Exploration of an anglerfish genome

Doctoral thesis (PhD) - Nord University, 2019The anglerfishes comprise an extremely diverse order of teleosts with unique adaptations. The most notable is sexually parasitism of reproduction where the male attaches to the female. This can result in fusion of two genetically distinct organisms, which would in most vertebrate species result in an immune rejection. However, in sexually parasitic anglerfish fusion occurs with no immune rejection. The mechanisms that have allowed the evolution of such adaptations are of interest not just to evolutionary biology, but perhaps also to biomedical research related to the prevention of allogenic rejection after transplantation. Nevertheless, anglerfishes remain poorly understood. In this project we have produced the first chromosome level assembly of an anglerfish (Lophius piscatorius). We also provide an annotation of this genome based on orthology inference and believe that this will provide a comprehensive genetic resource for the study of anglerfish biology facilitating research addressing the evolution of anglerfish specific properties. As part of an analysis of the initial contig level assembly we characterized the L. piscatorius mitochondrial genome and transcriptome. This identified low-level heteroplasmic sites, a species-specific control region indel, as well as a novel long non- coding RNA derived from the Cytochrome Oxidase I locus. Furthermore, we observed a remarkable sequence conservation of the mitochondrial-derived peptide Humanin. These findings contribute to our understanding of mitochondrial regulation and function, and are of interest not only to anglerfish research. It is thought that sexual parasitism has evolved independently multiple times within the Ceratioidei suborder, suggesting that they may share a common genetic predisposition that facilitates sexual parasitism. As the removal of immune rejection is a requirement for the fusion of two individuals it is possible that this predisposition arises from a modified immune system that may be shared with the non-parasitic anglerfish taxons. Given that two teleost taxons (Gadiformes and Syngnathus) have previously been reported to lack the MHC II arm of the adaptive immune system we made use of the initial draft genome assemblies to establish the absence or presence of MHC II in L. piscatorius. Surprisingly we found an absence of exactly the same five (of 30 assayed) genes absent in Gadiformes. This observation implies that these five genes (CD4, CD74 A/B, MHCIIα/β) comprise a core set of MHCII genes that have no essential functions external to MHC II, and suggests the possibility that loss of MHCII may have been one of the events that enabled the development of sexual parasitism in anglerfish. To annotate the final chromosome level assemblies, we made use of in silico gene predictions supported by evidence from RNA followed by an orthology based functional annotation. An analysis of the resulting annotation confirmed that L. piscatorius has a fairly typical teleost genome in terms of genome size, global gene repertoire and gene feature composition. We also observed a chromosomal orthology with several teleost species that argues that the scaffolds reported here do indeed represent physical chromosomes. These analyses also revealed a teleost specific bimodality in intron length distribution that could be correlated to genome size within the teleosts, suggesting a coupling between the mechanisms governing intron and genome size in teleosts. The work presented in this thesis not only provides new genome resources that should facilitate further research into the weird and wonderful world of the anglerfishes, but also confirms an unexpected plasticity in teleost adaptive immunity. Surprisingly we were also able to observe fundamental genome properties related to intron size that have not previously been reported. Our work thus touches not only on the specifics of teleost immunology but also on general mechanisms underlying genome evolution in the teleosts.publishedVersio

Molecular characterization of the deep-water sea anemone Protanthea simplex reveals true novel features in mitogenome organization

Masteroppgave i marin økologi - Universitetet i Nordland, 2015Sperra for utlån til 2020-06-2

The mitochondrial transcriptome of the anglerfish Lophius piscatorius

Objective Analyze key features of the anglerfish Lophius piscatorius mitochondrial transcriptome based on high-throughput total RNA sequencing. Results We determined the complete mitochondrial DNA and corresponding transcriptome sequences of L. piscatorius. Key features include highly abundant mitochondrial ribosomal RNAs (10–100 times that of mRNAs), and that cytochrome oxidase mRNAs appeared > 5 times more abundant than both NADH dehydrogenase and ATPase mRNAs. Unusual for a vertebrate mitochondrial mRNA, the polyadenylated COI mRNA was found to harbor a 75 nucleotide 3′ untranslated region. The mitochondrial genome expressed several non-canonical genes, including the long noncoding RNAs lncCR-H, lncCR-L and lncCOI. Whereas lncCR-H and lncCR-L mapped to opposite strands in a non-overlapping organization within the control region, lncCOI appeared novel among vertebrates. We found lncCOI to be a highly abundant mitochondrial RNA in antisense to the COI mRNA. Finally, we present the coding potential of a humanin-like peptide within the large subunit ribosomal RNA.publishedVersio

The Mitochondrial Genome of the Sea Anemone Stichodactyla haddoni Reveals Catalytic Introns, Insertion-Like Element, and Unexpected Phylogeny

A hallmark of sea anemone mitochondrial genomes (mitogenomes) is the presence of complex catalytic group I introns. Here, we report the complete mitogenome and corresponding transcriptome of the carpet sea anemone Stichodactyla haddoni (family Stichodactylidae). The mitogenome is vertebrate-like in size, organization, and gene content. Two mitochondrial genes encoding NADH dehydrogenase subunit 5 (ND5) and cytochrome c oxidase subunit I (COI) are interrupted with complex group I introns, and one of the introns (ND5-717) harbors two conventional mitochondrial genes (ND1 and ND3) within its sequence. All the mitochondrial genes, including the group I introns, are expressed at the RNA level. Nonconventional and optional mitochondrial genes are present in the mitogenome of S. haddoni. One of these gene codes for a COI-884 intron homing endonuclease and is organized in-frame with the upstream COI exon. The insertion-like orfA is expressed as RNA and translocated in the mitogenome as compared with other sea anemones. Phylogenetic analyses based on complete nucleotide and derived protein sequences indicate that S. haddoni is embedded within the family Actiniidae, a finding that challenges current taxonomy

Intron size minimisation in teleosts

Background: Spliceosomal introns are parts of primary transcripts that are removed by RNA splicing. Although introns apparently do not contribute to the function of the mature transcript, in vertebrates they comprise the majority of the transcribed region increasing the metabolic cost of transcription. The persistence of long introns across evolutionary time suggests functional roles that can offset this metabolic cost. The teleosts comprise one of the largest vertebrate clades. They have unusually compact and variable genome sizes and provide a suitable system for analysing intron evolution. Results: We have analysed intron lengths in 172 vertebrate genomes and show that teleost intron lengths are relatively short, highly variable and bimodally distributed. Introns that were long in teleosts were also found to be long in mammals and were more likely to be found in regulatory genes and to contain conserved sequences. Our results argue that intron length has decreased in parallel in a non-random manner throughout teleost evolution and represent a deviation from the ancestral state. Conclusion: Our observations indicate an accelerated rate of intron size evolution in the teleosts and that teleost introns can be divided into two classes by their length. Teleost intron sizes have evolved primarily as a side-effect of genome size evolution and small genomes are dominated by short introns (<256 base pairs). However, a non-random subset of introns has resisted this process across the teleosts and these are more likely have functional roles in all vertebrate clades

Complete loss of the MHC II pathway in an anglerfish, Lophius piscatorius

Genome studies in fish provide evidence for the adaptability of the vertebrate immune system, revealing alternative immune strategies. The reported absence of the major compatibility complex (MHC) class II pathway components in certain species of pipefish (genus Syngnathus) and cod-like fishes (order Gadiformes) is of particular interest. The MHC II pathway is responsible for immunization and defence against extracellular threats through the presentation of exogenous peptides to T helper cells. Here, we demonstrate the absence of all genes encoding MHC II components (CD4, CD74 A/B, and both classical and non-classical MHC II α/β) in the genome of an anglerfish, Lophius piscatorius, indicating loss of the MHC II pathway. By contrast, it has previously been reported that another anglerfish, Antennarius striatus, retains all MHC II genes, placing the loss of MHC II in the Lophius clade to their most recent common ancestor. In the three taxa where MHC II loss has occurred, the gene loss has been restricted to four or five core MHC II components, suggesting that, in teleosts, only these genes have functions that are restricted to the MHC II pathway.publishedVersionPaid Open Acces

Navigating sex and sex roles: deciphering sex-biased gene expression in a species with sex-role reversal (Syngnathus typhle)

Sexual dimorphism, the divergence in morphological traits between males and females of the same species, is often accompanied by sex-biased gene expression. However, the majority of research has focused on species with conventional sex roles, where females have the highest energy burden with both egg production and parental care, neglecting the diversity of reproductive roles found in nature. We investigated sex-biased gene expression in Syngnathus typhle, a sex-role reversed species with male pregnancy, allowing us to separate two female traits: egg production and parental care. Using RNA sequencing, we examined gene expression across organs (brain, head kidney and gonads) at various life stages, encompassing differences in age, sex and reproductive status. While some gene groups were more strongly associated with sex roles, such as stress resistance and immune defence, others were driven by biological sex, such as energy and lipid storage regulation in an organ- and age-specific manner. By investigating how genes regulate and are regulated by changing reproductive roles and resource allocation in a model system with an unconventional life-history strategy, we aim to better understand the importance of sex and sex role in regulating gene expression patterns, broadening the scope of this discussion to encompass a wide range of organisms

The effects of primary and secondary bacterial exposure on the seahorse (Hippocampus erectus) immune response

Highlights: • Transcriptomic immune response assessments in seahorse (Hippocampus erectus). • Seahorses exposed in two phases to heat-killed Vibrio and Tenacibaculum strains. • Adaptive immune memory evidence (double-exposed) and increased naivety to Tenacibaculum. • Upregulated gene expression pertaining to potential innate ‘trained immunity’. • Trained immunity potential compensator for deduced MHC II loss of function. Evolutionary adaptations in the Syngnathidae teleost family (seahorses, pipefish and seadragons) culminated in an array of spectacular morphologies, key immune gene losses, and the enigmatic male pregnancy. In seahorses, genome modifications associated with immunoglobulins, complement, and major histocompatibility complex (MHC II) pathway components raise questions concerning their immunological efficiency and the evolution of compensatory measures that may act in their place. In this investigation heat-killed bacteria (Vibrio aestuarianus and Tenacibaculum maritimum) were used in a two-phased experiment to assess the immune response dynamics of Hippocampus erectus. Gill transcriptomes from double and single-exposed individuals were analysed in order to determine the differentially expressed genes contributing to immune system responses towards immune priming. Double-exposed individuals exhibited a greater adaptive immune response when compared with single-exposed individuals, while single-exposed individuals, particularly with V. aestuarianus replicates, associated more with the innate branch of the immune system. T. maritimum double-exposed replicates exhibited the strongest immune reaction, likely due to their immunological naivety towards the bacterium, while there are also potential signs of innate trained immunity. MHC II upregulated expression was identified in selected V. aestuarianus-exposed seahorses, in the absence of other pathway constituents suggesting a possible alternative or non-classical MHC II immune function in seahorses. Gene Ontology (GO) enrichment analysis highlighted prominent angiogenesis activity following secondary exposure, which could be linked to an adaptive immune process in seahorses. This investigation highlights the prominent role of T-cell mediated adaptive immune responses in seahorses when exposed to sequential foreign bacteria exposures. If classical MHC II pathway function has been lost, innate trained immunity in syngnathids could be a potential compensatory mechanism