloration and female mate preference in the guppy.

OBJECTIVE: The purpose of this study was to segregate the genetic lines responsible for the orange area of coloration in males and the response to orange coloration exhibited by females in the guppy (Poecilia reticulata) through artificial selection. This study is part of a project that uses QTL-seq to search for candidate genes involved in male orange coloration and female response to male coloration. We created two lines: high-selected lines of males having large areas of orange spots and of females with high response to male orange coloration; and low-selected lines of males having small areas of orange spots and of females with low response to male orange coloration. RESULTS: The male orange area and the female response became significantly different between high- and low-selected lines after three generations of artificial selection. This indicates that the differences in the frequencies of alleles at loci affecting the orange area and the female response between the lines increased over the generations through selection.

Environmental factors explain spawning day deviation from full moon in the scleractinian coral Acropora.

Broadcast-spawning scleractinian corals annually release their gametes with high levels of synchrony, both within and among species. However, the timing of spawning can vary inter-annually. In particular, the night of spawning relative to the full moon phase can vary considerably among years at some locations. Although multiple environmental factors can affect the night of spawning, their effects have not been quantitatively assessed at the multi-regional level. In this study, we analysed environmental factors that are potentially correlated with spawning day deviation, in relation to the full moon phase, in Acropora corals inhabiting seven reefs in Australia and Japan. We accordingly found that sea surface temperature and wind speed within one to two months prior to the full moon of the spawning month were strongly correlated with spawning day deviations. In addition, solar flux had a weak effect on the night of spawning. These findings indicate that Acropora have the capacity to adjust their development and physiology in response to environmental factors for fine-tuning the timing of synchronous spawning, thereby maximizing reproductive success and post-fertilization survival.