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Whole genome duplication (WGD) is a dramatic evolutionary event generating many new genes and which may play a role in survival through mass extinctions. Paddlefish and sturgeon are sister lineages that both show genomic evidence for ancient WGD. Until now this has been interpreted as two independent WGD events due to a preponderance of duplicate genes with independent histories. Here we show that although there is indeed a plurality of apparently ‘independent’ gene duplications, these derive from a shared genome duplication event occurring close to the Permian-Triassic mass extinction period, followed by a prolonged process of reversion to stable diploid inheritance (rediploidisation). We show that the sharing of this WGD is masked by the fact that paddlefish and sturgeon lineage divergence occurred before rediploidisation had proceeded even half-way. Thus, for most genes the resolution to diploidy was lineage-specific. Because genes are only truly duplicated once diploid inheritance is established, the paddlefish and sturgeon genomes are a mosaic of shared and non-shared gene duplications resulting from a shared genome duplication event. This is the first time that lineage-specific resolution of genes from a common WGD event has been shown to affect such a large proportion of the genome.

Cytokines of the TNF superfamily (TNFSF) control many immunological processes and are implicated in the etiology of many immune disorders and diseases. Despite their obvious biological importance, the TNFSF repertoires of many species remain poorly characterized. In this study, we perform detailed bioinformatic, phylogenetic, and syntenic analyses of five cartilaginous fish genomes to identify their TNFSF repertoires. Strikingly, we find that shark genomes harbor ∼30 TNFSF genes, more than any other vertebrate examined to date and substantially more than humans. This is due to better retention of the ancestral jawed vertebrate TNFSF repertoire than any other jawed vertebrate lineage, combined with lineage-specific gene family expansions. All human TNFSFs appear in shark genomes, except for lymphotoxin-α (LTA; TNFSF1) and TNF (TNFSF2), and CD70 (TNFSF7) and 4-1BBL (TNFSF9), which diverged by tandem duplications early in tetrapod and mammalian evolution, respectively. Although lacking one-to-one LTA and TNF orthologs, sharks have evolved lineage-specific clusters of LTA/TNF co-orthologs. Other key findings include the presence of two BAFF (TNFSF13B) genes along with orthologs of APRIL (TNFSF13) and BALM (TNFSF13C) in sharks, and that all cartilaginous fish genomes harbor an ∼400-million-year-old cluster of multiple FASLG (TNFSF6) orthologs. Finally, sharks have retained seven ancestral jawed vertebrate TNFSF genes lost in humans. Taken together, our data indicate that the jawed vertebrate ancestor possessed a much larger and diverse TNFSF repertoire than previously hypothesized and oppose the idea that the cartilaginous fish immune system is “primitive” compared with that of mammals.

Due to their key phylogenetic position, cartilaginous fishes, which includes the largest fish species Rhincodon typus (whale shark), are an important vertebrate lineage for understanding the origin and evolution of vertebrates. However, until recently, this lineage has been understudied in vertebrate genomics. Using newly-generated long read sequences, we produced the best gapless cartilaginous fish genome assembly to date. The assembly has fewer missing ancestral genes than Callorhinchus milii , which has been widely-used for evolutionary studies up to now. We used the new assembly to study the evolution of gene families in the whale shark and other vertebrates, focusing on historical patterns of gene family origins and loss across early vertebrate evolution, innate immune receptor repertoire evolution, and dynamics of gene family evolution size in relation to gigantism. From inferring the pattern of origin of gene families across the most recent common ancestors of major vertebrate clades, we found that there were many shared gene families between the whale shark and bony vertebrates that were present in the most recent common ancestor of jawed vertebrates, with a large increase in novel genes at the origin of jawed vertebrates independent of whole genome duplication events. The innate immune system in the whale shark, which consisted of diverse pathogen recognition receptors (PRRs) including NOD-like receptors, RIG-like receptors, and Toll-like receptors. We discovered a unique complement of Toll-like receptors and triplication of NOD1 in the whale shark genome. Further, we found diverse patterns of gene family evolution between PRRs within vertebrates demonstrating that the origin of adaptive immunity in jawed vertebrates is more complicated than simply replacing the need for a vast repertoire of germline encoded PRRs. We then studied rates of amino acid substitution and gene family size evolution across origins of vertebrate gigantism. While we found that cartilaginous fishes and giant vertebrates tended to have slower substitution rates than the background rate in vertebrates, the whale shark genome substitution rate was not significantly slower than Callorhinchus . Furthermore, rates of gene family size evolution varied among giants and the background, suggesting that differences in rate of substitution and gene family size evolution relative to gigantism are decoupled. We found that the gene families that have shifted in duplication rate in whale shark are enriched for genes related to driving cancer in humans, consistent with studies in other giant vertebrates than support the hypothesis that evolution of increased body size requires adaptations that result in reduction of per cell cancer rate.

In mammals, haptoglobin (Hp) is an acute-phase plasma protein that binds with high affinity to hemoglobin (Hb) released by intravascular hemolysis. The resultant Hp–Hb complexes are bound and cleared by the scavenger receptor CD163, limiting Hb-induced oxidative damage. In this study, we show that Hp is a divergent member of the complement-initiating MASP family of proteins, which emerged in the ancestor of jawed vertebrates. We demonstrate that Hp has been independently lost from multiple vertebrate lineages, that characterized Hb-interacting residues of mammals are poorly conserved in nonmammalian species maintaining Hp, and that the extended loop 3 region of Hp, which mediates CD163 binding, is present only in mammals. We show that the Hb-binding ability of cartilaginous fish (nurse shark, Ginglymostoma cirratum; small-spotted catshark, Scyliorhinus canicula; and thornback ray, Raja clavata) and teleost fish (rainbow trout, Oncorhynchus mykiss) Hp is species specific, and where binding does occur it is likely mediated through a different structural mechanism to mammalian Hp. The continued, high-level expression of Hp in cartilaginous fishes in which Hb binding is not evident signals that Hp has (an)other, yet unstudied, role(s) in these species. Previous work indicates that mammalian Hp also has secondary, immunomodulatory functions that are independent of Hb binding; our work suggests these may be remnants of evolutionary more ancient functions, retained after Hb removal became the primary role of Hp in mammals.

In every jawed vertebrate species studied so far, the T cell receptor (TCR) complex is composed of two different TCR chains (α/β or γ/δ) and a number of CD3 subunits responsible for transmitting signals into the T cell. In this study, we characterised all of the TCR and CD3 genes of small-spotted catshark (Scyliorhinus canicula) and analysed their expression in a broad range of tissues. While the TCR complex is highly conserved across jawed vertebrates, we identified a number of differences in catshark, most notably the presence of two copies of both TCRβ and CD3γδ, and the absence of a functionally-important proline rich region from CD3ε. We also demonstrate that TCRβ has duplicated independently multiple times in jawed vertebrate evolution, bringing additional diversity to the TCR complex. This study reveals new insights about the evolutionary history of the TCR complex and raises new avenues for future exploration.

The molecular machinery required for lymphocyte development and differentiation appears to have emerged concomitantly with distinct B- and T-like lymphocyte subsets in the ancestor of all vertebrates. The TNFSF superfamily (TNFSF) members BAFF (TNFSF13/Blys) and APRIL (TNFSF13) are key regulators of B cell development survival, and activation in mammals, but the temporal emergence of these molecules, and their precise relationship to the newly identified TNFSF gene BALM (BAFF and APRIL-like molecule), have not yet been elucidated. Here, to resolve the early evolutionary history of this family, we improved outgroup sampling and alignment quality, and applied better fitting substitution models compared to past studies. Our analyses reveal that BALM is a definitive TNFSF13 family member, which split from BAFF in the gnathostome (jawed vertebrate) ancestor. Most importantly, however, we show that both the APRIL and BAFF lineages existed in the ancestors of all extant vertebrates. This implies that APRIL has been lost, or is yet to be found, in cyclostomes (jawless vertebrates). Our results suggest that lineage-specific gene duplication and loss events have caused lymphocyte regulation, despite shared origins, to become secondarily distinct between gnathostomes and cyclostomes. Finally, the structure of lamprey BAFF-like, and its phylogenetic placement as sister to BAFF and BALM, but not the more slowly evolving APRIL, indicates that the primordial lymphocyte regulator was more APRIL-like than BAFF-like.

BACKGROUND: The cartilaginous fishes diverged from other jawed vertebrates ~ 450 million years ago (mya). Despite this key evolutionary position, the only high-quality cartilaginous fish genome available is for the elephant shark (Callorhinchus milii), a chimaera whose ancestors split from the elasmobranch lineage ~ 420 mya. Initial analysis of this resource led to proposals that key components of the cartilaginous fish adaptive immune system, most notably their array of T cell subsets, was primitive compared to mammals. This proposal is at odds with the robust, antigen-specific antibody responses reported in elasmobranchs following immunization. To explore this discrepancy, we generated a multi-tissue transcriptome for small-spotted catshark (Scyliorhinus canicula), a tractable elasmobranch model for functional studies. We searched this, and other newly available sequence datasets, for CD4+ T cell subset-defining genes, aiming to confirm the presence or absence of each subset in cartilaginous fishes.RESULTS: We generated a new transcriptome based on a normalised, multi-tissue RNA pool, aiming to maximise representation of tissue-specific and lowly expressed genes. We utilized multiple transcriptomic datasets and assembly variants in phylogenetic reconstructions to unambiguously identify several T cell subset-specific molecules in cartilaginous fishes for the first time, including interleukins, interleukin receptors, and key transcription factors. Our results reveal the inability of standard phylogenetic reconstruction approaches to capture the site-specific evolutionary processes of fast-evolving immune genes but show that site-heterogeneous mixture models can adequately do so.CONCLUSIONS: Our analyses reveal that cartilaginous fishes are capable of producing a range of CD4+ T cell subsets comparable to that of mammals. Further, that the key molecules required for the differentiation and functioning of these subsets existed in the jawed vertebrate ancestor. Additionally, we highlight the importance of considering phylogenetic diversity and, where possible, utilizing multiple datasets for individual species, to accurately infer gene presence or absence at higher taxonomic levels.

The tumour necrosis factor superfamily (TNFSF) members CD40L and BAFF play critical roles in mammalian B cell survival, proliferation and maturation, however little is known about these key cytokines in the oldest jawed vertebrates, the cartilaginous fishes. Here we report the cloning of CD40L and BAFF orthologues (designated ScCD40L and ScBAFF) in the small-spotted catshark (Scyliorhinus canicula). As predicted both proteins are type II membrane-bound proteins with a TNF homology domain in their extracellular region and both are highly expressed in shark immune tissues. ScCD40L transcript levels correlate with those of TCRα and transcription of both genes is modulated in peripheral blood leukocytes following in vitro stimulation. Although a putative CD40L orthologue was identified in the elephant shark genome the work herein is the first molecular characterisation and transcriptional analysis of CD40L in a cartilaginous fish. ScBAFF was also cloned and its transcription characterised in an attempt to resolve the discrepancies observed between spiny dogfish BAFF and bamboo shark BAFF in previously published studies.

This article will review current knowledge on CXCR in fish, that represent three distinct vertebrate groups: Agnatha (jawless fishes), Chondrichthyes (cartilaginous fishes) and Osteichthyes (bony fishes). With the sequencing of many fish genomes, information on CXCR in these species in particular has expanded considerably. In mammals, 6 CXCRs have been described, and their homologues will be initially reviewed before considering a number of atypical CXCRs and a discussion of CXCR evolution.

Background The functional divergence of duplicate genes (ohnologues) retained from whole genome duplication (WGD) is thought to promote evolutionary diversification. However, species radiation and phenotypic diversification are often temporally separated from WGD. Salmonid fish, whose ancestor underwent WGD by autotetraploidization ~95 million years ago, fit such a ‘time-lag’ model of post-WGD radiation, which occurred alongside a major delay in the rediploidization process. Here we propose a model, ‘lineage-specific ohnologue resolution’ (LORe), to address the consequences of delayed rediploidization. Under LORe, speciation precedes rediploidization, allowing independent ohnologue divergence in sister lineages sharing an ancestral WGD event. Results Using cross-species sequence capture, phylogenomics and genome-wide analyses of ohnologue expression divergence, we demonstrate the major impact of LORe on salmonid evolution. One-quarter of each salmonid genome, harbouring at least 4550 ohnologues, has evolved under LORe, with rediploidization and functional divergence occurring on multiple independent occasions >50 million years post-WGD. We demonstrate the existence and regulatory divergence of many LORe ohnologues with functions in lineage-specific physiological adaptations that potentially facilitated salmonid species radiation. We show that LORe ohnologues are enriched for different functions than ‘older’ ohnologues that began diverging in the salmonid ancestor. Conclusions LORe has unappreciated significance as a nested component of post-WGD divergence that impacts the functional properties of genes, whilst providing ohnologues available solely for lineage-specific adaptation. Under LORe, which is predicted following many WGD events, the functional outcomes of WGD need not appear ‘explosively’, but can arise gradually over tens of millions of years, promoting lineage-specific diversification regimes under prevailing ecological pressures. Electronic supplementary material The online version of this article (doi:10.1186/s13059-017-1241-z) contains supplementary material, which is available to authorized users.