Despite the widely accepted involvement of DNA methylation in the regulation of rDNA transcription, the relative participation of different cytosine methylation pathways is currently described only for a few model plants. Using PacBio, Bisulfite, and RNA sequencing; PCR; Southern hybridizations; and FISH, the epigenetic consequences of rDNA copy number variation were estimated in two T. porrifolius lineages, por1 and por2, the latter with more than twice the rDNA copy numbers distributed approximately equally between NORs on chromosomes A and D. The lower rDNA content in por1 correlated with significantly reduced (>90%) sizes of both D-NORs. Moreover, two (L and S) prominent rDNA variants, differing in the repetitive organization of intergenic spacers, were detected in por2, while only the S-rDNA variant was detected in por1. Transcriptional activity of S-rDNA in por1 was associated with secondary constriction of both A-NORs. In contrast, silencing of S-rDNA in por2 was accompanied by condensation of A-NORs, secondary constriction on D-NORs, and L-rDNA transcriptional activity, suggesting (i) bidirectional nucleolar dominance and (ii) association of S-rDNAs with A-NORs and L-rDNAs with D-NORs in T. porrifolius. Each S- and L-rDNA array was formed of several sub-variants differentiating both genetically (specific SNPs) and epigenetically (transcriptional efficiency and cytosine methylation). The most significant correlations between rDNA silencing and methylation were detected for symmetric CWG motifs followed by CG motifs. No correlations were detected for external cytosine in CCGs or asymmetric CHHs, where methylation was rather position-dependent, particularly for AT-rich variants. We conclude that variations in rDNA copy numbers in plant diploids can be accompanied by prompt epigenetic responses to maintain an appropriate number of active rDNAs. The methylation dynamics of CWGs are likely to be the most responsible for regulating silent and active rDNA states.

• Klíčová slova
• 35S rDNA copy number variations, CCGs and CHHs, CWGs, Tragopogon porrifolius ssp. porrifolius, bidirectional nucleolar dominance, methylation dynamics of CGs, transcriptional silencing/activation,
• MeSH
• chromozomy rostlin genetika   MeSH
• cytosin * metabolismus   MeSH
• epigeneze genetická MeSH
• genetická transkripce MeSH
• metylace DNA * MeSH
• regulace genové exprese u rostlin MeSH
• ribozomální DNA * genetika   MeSH
• umlčování genů * MeSH
• variabilita počtu kopií segmentů DNA MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• cytosin * MeSH
• ribozomální DNA * MeSH

Although both are salient features of genomes, at first glance ribosomal DNAs and transposable elements are genetic elements with not much in common: whereas ribosomal DNAs are mainly viewed as housekeeping genes that uphold all prime genome functions, transposable elements are generally portrayed as selfish and disruptive. These opposing characteristics are also mirrored in other attributes: organization in tandem (ribosomal DNAs) versus organization in a dispersed manner (transposable elements); evolution in a concerted manner (ribosomal DNAs) versus evolution by diversification (transposable elements); and activity that prolongs genomic stability (ribosomal DNAs) versus activity that shortens it (transposable elements). Re-visiting relevant instances in which ribosomal DNA-transposable element interactions have been reported, we note that both repeat types share at least four structural and functional hallmarks: (1) they are repetitive DNAs that shape genomes in evolutionary timescales, (2) they exchange structural motifs and can enter co-evolution processes, (3) they are tightly controlled genomic stress sensors playing key roles in senescence/aging, and (4) they share common epigenetic marks such as DNA methylation and histone modification. Here, we give an overview of the structural, functional, and evolutionary characteristics of both ribosomal DNAs and transposable elements, discuss their roles and interactions, and highlight trends and future directions as we move forward in understanding ribosomal DNA-transposable element associations.

• Klíčová slova
• concerted evolution, genome size, genome stability, homogenization, housekeeping genes, long-read sequencing, molecular cytogenetics, recombination, repetitive DNA, ribosomal DNA, transposable elements, transposition,
• MeSH
• cytogenetické vyšetření MeSH
• genomika * MeSH
• metylace DNA MeSH
• molekulární evoluce MeSH
• ribozomální DNA MeSH
• transpozibilní elementy DNA * MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• ribozomální DNA MeSH
• transpozibilní elementy DNA * MeSH

Genes for major ribosomal RNAs (rDNA) are present in multiple copies mainly organized in tandem arrays. The number and position of rDNA loci can change dynamically and their repatterning is presumably driven by other repetitive sequences. We explored a peculiar rDNA organization in several representatives of Lepidoptera with either extremely large or numerous rDNA clusters. We combined molecular cytogenetics with analyses of second- and third-generation sequencing data to show that rDNA spreads as a transcription unit and reveal association between rDNA and various repeats. Furthermore, we performed comparative long read analyses among the species with derived rDNA distribution and moths with a single rDNA locus, which is considered ancestral. Our results suggest that satellite arrays, rather than mobile elements, facilitate homology-mediated spread of rDNA via either integration of extrachromosomal rDNA circles or ectopic recombination. The latter arguably better explains preferential spread of rDNA into terminal regions of lepidopteran chromosomes as efficiency of ectopic recombination depends on the proximity of homologous sequences to telomeres.

• Klíčová slova
• Lepidoptera, major ribosomal RNA genes, mobile elements, satellite,
• MeSH
• chromozomy MeSH
• můry * genetika   MeSH
• repetitivní sekvence nukleových kyselin * MeSH
• ribozomální DNA genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ribozomální DNA MeSH

Genes encoding ribosomal RNA (rDNA) are essential for cell survival and are particularly sensitive to factors leading to genomic instability. Their repetitive character makes them prone to inappropriate recombinational events arising from collision of transcriptional and replication machineries, resulting in unstable rDNA copy numbers. In this review, we summarize current knowledge on the structure and organization of rDNA, its role in sensing changes in the genome, and its linkage to aging. We also review recent findings on the main factors involved in chromatin assembly and DNA repair in the maintenance of rDNA stability in the model plants Arabidopsis thaliana and the moss Physcomitrella patens, providing a view across the plant evolutionary tree.

• Klíčová slova
• CAF-1, RAD51, RTEL1, genome stability, rDNA organization, rRNA genes, ribosome,
• MeSH
• Arabidopsis genetika   MeSH
• DNA rostlinná genetika   MeSH
• genetická transkripce MeSH
• genová dávka MeSH
• lidé MeSH
• mechy genetika   MeSH
• nestabilita genomu MeSH
• oprava DNA * MeSH
• replikace DNA MeSH
• restrukturace chromatinu MeSH
• ribozomální DNA genetika   MeSH
• stárnutí genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• DNA rostlinná MeSH
• ribozomální DNA MeSH

Genes encoding 45S ribosomal RNA (rDNA) are known for their abundance within eukaryotic genomes and for their unstable copy numbers in response to changes in various genetic and epigenetic factors. Commonly, we understand as epigenetic factors (affecting gene expression without a change in DNA sequence), namely DNA methylation, histone posttranslational modifications, histone variants, RNA interference, nucleosome remodeling and assembly, and chromosome position effect. All these were actually shown to affect activity and stability of rDNA. Here, we focus on another phenomenon - the potential of DNA containing shortly spaced oligo-guanine tracts to form quadruplex structures (G4). Interestingly, sites with a high propensity to form G4 were described in yeast, animal, and plant rDNAs, in addition to G4 at telomeres, some gene promoters, and transposons, suggesting the evolutionary ancient origin of G4 as a regulatory module. Here, we present examples of rDNA promoter regions with extremely high potential to form G4 in two model plants, Arabidopsis thaliana and Physcomitrella patens. The high G4 potential is balanced by the activity of G4-resolving enzymes. The ability of rDNA to undergo these "structural gymnastics" thus represents another layer of the rich repertoire of epigenetic regulations, which is pronounced in rDNA due to its highly repetitive character.

• Klíčová slova
• G4, quadruplex DNA, rDNA stability, replication, ribosomal RNA genes, transcription,
• Publikační typ
• časopisecké články MeSH

The intergenic spacer (IGS) of rDNA is frequently built of long blocks of tandem repeats. To estimate the intragenomic variability of such knotty regions, we employed PacBio sequencing of the Cucurbita moschata genome, in which thousands of rDNA copies are distributed across a number of loci. The rRNA coding regions are highly conserved, indicating intensive interlocus homogenization and/or high selection pressure. However, the IGS exhibits high intragenomic structural diversity. Two repeated blocks, R1 (300-1250 bp) and R2 (290-643 bp), account for most of the IGS variation. They exhibit minisatellite-like features built of multiple periodically spaced short GC-rich sequence motifs with the potential to adopt non-canonical DNA conformations, G-quadruplex-folded and left-handed Z-DNA. The mutual arrangement of these motifs can be used to classify IGS variants into five structural families. Subtle polymorphisms exist within each family due to a variable number of repeats, suggesting the coexistence of an enormous number of IGS variants. The substantial length and structural heterogeneity of IGS minisatellites suggests that the tempo of their divergence exceeds the tempo of the homogenization of rDNA arrays. As frequently occurring among plants, we hypothesize that their instability may influence transcription regulation and/or destabilize rDNA units, possibly spreading them across the genome.

• Klíčová slova
• Cucurbita moschata, DNA-minisatellite, intragenomic structural heterogeneity, non-canonical DNA conformations, ribosomal DNA intergenic spacer,
• MeSH
• Cucurbita genetika   MeSH
• genetická variace * MeSH
• konformace nukleové kyseliny * MeSH
• mezerníky ribozomální DNA chemie   genetika   metabolismus   MeSH
• minisatelitní repetice * MeSH
• sekvenční analýza DNA MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• mezerníky ribozomální DNA MeSH

BACKGROUND: Pikes 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. RESULTS: The 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. CONCLUSIONS: Extreme 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.

• Klíčová slova
• Chromosome, Esox, Evolution, Fish, Single cell PacBio sequencing, rDNA,
• MeSH
• Esocidae genetika   MeSH
• fylogeneze MeSH
• genetické lokusy genetika   MeSH
• genomika * MeSH
• genová dávka MeSH
• heterochromatin metabolismus   MeSH
• konzervovaná sekvence MeSH
• metylace DNA * MeSH
• ribozomální DNA genetika   MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza hybridizací s uspořádaným souborem oligonukleotidů MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• heterochromatin MeSH
• ribozomální DNA MeSH

BACKGROUND: Ribosomal RNA (rRNA) accounts for the majority of the RNA in eukaryotic cells, and is encoded by hundreds to thousands of nearly identical gene copies, only a subset of which are active at any given time. In Arabidopsis thaliana, 45S rRNA genes are found in two large ribosomal DNA (rDNA) clusters and little is known about the contribution of each to the overall transcription pattern in the species. RESULTS: By taking advantage of genome sequencing data from the 1001 Genomes Consortium, we characterize rRNA gene sequence variation within and among accessions. Notably, variation is not restricted to the pre-rRNA sequences removed during processing, but it is also present within the highly conserved ribosomal subunits. Through linkage mapping we assign these variants to a particular rDNA cluster unambiguously and use them as reporters of rDNA cluster-specific expression. We demonstrate that rDNA cluster-usage varies greatly among accessions and that rDNA cluster-specific expression and silencing is controlled via genetic interactions between entire rDNA cluster haplotypes (alleles). CONCLUSIONS: We show that rRNA gene cluster expression is controlled via complex epistatic and allelic interactions between rDNA haplotypes that apparently regulate the entire rRNA gene cluster. Furthermore, the sequence polymorphism we discovered implies that the pool of rRNA in a cell may be heterogeneous, which could have functional consequences.

• Klíčová slova
• Dominance, Epistasis, Ribosomes, Transcription, rRNA genes,
• MeSH
• alely MeSH
• Arabidopsis genetika   MeSH
• genetická epistáze * MeSH
• haplotypy MeSH
• multigenová rodina * MeSH
• regulace genové exprese u rostlin * MeSH
• RNA ribozomální genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Názvy látek
• RNA ribozomální MeSH
• RNA, ribosomal, 45S MeSH Prohlížeč