Higher-order organisation of extremely amplified, potentially functional and massively methylated 5S rDNA in European pikes (Esox sp.)

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.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-017-3774-7) contains supplementary material, which is available to authorized users.

Satellite DNA landscapes after allotetraploidisation of quinoa (Chenopodium quinoa) reveal unique A and B subgenomes

“…For C. quinoa, we identified arrays of the satellite families ChenSat-1a, ChenSat-1b, ChenSat-2b, ChenSat-2c, ChenSat-2d, and ChenSat-2e on SMRT reads; all were arranged in short or long homogeneous arrays, but also in structures of higher order (ChenSat-1a, ChenSat-1b, ChenSat-2c, and ChenSat-2e) and head-to-head arrangements (ChenSat-1a, ChenSat-2b, and ChenSat-2e). Similar to observations in the pike genome (Symonová et al, 2017), we also detect inversions in the 5S rDNA, and long as well as short arrays. Higher-order arrangement of ChenSat-1a has also been detected on clones of multimers as reported very recently (Belyayev et al, 2019).…”

“…For non-model organisms, assemblies often contain even less information making the study of repetitive regions laborious (Peona et al, 2018). The availability of thirdgeneration long reads opens the way to solve genomic and evolutionary questions targeting satellite and ribosomal DNAs, such as array length, abundance and organisation in higher-15 order structures or head-to-head arrangements (Sevim et al, 2016, Khost et al, 2017, Symonová et al, 2017, Lower et al, 2018, Cechova et al, 2019, Vondrak et al, 2019. For C. quinoa, we identified arrays of the satellite families ChenSat-1a, ChenSat-1b, ChenSat-2b, ChenSat-2c, ChenSat-2d, and ChenSat-2e on SMRT reads; all were arranged in short or long homogeneous arrays, but also in structures of higher order (ChenSat-1a, ChenSat-1b, ChenSat-2c, and ChenSat-2e) and head-to-head arrangements (ChenSat-1a, ChenSat-2b, and ChenSat-2e).…”

“…Its short monomer size of 40 bp is unusual for a tandem repeats of high abundances, which usually consist of 160-180 bp or 320 to 360 bp monomers (Hemleben et al, 2007). SatDNAs have been split into micro-(2-5 bp monomer), mini-(6-100 bp monomer), and conventional satellites (>100 bp monomer) (Vergnaud andDenoeud, 2000, Mehrotra andGoyal, 2014). In recent times, the focus has shifted to recognise not only the satDNA's monomer size, but its genomic organisation of repeating units into long arrays as the main molecular hallmark of satDNA (Richard et al, 2008).…”

The Utility of Graph Clustering of 5S Ribosomal DNA Homoeologs in Plant Allopolyploids, Homoploid Hybrids, and Cryptic Introgressants

“…Although introgression and hybridization is also relatively frequent in the banana genus (Němečková et al, 2018) a more likely explanation for complex graph structures in Musa acuminata ( Figure 6C) is an exceptionally high number of 5S rDNA loci in this species (six per diploid genome) (Valarik et al, 2002;Garcia et al, 2012a) and probably inefficient interlocus recombination ( Schlotterer and Tau tz,19 94) lea ding to poor homogenization. Actually, the mechanisms of amplification of 5S rDNA loci across the chromosomes are still poorly understood (Schubert and Wobus, 1985;Symonova et al, 2017;Joachimiak et al, 2018;Souza et al, 2019). 3.…”

Repetitive DNA profiles Reveal Evidence of Rapid Genome Evolution and Reflect Species Boundaries in Ground Beetles

“…These mobilized fragments of rDNA can be thought of newly birthed, rDNA-like repetitive elements. Mobilization of rDNA has been documented using cytogenetic methods in many groups including plants (Raskina et al 2004;Qi et al 2015;Ding et al 2016;Wang et al 2016), fish (Martins et al 2006;Da Silva et al 2012;Symonová et al 2013Symonová et al , 2017, protists (Gong et al 2013), insects (Cabral-de-Mello et al 2010, 2011Nguyen et al 2010;Panzera et al 2012;Palacios-Gimenez and Cabral-de-Mello 2015), bivalves (Pérez-García et al 2014), and mammals (Sotero-Caio et al 2015), and is regarded as strong evidence of rapid rearrangements over short time scales (Jiang and Gill 1994;Raskina et al 2004Raskina et al , 2008. Mobilization of such multicopy gene families into heterochromatic regions is thought to be mediated through processes such as retrotransposon activity (Dimitri et al 1997;Dimitri and Junakovic 1999;Symonová et al 2013;de Bello Cioffi et al 2015) and ectopic recombination (Nguyen et al 2010).…”