Teleost fish, which roughly make up half of extant vertebrate species, exhibit an amazing level of biodiversity affecting their morphology, ecology and behavior as well as most other aspects of their biology. This huge variability makes fish extremely attractive for the study of many biological questions, including those related to development and evolution. We are particularly interested in sex determination, reproduction and pigmentation, which are hypervariable in fish.
The genome sequences of the pufferfish species Takifugu rubripes (Fugu) and Tetraodon nigroviridis, the zebrafish Danio rerio, the medaka Oryzias latipes and the threespine stickleback Gasterosteus aculeatus together with genomic data from various other fish species have opened an important era of comparative genomics shedding a new light on the structure and evolution of vertebrate genomes. Upcoming genome sequences include those of the Nile Tilapia Oreochromis niloticus and of several salmonids species like the rainbow trout Oncorhynchus mykiss and the Atlantic salmon Salmo salar. Comparative analyses based on fish genomes have for example revealed the ancestral bony vertebrate genome, confirmed the occurrence of at least one event of genome duplication in the early history of vertebrates and allowed the identification of conserved regulatory and coding sequences in the human genome. Based on this huge mass of information, as well as on our own sequencing projects, we aim to identify through comparative functional genomics peculiar evolutionary mechanisms driving biodiversity in fish, with more emphasis on hypervariable traits such as sex determination, reproduction and pigmentation.
Sex determination and sex chromosome evolution in fish
In contrast to the rather stable regulatory regimes established over more that 100 million years in birds and mammals, sex determination in fish frequently underwent evolutionary changes bringing the sex-determining cascade under new master sex regulators. This phenomenon, possibly associated with the emergence of new sex chromosomes from autosomes, would explain the frequent switching between sex determination systems observed in fish.
Finally, the amazing diversity of sex determination systems and the plasticity of sex chromosomes observed in teleost might have been involved in both pre- and postmating reproductive isolation.
Even if studies are underway in different fish models, the only species for which the master sex-determining gene has been isolated so far is the medaka Oryzias latipes. In this fish, the Y-specific master sex-determining gene dmrt1bY has been formed through duplication of the autosomal gene dmrt1 onto another autosome, this generating a new Y chromosome. Dmrt1bY emerged about 10 million years ago and does not correspond to the universal master sex regulator in fish. We therefore aim to isolate master sex determining genes in other fish species, including the platyfish Xiphophorus maculatus. For this fish, we have already constructed bacterial artificial chromosome contigs covering several megabases of the X and Y chromosomes, which are currently being sequenced by Genoscope. Genes identified in silico are subsequently analyzed at the functional level in different fish models.
The identification of new sex-determining genes in these species will shed a new light on the exceptional evolutionary instability governing sex determination in fish.
In order to avoid reproduction-linked stress and to exploit sex-specific economical advantages (size, taste, etc), monosex cultures are generally used in the aquaculture. Therefore, aquaculture companies are interested in determining the sex of individual fishes at early stages of their development (molecular sexing) and even in modifying sex determination (for example through hormone treatments). We therefore plan to clone and analyse genes orthologous to the sex-determining gene SD of the platyfish in different aquaculture-relevant fish species, particularly in the Nile tilapia (collaboration with Jean-Francois Baroiller, CIRAD, Montpellier, France) and the turbot (collaboration with the aquaculture company France Turbot, Noirmoutier, France
There is now substantial evidence that a round of tetraploidization/rediploidization has taken place during the early evolution of the ray-finned fish lineage, and that hundreds of duplicate pairs generated by this event have been maintained over hundreds of millions of years of evolution. Differential loss or subfunction partitioning of such gene duplicates might have been involved in the generation of fish variability.
Importantly, major differences have been observed between teleost fish and mammalian genomes. There is now convincing evidence that all teleosts are derived from a common tetraploid fish ancestor. This tetraploidization event arose about 320-350 million years ago in the ray-finned fish lineage, followed by rediploidization and retention of hundreds of duplicate pairs. Divergent evolution of the resulting duplicates has been proposed to be involved in the species richness observed in teleost fishes.
We have identified several genes potentially involved in pigmentation in fish that are present as ancient duplicates in fish genomes but are single-copy in tetrapods. Such duplicates have been probably generated by the postulated fish-specific genome duplication. We have already started to compare the expression, function and evolution of these genes in several fish species (platyfish, guppies, medaka, tilapia, trout, pike, goldfish, zebrafish and eel) in order to identify divergent evolution of gene pairs in different fish groups. These genes will be also analyzed in more divergent species (sturgeon) in order to assess the expression pattern and function of the ancestral gene before duplication. We have access to different pigmentation variants from numerous fish species.
The genome of fishes, as those of other vertebrates and other organisms, has evolved through whole genome duplication(s). One important question concerns the evolution of paralogous genes (paranome) and genomic regions generated by this duplication in different fish lineages. We already have substantial evidence that for a given pair of duplicates, different individual copies have been maintained in different species. In order to better understand this phenomenon, we plan to isolate corresponding paralogous regions in different fish species representative of the fish lineage, as well as from ancestral species like the sturgeon, and to compare their gene content. Furthermore, we are currently developing an array hybridization method, which should allow the detection of differential gene loss in different fish species.
The evolutionary dynamics of the major families of active retrotransposable elements identified in fish will be analyzed in representative species of the teleost lineage. Particular emphasis will be given on their evolutionary history (assessed through comparison between transposable element- and host-based phylogenies), their activity, their genomic localization (assessed by fluorescence in situ hybridization) and their potential horizontal transfer. We will also investigate the general mechanisms controlling transposition in fish by testing the effect of host factors on transposable element activity.
In contrast to mammalian genomes, teleost genomes also contain multiple families of active transposable elements, which might have played a role in speciation by affecting hybrid sterility and viability.
Fish genomes also contain many more families of transposable elements than mammals and birds.
In addition, our studies have shown that fish present a high diversity of transposable elements not found in mammalian genomes. Transposable elements are mobile DNA sequences that can disrupt genes, induce the formation of diverse genomic rearrangements, influence gene expression, mobilize various types of non-autonomous sequences and move between species. Mobile DNA and transposition are considered as driving forces of genome evolution, and they are likely to play a role in the biological diversity observed in fish.
Our studies have shown that fishes present an amazing diversity of active transposable elements (TEs) not found in mammalian genomes. We plan to analyze the evolutionary impact of TEs on the diversification of gene function in fish and functionally determine (i) if transposable elements have contributed regulatory sequences important for the expression of resident genes, (ii) if transposable elements have contributed coding sequences important for the function of resident proteins and (iii) if transposable elements themselves evolved as bona fide resident genes. Particularly, le role of TEs in the diversification of gene function will be analyzed in different fish lineages. This project will be principally based on resources and tools developed by the medaka, zebrafish and pufferfish genome projects, as well as by our own platyfish sex chromosome sequencing project.