Convergent genomic signatures associated with vertebrate viviparity.

BACKGROUND: Viviparity-live birth-is a complex and innovative mode of reproduction that has evolved repeatedly across the vertebrate Tree of Life. Viviparous species exhibit remarkable levels of reproductive diversity, both in the amount of care provided by the parent during gestation, and the ways in which that care is delivered. The genetic basis of viviparity has garnered increasing interest over recent years; however, such studies are often undertaken on small evolutionary timelines, and thus are not able to address changes occurring on a broader scale. Using whole genome data, we investigated the molecular basis of this innovation across the diversity of vertebrates to answer a long held question in evolutionary biology: is the evolution of convergent traits driven by convergent genomic changes? RESULTS: We reveal convergent changes in protein family sizes, protein-coding regions, introns, and untranslated regions (UTRs) in a number of distantly related viviparous lineages. Specifically, we identify 15 protein families showing evidence of contraction or expansion associated with viviparity. We additionally identify elevated substitution rates in both coding and noncoding sequences in several viviparous lineages. However, we did not find any convergent changes-be it at the nucleotide or protein level-common to all viviparous lineages. CONCLUSIONS: Our results highlight the value of macroevolutionary comparative genomics in determining the genomic basis of complex evolutionary transitions. While we identify a number of convergent genomic changes that may be associated with the evolution of viviparity in vertebrates, there does not appear to be a convergent molecular signature shared by all viviparous vertebrates. Ultimately, our findings indicate that a complex trait such as viviparity likely evolves with changes occurring in multiple different pathways.

Comparing the potential for maternal-fetal signalling in oviparous and viviparous lizards.

The evolution of a placenta requires several steps including changing the timing of reproductive events, facilitating nutrient exchange, and the capacity for maternal-fetal communication. To understand the evolution of maternal-fetal communication, we used ligand-receptor gene expression as a proxy for the potential for cross-talk in a live-bearing lizard (Pseudemoia entrecasteauxii) and homologous tissues in a related egg-laying lizard (Lampropholis guichenoti). Approximately 70% of expressed ligand/receptor genes were shared by both species. Gene ontology (GO) analysis showed that there was no GO-enrichment in the fetal membranes of the egg-laying species, but live-bearing fetal tissues were significantly enriched for 50 GO-terms. Differences in enrichment suggest that the evolution of viviparity involved reinforcing specific signalling pathways, perhaps to support fetal control of placentation. One identified change was in transforming growth factor beta signalling. Using immunohistochemistry, we show the production of the signalling molecule inhibin beta B (INHBB) occurs in viviparous fetal membranes but was absent in closely related egg-laying tissues, suggesting that the evolution of viviparity may have involved changes to signalling via this pathway. We argue that maternal-fetal signalling evolved through co-opting expressed signalling molecules and recruiting new signalling molecules to support the complex developmental changes required to support a fetus in utero. This article is part of the theme issue 'Extraembryonic tissues: exploring concepts, definitions and functions across the animal kingdom'.

Understanding the evolution of viviparity using intraspecific variation in reproductive mode and transitional forms of pregnancy.

How innovations such as vision, flight and pregnancy evolve is a central question in evolutionary biology. Examination of transitional (intermediate) forms of these traits can help address this question, but these intermediate phenotypes are very rare in extant species. Here we explore the biology and evolution of transitional forms of pregnancy that are midway between the ancestral state of oviparity (egg-laying) and the derived state, viviparity (live birth). Transitional forms of pregnancy occur in only three vertebrates, all of which are lizard species that also display intraspecific variation in reproductive phenotype. In these lizards (Lerista bougainvillii, Saiphos equalis, and Zootoca vivipara), geographic variation of three reproductive forms occurs within a single species: oviparity, viviparity, and a transitional form of pregnancy. This phenomenon offers the valuable prospect of watching 'evolution in action'. In these species, it is possible to conduct comparative research using different reproductive forms that are not confounded by speciation, and are of relatively recent origin. We identify major proximate and ultimate questions that can be addressed in these species, and the genetic and genomic tools that can help us understand how transitional forms of pregnancy are produced, despite predicted fitness costs. We argue that these taxa represent an excellent prospect for understanding the major evolutionary shift between egg-laying and live birth, which is a fundamental innovation in the history of animals.

Expression atlas of avian neural crest proteins: Neurulation to migration.

Neural crest (NC) cells are a dynamic population of embryonic stem cells that create various adult tissues in vertebrate species including craniofacial bone and cartilage and the peripheral and enteric nervous systems. NC development is thought to be a conserved and complex process that is controlled by a tightly-regulated gene regulatory network (GRN) of morphogens, transcription factors, and cell adhesion proteins. While multiple studies have characterized the expression of several GRN factors in single species, a comprehensive protein analysis that directly compares expression across development is lacking. To address this lack in information, we used three closely related avian models, Gallus gallus (chicken), Coturnix japonica (Japanese quail), and Pavo cristatus (Indian peafowl), to compare the localization and timing of four GRN transcription factors, PAX7, SNAI2, SOX9, and SOX10, from the onset of neurulation to migration. While the spatial expression of these factors is largely conserved, we find that quail NC cells express SNAI2, SOX9, and SOX10 proteins at the equivalent of earlier developmental stages than chick and peafowl. In addition, quail NC cells migrate farther and more rapidly than the larger organisms. These data suggest that despite a conservation of NC GRN players, differences in the timing of NC development between species remain a significant frontier to be explored with functional studies.

Tbx15/18/22 shares a binding site with Tbx6-r.b to maintain expression of a muscle structural gene in ascidian late embryos.

The ascidian larval tail contains muscle cells for swimming. Most of these muscle cells differentiate autonomously. The genetic program behind this autonomy has been studied extensively and the genetic cascade from maternal factors to initiation of expression of a muscle structural gene, Myl.c, has been uncovered; Myl.c expression is directed initially by transcription factor Tbx6-r.b at the 64-cell stage and then by the combined actions of Tbx6-r.b and Mrf from the gastrula to early tailbud stages. In the present study, we showed that transcription of Myl.c continued in late tailbud embryos and larvae, although a fusion protein of Tbx6-r.b and GFP was hardly detectable in late tailbud embryos. A knockdown experiment, reporter assay, and in vitro binding assay indicated that an essential cis-regulatory element of Myl.c that bound Tbx6-r.b in early embryos bound Tbx15/18/22 in late embryos to maintain expression of Myl.c. We also found that Tbx15/18/22 was controlled by Mrf, which constitutes a regulatory loop with Tbx6-r.b. Therefore, our data indicated that Tbx15/18/22 was activated initially under control of this regulatory loop as in the case of Myl.c, and then Tbx15/18/22 maintained the expression of Myl.c after Tbx6-r.b had disappeared. RNA-sequencing of Tbx15/18/22 morphant embryos revealed that many muscle structural genes were regulated similarly by Tbx15/18/22. Thus, the present study revealed the mechanisms of maintenance of transcription of muscle structural genes in late embryos in which Tbx15/18/22 takes the place of Tbx6-r.b.

Independent evolutionary transitions to pueriparity across multiple timescales in the viviparous genus Salamandra.

The ability to bear live offspring, viviparity, has evolved multiple times across the tree of life and is a remarkable adaptation with profound life-history and ecological implications. Within amphibians the ancestral reproductive mode is oviparity followed by a larval life stage, but viviparity has evolved independently in all three amphibian orders. Two types of viviparous reproduction can be distinguished in amphibians; larviparity and pueriparity. Larviparous amphibians deliver larvae into nearby ponds and streams, while pueriparous amphibians deliver fully developed juveniles and thus do not require waterbodies for reproduction. Among amphibians, the salamander genus Salamandra is remarkable as it exhibits both inter- and intraspecific variation in the occurrence of larviparity and pueriparity. While the evolutionary relationships among Salamandra lineages have been the focus of several recent studies, our understanding of how often and when transitions between modes occurred is still incomplete. Furthermore, in species with intraspecific variation, the reproductive mode of a given population can only be confirmed by direct observation of births and thus the prevalence of pueriparous populations is also incompletely documented. We used sequence capture to obtain 1,326 loci from 94 individuals from across the geographic range of the genus, focusing on potential reproductive mode transition zones. We also report additional direct observations of pueriparous births for 20 new locations and multiple lineages. We identify at least five independent transitions from the ancestral mode of larviparity to pueriparity among and within species, occurring at different evolutionary timescales ranging from the Pliocene to the Holocene. Four of these transitions occurred within species. Based on a distinct set of markers and analyses, we also confirm previous findings of introgression between species and the need for taxonomic revisions in the genus. We discuss the implications of our findings with respect to the evolution of this complex trait, and the potential of using five independent convergent transitions for further studies on the ecological context in which pueriparity evolves and the genetic architecture of this specialized reproductive mode.

The Evolution of Viviparity in Vertebrates.

In the vertebrate tree of life, viviparity or live birth has independently evolved many times, resulting in a rich diversity of reproductive strategies. Viviparity is believed to be a mode of reproduction that evolved from the ancestral condition of oviparity or egg laying, where most of the fetal development occurs outside the body. Today, there is not a simple model of parity transition to explain this species-specific divergence in modes of reproduction. Most evidence points to a gradual series of evolutionary adaptations that account for this phenomenon of reproduction, elegantly displayed by various viviparous squamates that exhibit placentae formed by the appositions of maternal and embryonic tissues, which share significant homology with the tissues that form the placenta in therian mammals. In an era where the genomes of many vertebrate species are becoming available, studies are now exploring the molecular basis of this transition from oviparity to viviparity, and in some rare instances its possible reversibility, such as the Australian three-toed skink (Saiphos equalis). In contrast to the parity diversity in squamates, mammals are viviparous with the notable exception of the egg-laying monotremes. Advancing computational tools coupled with increasing genome availability across species that utilize different reproductive strategies promise to reveal the molecular underpinnings of the ancestral transition of oviparity to viviparity. As a result, the dramatic changes in reproductive physiology and anatomy that accompany these parity changes can be reinterpreted. This chapter will briefly explore the vertebrate modes of reproduction using a phylogenetic framework and where possible highlight the role of potential candidate genes that may help explain the polygenic origins of live birth.

Evolution of reproductive strategies in the species-rich land snail subfamily Phaedusinae (Stylommatophora: Clausiliidae).

Most of the present knowledge on animal reproductive mode evolution, and possible factors driving transitions between oviparity and viviparity is based on studies on vertebrates. The species rich door snail (Clausiliidae) subfamily Phaedusinae represents a suitable and unique model for further examining parity evolution, as three different strategies, oviparity, viviparity, and the intermediate mode of embryo-retention, occur in this group. The present study reconstructs the evolution of reproductive strategies in Phaedusinae based on time-calibrated molecular phylogenetics, reproductive mode examinations and ancestral state reconstruction. Our phylogenetic analysis employing multiple mitochondrial and nuclear markers identified a well-supported clade (including the tribes Phaedusini and Serrulinini) that contains species exhibiting various reproductive strategies. This clade evolved from an oviparous most recent common ancestor according to our reconstruction. All non-oviparous taxa are confined to a highly supported subclade, coinciding with the tribe Phaedusini. Both oviparity and viviparity occur frequently in different lineages of this subclade that are not closely related. During Phaedusini diversification, multiple transitions in reproductive strategy must have taken place, which could have been promoted by a high fitness of embryo-retaining species. The evolutionary success of this group might result from the maintenance of various strategies.

Genetic improvement of egg laying traits in Fayoumi chickens bred under conditions of heat stress through selection and gene expression studies.

Thermal stress has been shown to result in decreased egg production, decreased eggshell quality, and ultimately millions of dollars in losses to the industry. Therefore, there are many factors to consider when implementing genetic selection programs aimed at improving egg production under tropical conditions. So, trial is trying to improve the productivity and eggshell quality traits of the Fayoumi chicken under high ambient temperatures via selection programs and gene expression. In the present study, day-old Fayoumi chicks were raised either under normal temperature (control) or conditions of thermal stress (the heated group). At 35 weeks, male and female chickens from the control group were mated randomly and females selected for higher egg production and eggshell strength were mated to male siblings to obtain the progeny of the first generation (F1). F1 birds were further selected and mated to obtain the progeny of the second generation. Our results show that egg production and eggshell strength traits improved over successive generations via selection under conditions of heat stress. Furthermore, the reduction in egg production and eggshell strength as a result of heat stress declined from one generation to the next in birds selected for good heat tolerance, and an inverse relationship was observed between the OC-17 and eggshell strength. Additionally, levels of HSP90 and gene expression increased in the two successive generations, indicating that both productivity and heat tolerance were enhanced due to selection in birds raised under conditions of thermal stress. Moreover, generation exerted an important effect on this trait. Thus, desirable traits such as improved heat tolerance in producing lines were observed in Fayoumi chickens exposed to conditions of thermal stress via selection. Therefore, modern advances in studies of poultry breeding and genetics, such as gene expression studies, should be examined.

Emergence of an evolutionary innovation: Gene expression differences associated with the transition between oviparity and viviparity.

Our understanding of the evolution of complex biological traits is greatly advanced by examining taxa with intermediate phenotypes. The transition from oviparity (egg-laying) to viviparity (live-bearing) has occurred independently in many animal lineages, but there are few phenotypic intermediates. The lizard Saiphos equalis exhibits bimodal reproduction, with some viviparous populations, and other oviparous populations with long egg-retention, a rare trait where most of embryonic development occurs inside the mother prior to late ovipositioning. We posit that oviparous S. equalis represent an intermediate form between "true" oviparity and viviparity. We used transcriptomics to compare uterine gene expression in these two phenotypes, and provide a molecular model for the genetic control and evolution of reproductive mode. Many genes are differentially expressed throughout the reproductive cycle of both phenotypes, which have clearly different gene expression profiles overall. The differentially expressed genes within oviparous and viviparous individuals have broadly similar biological functions putatively important for sustaining embryos, including uterine remodelling, respiratory gas and water exchange, and immune regulation. These functional similarities indicate either that long egg-retention is an exaptation for viviparity, or might reflect parallel evolution of similar gravidity-related changes in gene expression in long egg-retention oviparity. In contrast, gene expression changes across the reproductive cycle of long egg-retaining oviparous S. equalis are dramatically different from those of "true" oviparous skinks (such as Lampropholis guichenoti), supporting our assertion that oviparous S. equalis exhibit an intermediate phenotype between "true" oviparity and viviparity.

Twins! Microsatellite analysis of two embryos within one egg case in oviparous elasmobranchs.

Elasmobranchs display various reproductive modes, which have been key to their evolutionary success. In recent decades there has been a rise in the number of reported cases of foetal abnormalities including fertilised, double-embryos held within one egg capsule, hereafter referred to as twins. Previously, the occurrences of twin egg cases have been reported in two batoid and one shark species. We report the first cases of twins in three species of oviparous elasmobranchs: the undulate ray (Raja undulata), the nursehound (Scyliorhinus stellaris), and the small-spotted catshark (Scyliorhinus canicula). We investigated the genetic relationships between the twins in S. stellaris, and S. canicula using microsatellite markers. Whilst the S. stellaris twins displayed the same genotypes, we found that the S. canicula twin individuals arose through heteropaternal superfecundation. This is the first reported incidence of such a paternity in elasmobranchs. The relationship between environmental change and reproductive strategy in elasmobranchs is unclear and further research is needed to determine its effect on the prevalence and mechanisms of formation of elasmobranch twins.

Comparative landscape genetics reveals the evolution of viviparity reduces genetic connectivity in fire salamanders.

Evolutionary changes in reproductive mode may affect co-evolving traits, such as dispersal, although this subject remains largely underexplored. The shift from aquatic oviparous or larviparous reproduction to terrestrial viviparous reproduction in some amphibians entails skipping the aquatic larval stage and, thus, greater independence from water. Accordingly, amphibians exhibiting terrestrial viviparous reproduction may potentially disperse across a wider variety of suboptimal habitats and increase population connectivity in fragmented landscapes compared to aquatic-breeding species. We investigated this hypothesis in the fire salamander (Salamandra salamandra), which exhibits both aquatic- (larviparity) and terrestrial-breeding (viviparity) strategies. We genotyped 426 larviparous and 360 viviparous adult salamanders for 13 microsatellite loci and sequenced a mitochondrial marker for 133 larviparous and 119 viviparous individuals to compare population connectivity and landscape resistance to gene flow within a landscape genetics framework. Contrary to our predictions, viviparous populations exhibited greater differentiation and reduced genetic connectivity compared to larviparous populations. Landscape genetic analyses indicate viviparity may be partially responsible for this pattern, as water courses comprised a significant barrier only in viviparous salamanders, probably due to their fully terrestrial life cycle. Agricultural areas and, to a lesser extent, topography also decreased genetic connectivity in both larviparous and viviparous populations. This study is one of very few to explicitly demonstrate the evolution of a derived reproductive mode affects patterns of genetic connectivity. Our findings open avenues for future research to better understand the eco-evolutionary implications underlying the emergence of terrestrial reproduction in amphibians.

Genomic and transcriptomic investigations of the evolutionary transition from oviparity to viviparity.

Viviparous (live-bearing) vertebrates have evolved repeatedly within otherwise oviparous (egg-laying) clades. Over two-thirds of these changes in vertebrate reproductive parity mode happened in squamate reptiles, where the transition has happened between 98 and 129 times. The transition from oviparity to viviparity requires numerous physiological, morphological, and immunological changes to the female reproductive tract, including eggshell reduction, delayed oviposition, placental development for supply of water and nutrition to the embryo by the mother, enhanced gas exchange, and suppression of maternal immune rejection of the embryo. We performed genomic and transcriptomic analyses of a closely related oviparous-viviparous pair of lizards (Phrynocephalus przewalskii and Phrynocephalus vlangalii) to examine these transitions. Expression patterns of maternal oviduct through reproductive development of the egg and embryo differ markedly between the two species. We found changes in expression patterns of appropriate genes that account for each of the major aspects of the oviparity to viviparity transition. In addition, we compared the gene sequences in transcriptomes of four oviparous-viviparous pairs of lizards in different genera (Phrynocephalus, Eremias, Scincella, and Sphenomorphus) to look for possible gene convergence at the sequence level. We discovered low levels of convergence in both amino acid replacement and evolutionary rate shift. This suggests that most of the changes that produce the oviparity-viviparity transition are changes in gene expression, so occasional reversals to oviparity from viviparity may not be as difficult to achieve as has been previously suggested.

Embryonic estrogen exposure recapitulates persistent ovarian transcriptional programs in a model of environmental endocrine disruption.

Estrogens regulate key aspects of sexual determination and differentiation, and exposure to exogenous estrogens can alter ovarian development. Alligators inhabiting Lake Apopka, FL, are historically exposed to estrogenic endocrine disrupting contaminants and are characterized by a suite of reproductive abnormalities, including altered ovarian gene expression and abated transcriptional responses to follicle stimulating hormone. Here, we test the hypothesis that disrupting estrogen signaling during gonadal differentiation results in persistent alterations to ovarian gene expression that mirror alterations observed in alligators from Lake Apopka. Alligator embryos collected from a reference site lacking environmental contamination were exposed to estradiol-17 beta or a nonaromatizable androgen in ovo and raised to the juvenile stage. Changes in basal and gonadotropin-challenged ovarian gene expression were then compared to Apopka juveniles raised under identical conditions. Assessing basal transcription in untreated reference and Apopka animals revealed a consistent pattern of differential expression of key ovarian genes. For each gene where basal expression differed across sites, in ovo estradiol treatment in reference individuals recapitulated patterns observed in Apopka alligators. Among those genes affected by site and estradiol treatment were three aryl hydrocarbon receptor (AHR) isoforms, suggesting that developmental estrogen signaling might program sensitivity to AHR ligands later in life. Treatment with gonadotropins stimulated strong ovarian transcriptional responses; however, the magnitude of responses was not strongly affected by steroid hormone treatment. Collectively, these findings demonstrate that precocious estrogen signaling in the developing ovary likely underlies altered transcriptional profiles observed in a natural population exposed to endocrine disrupting contaminants.

Comparative proteomic analysis of hypothalamus tissue from Huoyan geese between pre-laying period and laying period using an iTRAQ-based approach.

The hypothalamus plays a central role in controlling poultry endocrine and reproductive activities. So far there is limited information focused on the proteome profiles of the hypothalamus from geese during different stages of the egg-laying cycle. In order to identify proteins regulating the egg-laying process of Huoyan geese, we investigated the proteome profiles of the hypothalamus from Huoyan geese during the laying period and pre-laying period by applying an isobaric tags for relative and absolute quantitation (iTRAQ)-based proteomic technology. A total number of 3,337 were identified and quantified, of which 18 were significantly up-regulated and 16 were significantly down-regulated. These differentially expressed proteins were subjected to bioinformatics analyses based on the Gene Ontology annotation and Kyoto Encyclopedia of Genes and Genomes pathway. Some of these were revealed to be involved in hormone and neurotransmitter secretion, exocytosis, calcium ion transport and synaptic transmission. Subsequently, excitatory amino acid transporter 2, complexin-1 and inositol 1,4,5-trisphosphate receptor, type 3 were confirmed at the messenger RNA level using quantitative real-time RT-PCR. Then, the abundance change of these proteins was verified further using Western blotting analysis. These data may aid in elucidating the molecular mechanism of higher laying performance in Huoyan geese.

Molecular characterization, expression and cellular localization of CYP17 gene during geese (Anser cygnoides) follicular development.

As a key member of the cytochrome P450 gene superfamily, CYP17 gene encodes 17α-hydroxylase/C17,20-lyase that is critical for directing androgen synthesis. The CYP17 gene has been identified in several species, yet little is known about its distribution and expression profile during goose follicular development. In the present study, we obtained the full-length coding sequence of goose CYP17 (gCYP17) gene for the first time using RACE method. Its sequence alignment and phylogenetic analysis suggested that gCYP17 was highly conserved with those of other birds and consisted of four main functional domains like other species. Results from immunohistochemistry, quantitative real-time PCR and Western blotting suggested that gCYP17 was predominantly located in theca interna throughout follicular development. Furthermore, levels of gCYP17 reached the maximum in theca layer of the 6-8 mm follicles which were significantly higher than in those of other follicles (P < 0.05). In addition, gCYP17 was expressed at much higher levels in the F4 theca layer than the F1 follicle (P < 0.05). Therefore, these results indicated that the fluctuating expression pattern and specific cellular localization of gCYP17 during follicular development might be closely related to androgen secretion, and thereby follicular maturation.

The genetics of egg retention and fertilization success in Drosophila: One step closer to understanding the transition from facultative to obligate viviparity.

Oviparous, facultative egg retention enables Drosophila females to withhold fertilized eggs in their reproductive tracts until circumstances favor oviposition. The propensity to retain fertilized eggs varies greatly between species, and is correlated with other reproductive traits, such as egg size and ovariole number. While previous studies have described the phenomenon, no study to date has characterized within-species variation or the genetic basis of the trait. Here, we develop a novel microscope-based method for measuring egg retention in Drosophila females and determine the range of phenotypic variation in mated female egg retention in a subset of 91 Drosophila Genetic Reference Panel (DGRP) lines. We inferred the genetic basis of egg retention using a genome-wide association study (GWAS). Further, the scoring of more than 95,000 stained, staged eggs enabled estimates of fertilization success for each line. We found evidence that ovary- and spermathecae-related genes as well as genes affecting olfactory behavior, male mating behavior, male-female attraction and sperm motility may play a crucial role in post-mating physiology. Based on our findings we also propose potential evolutionary routes toward obligate viviparity. In particular, we propose that the loss of fecundity incurred by viviparity could be offset by benefits arising from enhanced mate discrimination, resource specialization, or modified egg morphology.

Crustacean cardioactive peptides: Expression, localization, structure, and a possible involvement in regulation of egg-laying in the cuttlefish Sepia officinalis.

The cuttlefish (Sepia officinalis) is a cephalopod mollusk distributed on the western European coast, in the West African Ocean and in the Mediterranean Sea. On the Normandy coast (France), cuttlefish is a target species of professional fishermen, so its reproduction strategy is of particular interest in the context of stock management. Egg-laying, which is coastal, is controlled by several types of regulators among which neuropeptides. The cuttlefish neuropeptidome was recently identified by Zatylny-Gaudin et al. (2016). Among the 38 neuropeptide families identified, some were significantly overexpressed in egg-laying females as compared to mature males. This study is focused on crustacean cardioactive peptides (CCAPs), a highly expressed neuropeptide family strongly suspected of being involved in the control of egg-laying. We investigated the functional and structural characterization and tissue mapping of CCAPs, as well as the expression patterns of their receptors. CCAPs appeared to be involved in oocyte transport through the oviduct and in mechanical secretion of capsular products. Immunocytochemistry revealed that the neuropeptides were localized throughout the central nervous system (CNS) and in the nerve endings of the glands involved in egg-capsule synthesis and secretion, i.e. the oviduct gland and the main nidamental glands. The CCAP receptor was expressed in these glands and in the subesophageal mass of the CNS. Multiple sequence alignments revealed a high level of conservation of CCAP protein precursors in Sepia officinalis and Loligo pealei, two cephalopod decapods. Primary sequences of CCAPs from the two species were fully conserved, and cryptic peptides detected in the nerve endings were also partially conserved, suggesting biological activity that remains unknown for the time being.

An integrated omic analysis of hepatic alteration in medaka fish chronically exposed to cyanotoxins with possible mechanisms of reproductive toxicity.

Cyanobacterial blooms threaten human health as well as the population of other living organisms in the aquatic environment, particularly due to the production of natural toxic components, the cyanotoxin. So far, the most studied cyanotoxins are microcystins (MCs). In this study, the hepatic alterations at histological, proteome and transcriptome levels were evaluated in female and male medaka fish chronically exposed to 1 and 5 µg L-1 microcystin-LR (MC-LR) and to the extract of MC-producing Microcystis aeruginosa PCC 7820 (5 µg L-1 of equivalent MC-LR) by balneation for 28 days, aiming at enhancing our understanding of the potential reproductive toxicity of cyanotoxins in aquatic vertebrate models. Indeed, both MC and Microcystis extract adversely affect reproductive parameters including fecundity and egg hatchability. The liver of toxin treated female fish present glycogen storage loss and cellular damages. The quantitative proteomics analysis revealed that the quantities of 225 hepatic proteins are dysregulated. In particular, a notable decrease in protein quantities of vitellogenin and choriogenin was observed, which could explain the decrease in reproductive output. Liver transcriptome analysis through Illumina RNA-seq reveals that over 100-400 genes are differentially expressed under 5 µg L-1 MC-LR and Microcystis extract treatments, respectively. Ingenuity pathway analysis of the omic data attests that various metabolic pathways, such as energy production, protein biosynthesis and lipid metabolism, are disturbed by both MC-LR and the Microcystis extract, which could provoke the observed reproductive impairment. The transcriptomics analysis also constitutes the first report of the impairment of circadian rhythm-related gene induced by MCs. This study contributes to a better understanding of the potential consequences of chronic exposure of fish to environmental concentrations of cyanotoxins, suggesting that Microcystis extract could impact a wider range of biological pathways, compared with pure MC-LR, and even 1 µg L-1 MC-LR potentially induces a health risk for aquatic organisms.

Reptile Pregnancy Is Underpinned by Complex Changes in Uterine Gene Expression: A Comparative Analysis of the Uterine Transcriptome in Viviparous and Oviparous Lizards.

The evolution of new organs is difficult to study because most vertebrate organs evolved only once, more than 500 million years ago. An ideal model for understanding complex organ evolution is the placenta, a structure that is present in live bearing reptiles and mammals (amniotes), which has evolved independently more than 115 times. Using transcriptomics, we characterized the uterine gene expression patterns through the reproductive cycle of a viviparous skink lizard, Pseudemoia entrecasteauxii Then we compare these patterns with the patterns of gene expression from two oviparous skinks Lampropholis guichenoti and Lerista bougainvillii While thousands of genes are differentially expressed between pregnant and non-pregnant uterine tissue in the viviparous skink, few differentially expressed genes were identified between gravid and non-gravid oviparous skinks. This finding suggests that in P. entrecasteauxii, a pregnant-specific gene expression profile has evolved, allowing for the evolution of pregnancy-specific innovations in the uterus. We find substantial gene expression differences between the uterus of the chorioallantoic and the yolk sac placenta in P. entrecasteauxii, suggesting these placental regions are specialized for different placental functions. In particular, the chorioallantoic placenta is likely a major site of nutrient transport by membrane-bound transport proteins, while the yolk sac placenta also likely transports nutrients but via apocrine secretions. We discuss how the evolution of transcription factor networks is likely to underpin the evolution of the new transcriptional states in the uterine tissue of viviparous reptiles.