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Amphibians evolved in the Devonian period about 400 Mya and represent a transition step in tetrapod evolution. Among amphibians, high-throughput sequencing data are very limited for Caudata, due to their largest genome sizes among terrestrial vertebrates. In this paper we present the transcriptome from the fire bellied newt Cynops orientalis. Data here presented display a high level of completeness, comparable to the fully sequenced genomes available from other amphibians. Moreover, this work focused on genes involved in gametogenesis and sexual development. Surprisingly, the gsdf gene was identified for the first time in a tetrapod species, so far known only from bony fish and basal sarcopterygians. Our analysis failed to isolate fgf24 and foxl3, supporting the possible loss of both genes in the common ancestor of Rhipidistians. In Cynops, the expression analysis of genes described to be sex-related in vertebrates singled out an expected functional role for some genes, while others displayed an unforeseen behavior, confirming the high variability of the sex-related pathway in vertebrates.
Electrophysiological analyses conducted about 25 years ago detected two types of anion channels in the plasma membrane of guard cells. One type of channel responds slowly to changes in membrane voltage while the other responds quickly. Consequently, they were named SLAC, for SLow Anion Channel, and QUAC, for QUick Anion Channel. Recently, genes SLAC1 and QUAC1/ALMT12, underlying the two different anion current components, could be identified in the model plant Arabidopsis thaliana. Expression of the gene products in Xenopus oocytes confirmed the quick and slow current kinetics. In this study we provide an overview on our current knowledge on slow and quick anion channels in plants and analyze the molecular evolution of ALMT/QUAC-like and SLAC-like channels. We discovered fingerprints that allow screening databases for these channel types and were able to identify 192 (177 non-redundant) SLAC-like and 422 (402 non-redundant) ALMT/QUAC-like proteins in the fully sequenced genomes of 32 plant species. Phylogenetic analyses provided new insights into the molecular evolution of these channel types. We also combined sequence alignment and clustering with predictions of protein features, leading to the identification of known conserved phosphorylation sites in SLAC1-like channels along with potential sites that have not been yet experimentally confirmed. Using a similar strategy to analyze the hydropathicity of ALMT/QUAC-like channels, we propose a modified topology with additional transmembrane regions that integrates structure and function of these membrane proteins. Our results suggest that cross-referencing phylogenetic analyses with position-specific protein properties and functional data could be a very powerful tool for genome research approaches in general.
A comparative view on sex differentiation and gametogenesis genes in lungfish and coelacanths
(2018)
Gonadal sex differentiation and reproduction are the keys to the perpetuation of favorable gene combinations and positively selected traits. In vertebrates, several gonad development features that differentiate tetrapods and fishes are likely to be, at least in part, related to the water-to-land transition. The collection of information from basal sarcopterygians, coelacanths, and lungfishes, is crucial to improve our understanding of the molecular evolution of pathways involved in reproductive functions, since these organisms are generally regarded as “living fossils” and as the direct ancestors of tetrapods. Here, we report for the first time the characterization of >50 genes related to sex differentiation and gametogenesis in Latimeria menadoensis and Protopterus annectens. Although the expression profiles of most genes is consistent with the intermediate position of basal sarcopterygians between actinopterygian fish and tetrapods, their phylogenetic placement and presence/absence patterns often reveal a closer affinity to the tetrapod orthologs. On the other hand, particular genes, for example, the male gonad factor gsdf (Gonadal Soma-Derived Factor), provide examples of ancestral traits shared with actinopterygians, which disappeared in the tetrapod lineage.
How allopolyploids are able not only to cope but profit from their condition is a question that remains elusive, but is of great importance within the context of successful allopolyploid evolution. One outstanding example of successful allopolyploidy is the endemic Iberian cyprinid Squalius alburnoides. Previously, based on the evaluation of a few genes, it was reported that the transcription levels between diploid and triploid S. alburnoides were similar. If this phenomenon occurs on a full genomic scale, a wide functional "diploidization'' could be related to the success of these polyploids. We generated RNA-seq data from whole juvenile fish and from adult livers, to perform the first comparative quantitative transcriptomic analysis between diploid and triploid individuals of a vertebrate allopolyploid. Together with an assay to estimate relative expression per cell, it was possible to infer the relative sizes of transcriptomes. This showed that diploid and triploid S. alburnoides hybrids have similar liver transcriptome sizes. This in turn made it valid to directly compare the S. alburnoides RNA-seq transcript data sets and obtain a profile of dosage responses across the S. alburnoides transcriptome. We found that 64% of transcripts in juveniles' samples and 44% in liver samples differed less than twofold between diploid and triploid hybrids (similar expression). Yet, respectively 29% and 15% of transcripts presented accurate dosage compensation (PAA/PA expression ratio of 1 instead of 1.5). Therefore, an exact functional diploidization of the triploid genome does not occur, but a significant down regulation of gene expression in triploids was observed. However, for those genes with similar expression levels between diploids and triploids, expression is not globally strictly proportional to gene dosage nor is it set to a perfect diploid level. This quantitative expression flexibility may be a strong contributor to overcome the genomic shock, and be an immediate evolutionary advantage of allopolyploids.
All Staphylococcus aureus genomes contain a genomic island, which is termed vSa\(\alpha\) and characterized by two clusters of tandem repeat sequences, i.e. the exotoxin (set) and 'lipoprotein-like' genes (lpl). Based on their structural similarities the vSa\(\alpha\) islands have been classified as type I to IV. The genomes of highly pathogenic and particularly epidemic S. aureus strains (USA300, N315, Mu50, NCTC8325, Newman, COL, JH1 or JH9) belonging to the clonal complexes CC5 and CC8 bear a type I vSa\(\alpha\) island. Since the contribution of the lpl gene cluster encoded in the vSa\(\alpha\) island to virulence is unclear to date, we deleted the entire lpl gene cluster in S. aureus USA300. The results showed that the mutant was deficient in the stimulation of pro-inflammatory cytokines in human monocytes, macrophages and keratinocytes. Purified lipoprotein Lpl1 was further shown to elicit a TLR2-dependent response. Furthermore, heterologous expression of the USA300 lpl cluster in other S. aureus strains enhanced their immune stimulatory activity. Most importantly, the lpl cluster contributed to invasion of S. aureus into human keratinocytes and mouse skin and the non-invasive S. carnosus expressing the lpl gene cluster became invasive. Additionally, in a murine kidney abscess model the bacterial burden in the kidneys was higher in wild type than in mutant mice. In this infection model the lpl cluster, thus, contributes to virulence. The present report is one of the first studies addressing the role of the vSa\(\alpha\) encoded lpl gene cluster in staphylococcal virulence. The finding that the lpl gene cluster contributes to internalization into non-professional antigen presenting cells such as keratinocytes high-lights the lpl as a new cell surface component that triggers host cell invasion by S. aureus. Increased invasion in murine skin and an increased bacterial burden in a murine kidney abscess model suggest that the lpl gene cluster serves as an important virulence factor.
Mimicking female insects to attract male pollinators is an important strategy in sexually deceptive orchids of the genus Ophrys, and some species possess flowers with conspicuous labellum patterns. The function of the variation of the patterns remains unresolved, with suggestions that these enhance pollinator communication. We investigated the possible function of the labellum pattern in Ophrys heldreichii, an orchid species in which the conspicuous and complex labellum pattern contrasts with a dark background. The orchid is pollinated exclusively by males of the solitary bee, Eucera berlandi. Comparisons of labellum patterns revealed that patterns within inflorescences are more similar than those of other conspecific plants. Field observations showed that the males approach at a great speed and directly land on flowers, but after an unsuccessful copulation attempt, bees hover close and visually scan the labellum pattern for up to a minute. Learning experiments conducted with honeybees as an accessible model of bee vision demonstrated that labellum patterns of different plants can be reliably learnt; in contrast, patterns of flowers from the same inflorescence could not be discriminated. These results support the hypothesis that variable labellum patterns in O. heldreichii are involved in flower-pollinator communication which would likely help these plants to avoid geitonogamy.
Background: Males in some species of the genus Xiphophorus, small freshwater fishes from Meso-America, have an extended caudal fin, or sword - hence their common name "swordtails". Longer swords are preferred by females from both sworded and - surprisingly also, non-sworded (platyfish) species that belong to the same genus. Swordtails have been studied widely as models in research on sexual selection. Specifically, the pre-existing bias hypothesis was interpreted to best explain the observed bias of females in presumed ancestral lineages of swordless species that show a preference for assumed derived males with swords over their conspecific swordless males. However, many of the phylogenetic relationships within this genus still remained unresolved. Here we construct a comprehensive molecular phylogeny of all 26 known Xiphophorus species, including the four recently described species (X. kallmani, X. mayae, X. mixei and X. monticolus). We use two mitochondrial and six new nuclear markers in an effort to increase the understanding of the evolutionary relationships among the species in this genus. Based on the phylogeny, the evolutionary history and character state evolution of the sword was reconstructed and found to have originated in the common ancestral lineage of the genus Xiphophorus and that it was lost again secondarily.
Results: We estimated the evolutionary relationships among all known species of the genus Xiphophorus based on the largest set of DNA markers so far. The phylogeny indicates that one of the newly described swordtail species, Xiphophorus monticolus, is likely to have arisen through hybridization since it is placed with the southern platyfish in the mitochondrial phylogeny, but with the southern swordtails in the nuclear phylogeny. Such discordance between these two types of markers is a strong indication for a hybrid origin. Additionally, by using a maximum likelihood approach the possession of the sexually selected sword trait is shown to be the most likely ancestral state for the genus Xiphophorus. Further, we provide a well supported estimation of the phylogenetic relationships between the previously unresolved northern swordtail groups.
Conclusions: This comprehensive molecular phylogeny of the entire genus Xiphophorus provides evidence that a second swordtail species, X. monticolus, arose through hybridization. Previously, we demonstrated that X. clemenciae, another southern swordtail species, arose via hybridization. These findings highlight the potential key role of hybridization in the evolution of this genus and suggest the need for further investigations into how hybridization contributes to speciation more generally.
Background: Heterococcus is a microalgal genus of Xanthophyceae (Stramenopiles) that is common and widespread in soils, especially from cold regions. Species are characterized by extensively branched filaments produced when grown on agarized culture medium. Despite the large number of species described exclusively using light microscopic morphology, the assessment of species diversity is hampered by extensive morphological plasticity.
Results: Two independent types of molecular data, the chloroplast-encoded psbA/rbcL spacer complemented by rbcL gene and the internal transcribed spacer 2 of the nuclear rDNA cistron (ITS2), congruently recovered a robust phylogenetic structure. With ITS2 considerable sequence and secondary structure divergence existed among the eight species, but a combined sequence and secondary structure phylogenetic analysis confined to helix II of ITS2 corroborated relationships as inferred from the rbcL gene phylogeny. Intra-genomic divergence of ITS2 sequences was revealed in many strains. The 'monophyletic species concept', appropriate for microalgae without known sexual reproduction, revealed eight different species. Species boundaries established using the molecular-based monophyletic species concept were more conservative than the traditional morphological species concept. Within a species, almost identical chloroplast marker sequences (genotypes) were repeatedly recovered from strains of different origins. At least two species had widespread geographical distributions; however, within a given species, genotypes recovered from Antarctic strains were distinct from those in temperate habitats. Furthermore, the sequence diversity may correspond to adaptation to different types of habitats or climates.
Conclusions: We established a method and a reference data base for the unambiguous identification of species of the common soil microalgal genus Heterococcus which uses DNA sequence variation in markers from plastid and nuclear genomes. The molecular data were more reliable and more conservative than morphological data.
Background: Teleost fish present a high diversity of sex determination systems, with possible frequent evolutionary turnover of sex chromosomes and sex-determining genes. In order to identify genes involved in male sex determination and differentiation in the platyfish Xiphophorus maculatus, bacterial artificial chromosome contigs from the sex-determining region differentiating the Y from the X chromosome have been assembled and analyzed.
Results: A novel three-copy gene called teximY (for testis-expressed in Xiphophorus maculatus on the Y) was identified on the Y but not on the X chromosome. A highly related sequence called texim1, probably at the origin of the Y-linked genes, as well as three more divergent texim genes were detected in (pseudo) autosomal regions of the platyfish genome. Texim genes, for which no functional data are available so far in any organism, encode predicted esterases/lipases with a SGNH hydrolase domain. Texim proteins are related to proteins from very different origins, including proteins encoded by animal CR1 retrotransposons, animal platelet-activating factor acetylhydrolases (PAFah) and bacterial hydrolases. Texim gene distribution is patchy in animals. Texim sequences were detected in several fish species including killifish, medaka, pufferfish, sea bass, cod and gar, but not in zebrafish. Texim-like genes are also present in Oikopleura (urochordate), Amphioxus (cephalochordate) and sea urchin (echinoderm) but absent from mammals and other tetrapods. Interestingly, texim genes are associated with a Helitron transposon in different fish species but not in urochordates, cephalochordates and echinoderms, suggesting capture and mobilization of an ancestral texim gene in the bony fish lineage. RT-qPCR analyses showed that Y-linked teximY genes are preferentially expressed in testis, with expression at late stages of spermatogenesis (late spermatids and spermatozeugmata).
Conclusions: These observations suggest either that TeximY proteins play a role in Helitron transposition in the male germ line in fish, or that texim genes are spermatogenesis genes mobilized and spread by transposable elements in fish genomes.
Assessing allele-specific gene expression (ASE) on a large scale continues to be a technically challenging problem. Certain biological phenomena, such as X chromosome inactivation and parental imprinting, affect ASE most drastically by completely shutting down the expression of a whole set of alleles. Other more subtle effects on ASE are likely to be much more complex and dependent on the genetic environment and are perhaps more important to understand since they may be responsible for a significant amount of biological diversity. Tools to assess ASE in a diploid biological system are becoming more reliable. Non-diploid systems are, however, not uncommon. In humans full or partial polyploid states are regularly found in both healthy (meiotic cells, polynucleated cell types) and diseased tissues (trisomies, non-disjunction events, cancerous tissues). In this work we have studied ASE in the medaka fish model system. We have developed a method for determining ASE in polyploid organisms from RNAseq data and we have implemented this method in a software tool set. As a biological model system we have used nuclear transplantation to experimentally produce artificial triploid medaka composed of three different haplomes. We measured ASE in RNA isolated from the livers of two adult, triploid medaka fish that showed a high degree of similarity. The majority of genes examined (82%) shared expression more or less evenly among the three alleles in both triploids. The rest of the genes (18%) displayed a wide range of ASE levels. Interestingly the majority of genes (78%) displayed generally consistent ASE levels in both triploid individuals. A large contingent of these genes had the same allele entirely suppressed in both triploids. When viewed in a chromosomal context, it is revealed that these genes are from large sections of 4 chromosomes and may be indicative of some broad scale suppression of gene expression.