Abstract

BackgroundPikes represent an important genus (Esox) harbouring a pre-duplication karyotype (2n = 2x = 50) of economically important salmonid pseudopolyploids. Here, we have characterized the 5S ribosomal RNA genes (rDNA) in Esox lucius and its closely related E. cisalpinus using cytogenetic, molecular and genomic approaches. Intragenomic homogeneity and copy number estimation was carried out using Illumina reads. The higher-order structure of rDNA arrays was investigated by the analysis of long PacBio reads. Position of loci on chromosomes was determined by FISH. DNA methylation was analysed by methylation-sensitive restriction enzymes.ResultsThe 5S rDNA loci occupy exclusively (peri)centromeric regions on 30–38 acrocentric chromosomes in both E. lucius and E. cisalpinus. The large number of loci is accompanied by extreme amplification of genes (>20,000 copies), which is to the best of our knowledge one of the highest copy number of rRNA genes in animals ever reported. Conserved secondary structures of predicted 5S rRNAs indicate that most of the amplified genes are potentially functional. Only few SNPs were found in genic regions indicating their high homogeneity while intergenic spacers were more heterogeneous and several families were identified. Analysis of 10–30 kb-long molecules sequenced by the PacBio technology (containing about 40% of total 5S rDNA) revealed that the vast majority (96%) of genes are organised in large several kilobase-long blocks. Dispersed genes or short tandems were less common (4%). The adjacent 5S blocks were directly linked, separated by intervening DNA and even inverted. The 5S units differing in the intergenic spacers formed both homogeneous and heterogeneous (mixed) blocks indicating variable degree of homogenisation between the loci. Both E. lucius and E. cisalpinus 5S rDNA was heavily methylated at CG dinucleotides.ConclusionsExtreme amplification of 5S rRNA genes in the Esox genome occurred in the absence of significant pseudogenisation suggesting its recent origin and/or intensive homogenisation processes. The dense methylation of units indicates that powerful epigenetic mechanisms have evolved in this group of fish to silence amplified genes. We discuss how the higher-order repeat structures impact on homogenisation of 5S rDNA in the genome.

Highlights

  • Pikes represent an important genus (Esox) harbouring a pre-duplication karyotype (2n = 2x = 50) of economically important salmonid pseudopolyploids

  • Localisation of 5S and 45S Ribosomal RNA genes (rDNA) loci and heterochromatin on Esox chromosomes Both E. lucius and E. cisalpinus showed the same number of chromosomes (2n = 50) exhibiting strict acrocentric morphology (FN = 50)

  • In E. cisalpinus, the 5S rDNA probe hybridised to 30–34 sites (Fig. 1a, quantitative data are summarized in Additional file 1: Figure S1), all in thecentromeric regions

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Summary

Introduction

We have characterized the 5S ribosomal RNA genes (rDNA) in Esox lucius and its closely related E. cisalpinus using cytogenetic, molecular and genomic approaches. E. masquinongy and E. niger live naturally in North America, E. reicherti is the only Euroasian esocid endemic to the Amur River basin (Russia and China) while the E. cisalpinus (E. flaviae) and E. aquitanicus are native to Europe [2]. The Northern pike (E. lucius) occurs in North America and Eurasia. The Northern pike inhabits lakes, rivers and brackish waters. It is an important commercial and recreational species. Overexploitation of the natural stocks and climate change has resulted in the dramatic decline of some pike populations [3]

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