Nucleolar dominance (ND) consists of the reversible silencing of 35S/45S rDNA loci inherited from one of the ancestors of an allopolyploid. The molecular mechanisms by which one ancestral rDNA set is selected for silencing remain unclear. We applied a combination of molecular (Southern blot hybridization and reverse-transcription cleaved amplified polymorphic sequence analysis), genomic (analysis of variants) and cytogenetic (fluorescence in situ hybridization) approaches to study the structure, expression and epigenetic landscape of 35S rDNA in an allotetraploid grass that exhibits ND, Brachypodium hybridum (genome composition DDSS), and its putative progenitors, Brachypodium distachyon (DD) and Brachypodium stacei (SS). In progenitor genomes, B. stacei showed a higher intragenomic heterogeneity of rDNA compared with B. distachyon. In all studied accessions of B. hybridum, there was a reduction in the copy number of S homoeologues, which was accompanied by their inactive transcriptional status. The involvement of DNA methylation in CG and CHG contexts in the silencing of the S-genome rDNA loci was revealed. In the B. hybridum allotetraploid, ND is stabilized towards the D-genome units, irrespective of the polyphyletic origin of the species, and does not seem to be influenced by homoeologous 35S rDNA ratios and developmental stage.

Department of Field Crops Faculty of Agriculture Tekirdag Namik Kemal University Suleymanpasa Tekirdag 59030 Turkey

Department of Molecular Epigenetics Institute of Biophysics Academy of Sciences of the Czech Republic v v i Královopolská 135 Brno 612 65 Czech Republic

Plant Cytogenetics and Molecular Biology Group Institute of Biology Biotechnology and Environmental Protection Faculty of Natural Sciences University of Silesia in Katowice Jagiellonska 28 Katowice 40 032 Poland

Tekirdag Anatolian High School Suleymanpasa Tekirdag 59030 Turkey

University California Berkeley Berkeley CA 94720 USA

US Department of Energy Berkeley CA 94720 USA

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Báez, M. , Souza, G. and Guerra, M. (2019) Does the chromosomal position of 35S rDNA sites influence their transcription? A survey on Nothoscordum species (Amaryllidaceae). Genet. Mol. Biol. 10.1590/1678-4685-GMB-2018-0194. PubMed DOI PMC

Bockor, V.V. , Barisic, D. , Horvat, T. , Maglica, Z. , Vojta, A. and Zoldos, V. (2014) Inhibition of DNA methylation alters chromatin organization, nuclear positioning and activity of 45S rDNA loci in cycling cells of Q. robur . PLoS One, 9, e103954. PubMed PMC

Borisjuk, N.V. , Davidjuk, Y.M. , Kostishin, S.S. , Miroshnichenco, G.P. , Velasco, R. , Hemleben, V. and Volkov, R.A. (1997) Structural analysis of rDNA in the genus Nicotiana . Plant Mol. Biol. 35, 655–660. PubMed

Borowska‐Zuchowska, N. and Hasterok, R. (2017) Epigenetics of the preferential silencing of Brachypodium stacei‐originated 35S rDNA loci in the allotetraploid grass Brachypodium hybridum . Sci. Rep. 7, 5260. PubMed PMC

Borowska‐Zuchowska, N. , Kwasniewski, M. and Hasterok, R. (2016) Cytomolecular analysis of ribosomal DNA evolution in a natural allotetraploid Brachypodium hybridum and its putative ancestors ‐ dissecting complex repetitive structure of intergenic spacers. Front. Plant Sci. 7, 1499. PubMed PMC

Borowska‐Zuchowska, N. , Robaszkiewicz, E. , Wolny, E. , Betekhtin, A. and Hasterok, R. (2019) Ribosomal DNA loci derived from Brachypodium stacei are switched off for major parts of the life cycle of Brachypodium hybridum . J. Exp. Bot. 70, 805–815. PubMed PMC

Castilho, A. , Queiroz, A. , Neves, N. , Barao, A. , Silva, M. and Viegas, W. (1995) The developmental stage of inactivation of rye origin rRNA genes in the embryo and endosperm of wheat x rye F1 hybrids. Chromosome Res. 3, 169–174. PubMed

Catalan, P. , Muller, J. , Hasterok, R. , Jenkins, G. , Mur, L.A. , Langdon, T. , Betekhtin, A. , Siwinska, D. , Pimentel, M. and Lopez‐Alvarez, D. (2012) Evolution and taxonomic split of the model grass Brachypodium distachyon . Ann. Bot. 109, 385–405. PubMed PMC

Chen, Z.J. and Pikaard, C.S. (1997a) Epigenetic silencing of RNA polymerase I transcription: a role for DNA methylation and histone modification in nucleolar dominance. Gene Dev. 11, 2124–2136. PubMed PMC

Chen, Z.J. and Pikaard, C.S. (1997b) Transcriptional analysis of nucleolar dominance in polyploid plants: biased expression/silencing of progenitor rRNA genes is developmentally regulated in Brassica . Proc. Natl Acad. Sci. USA, 94, 3442–3447. PubMed PMC

Clarkson, J.J. , Lim, K.Y. , Kovarik, A. , Chase, M.W. , Knapp, S. and Leitch, A.R. (2005) Long‐term genome diploidization in allopolyploid Nicotiana section Repandae (Solanaceae). New Phytol. 168, 241–252. PubMed

Costa‐Nunes, P. , Kim, J.Y. , Hong, E. and Pontes, O. (2014) The cytological and molecular role of DOMAINS REARRANGED METHYLTRANSFERASE3 in RNA‐dependent DNA methylation of Arabidopsis thaliana . BMC Res. Notes, 7, 721. PubMed PMC

Costa‐Nunes, P. , Pontes, O. , Preuss, S.B. and Pikaard, C.S. (2010) Extra views on RNA‐dependent DNA methylation and MBD6‐dependent heterochromatin formation in nucleolar dominance. Nucleus, 1, 254–259. PubMed PMC

Dadejova, M. , Lim, K.Y. , Souckova‐Skalicka, K. , Matyasek, R. , Grandbastien, M.A. , Leitch, A. and Kovarik, A. (2007) Transcription activity of rRNA genes correlates with a tendency towards intergenomic homogenization in Nicotiana allotetraploids. New Phytol. 174, 658–668. PubMed

Diaz‐Perez, A. , Lopez‐Alvarez, D. , Sancho, R. and Catalan, P. (2018) Reconstructing the origins and the biogeography of species' genomes in the highly reticulate allopolyploid‐rich model grass genus Brachypodium using minimum evolution, coalescence and maximum likelihood approaches. Mol. Phylogenet. Evol. 127, 256–271. PubMed

Dobesova, E. , Malinska, H. , Matyasek, R. , Leitch, A.R. , Soltis, D.E. , Soltis, P.S. and Kovarik, A. (2015) Silenced rRNA genes are activated and substitute for partially eliminated active homeologs in the recently formed allotetraploid, Tragopogon mirus (Asteraceae). Heredity, 114, 356–365. PubMed PMC

Dolezel, J. , Bartos, J. , Voglmayr, H. and Greilhuber, J. (2003) Nuclear DNA content and genome size of trout and human. Cytometry A, 51, 127–128. PubMed

Doyle, J. (1991) DNA protocols for plants In Molecular Techniques in Taxonomy (Hewitt G.M., Johnston A.W.B. and Young J.P.W., eds). Berlin, Heidelberg: Springer, Berlin Heidelberg, pp. 283–293.

Earley, K. , Lawrence, R.J. , Pontes, O. , Reuther, R. , Enciso, A.J. , Silva, M. , Neves, N. , Gross, M. , Viegas, W. and Pikaard, C.S. (2006) Erasure of histone acetylation by Arabidopsis HDA6 mediates large‐scale gene silencing in nucleolar dominance. Gene Dev. 20, 1283–1293. PubMed PMC

Eickbush, T.H. and Eickbush, D.G. (2007) Finely orchestrated movements: evolution of the ribosomal RNA genes. Genetics, 175, 477–485. PubMed PMC

Frieman, M. , Chen, Z.J. , Saez‐Vasquez, J. , Shen, L.A. and Pikaard, C.S. (1999) RNA polymerase I transcription in a Brassica interspecific hybrid and its progenitors: tests of transcription factor involvement in nucleolar dominance. Genetics, 152, 451–460. PubMed PMC

Ganley, A.R.D. and Kobayashi, T. (2007) Highly efficient concerted evolution in the ribosomal DNA repeats: total rDNA repeat variation revealed by whole‐genome shotgun sequence data. Genome Res. 17, 184–191. PubMed PMC

Garcia, S. , Kovarik, A. , Leitch, A.R. and Garnatje, T. (2017) Cytogenetic features of rRNA genes across land plants: analysis of the Plant rDNA database. Plant J. 89, 1020–1030. PubMed

Gerlach, W.L. and Dyer, T.A. (1980) Sequence organization of the repeating units in the nucleus of wheat which contain 5S ribosomal‐RNA genes. Nucleic Acids Res. 8, 4851–4865. PubMed PMC

Guo, X. and Han, F. (2014) Asymmetric epigenetic modification and elimination of rDNA sequences by polyploidization in wheat. Plant Cell, 26, 4311–4327. PubMed PMC

Hasterok, R. , Draper, J. and Jenkins, G. (2004) Laying the cytotaxonomic foundations of a new model grass, Brachypodium distachyon (L.) Beauv. Chromosome Res. 12, 397–403. PubMed

Hasterok, R. and Maluszynska, J. (2000) Nucleolar dominance does not occur in root tip cells of allotetraploid Brassica species. Genome, 43, 574–579. PubMed

Houchins, K. , O'Dell, M. , Flavell, R.B. and Gustafson, J.P. (1997) Cytosine methylation and nucleolar dominance in cereal hybrids. Mol. Gen. Genet. 255, 294–301. PubMed

IBI (2010) Genome sequencing and analysis of the model grass Brachypodium distachyon . Nature, 463, 763–768. PubMed

Idziak, D. , Betekhtin, A. , Wolny, E. , Lesniewska, K. , Wright, J. , Febrer, M. , Bevan, M.W. , Jenkins, G. and Hasterok, R. (2011) Painting the chromosomes of Brachypodium: current status and future prospects. Chromosoma, 120, 469–479. PubMed PMC

Idziak, D. and Hasterok, R. (2008) Cytogenetic evidence of nucleolar dominance in allotetraploid species of Brachypodium . Genome, 51, 387–391. PubMed

Jenkins, G. and Hasterok, R. (2007) BAC 'landing' on chromosomes of Brachypodium distachyon for comparative genome alignment. Nat. Protoc. 2, 88–98. PubMed

Kiss, T. , Kis, M. , Abel, S. and Solymosy, F. (1988) Nucleotide sequence of the 17S–25S spacer region from tomato rDNA. Nucleic Acids Res. 16, 7179. PubMed PMC

Kolano, B. , Mccann, J. , Oskędra, M. , Chrapek, M. , Rojek, M. , Nobis, A. and Weiss‐Schneeweiss, H. (2019) Parental origin and genome evolution of several Eurasian hexaploid species of Chenopodium (Chenopodiaceae). Phytotaxa, 392, 163–185.

Komarova, N.Y. , Grabe, T. , Huigen, D.J. , Hemleben, V. and Volkov, R.A. (2004) Organization, differential expression and methylation of rDNA in artificial Solanum allopolyploids. Plant Mol. Biol. 56, 439–463. PubMed

Kotseruba, V. , Gernand, D. , Meister, A. and Houben, A. (2003) Uniparental loss of ribosomal DNA in the allotetraploid grass Zingeria trichopoda (2n = 8). Genome, 46, 156–163. PubMed

Kovarik, A. , Dadejova, M. , Lim, Y.K. , Chase, M.W. , Clarkson, J.J. , Knapp, S. and Leitch, A.R. (2008) Evolution of rDNA in Nicotiana allopolyploids: a potential link between rDNA homogenization and epigenetics. Ann. Bot. 101, 815–823. PubMed PMC

Kovarik, A. , Pires, J.C. , Leitch, A.R. , Lim, K.Y. , Sherwood, A.M. , Matyasek, R. , Rocca, J. , Soltis, D.E. and Soltis, P.S. (2005) Rapid concerted evolution of nuclear ribosomal DNA in two Tragopogon allopolyploids of recent and recurrent origin. Genetics, 169, 931–944. PubMed PMC

Lawrence, R.J. , Earley, K. , Pontes, O. , Silva, M. , Chen, Z.J. , Neves, N. , Viegas, W. and Pikaard, C.S. (2004) A concerted DNA methylation/histone methylation switch regulates rRNA gene dosage control and nucleolar dominance. Mol. Cell, 13, 599–609. PubMed

Leitch, A.R. and Leitch, I.J. (2008) Genomic plasticity and the diversity of polyploid plants. Science, 320, 481–483. PubMed

Lim, K.Y. , Matyasek, R. , Kovarik, A. and Leitch, A. (2007) Parental origin and genome evolution in the allopolyploid Iris versicolor . Ann. Bot. 100, 219–224. PubMed PMC

Liu, B. and Wendel, J.F. (2003) Epigenetic phenomena and the evolution of plant allopolyploids. Mol. Phylogenet. Evol. 29, 365–379. PubMed

Liu, Y. , Forrest, L.L. , Bainard, J.D. , Budke, J.M. and Goffinet, B. (2013) Organellar genome, nuclear ribosomal DNA repeat unit, and microsatellites isolated from a small‐scale of 454 GS FLX sequencing on two mosses. Mol. Phylogenet. Evol. 66, 1089–1094. PubMed

Lunerova, J. , Renny‐Byfield, S. , Matyasek, R. , Leitch, A. and Kovarik, A. (2017) Concerted evolution rapidly eliminates sequence variation in rDNA coding regions but not in intergenic spacers in Nicotiana tabacum allotetraploid. Plant Syst. Evol. 303, 1043–1060.

Lusinska, J. , Majka, J. , Betekhtin, A. , Susek, K. , Wolny, E. and Hasterok, R. (2018) Chromosome identification and reconstruction of evolutionary rearrangements in Brachypodium distachyon, B. stacei and B. hybridum . Ann. Bot. 122, 445–459. PubMed PMC

Mahelka, V. , Kopecky, D. and Baum, B.R. (2013) Contrasting patterns of evolution of 45S and 5S rDNA families uncover new aspects in the genome constitution of the agronomically important grass Thinopyrum intermedium (Triticeae). Mol. Biol. Evol. 30, 2065–2086. PubMed

Mathieu, O. , Picard, G. and Tourmente, S. (2002) Methylation of a euchromatin‐heterochromatin transition region in Arabidopsis thaliana chromosome 5 left arm. Chromosome Res. 10, 455–466. PubMed

Mohannath, G. , Pontvianne, F. and Pikaard, C.S. (2016) Selective nucleolus organizer inactivation in Arabidopsis is a chromosome position‐effect phenomenon. Proc. Natl. Acad. Sci. USA, 113, 13426–13431. PubMed PMC

Naidoo, K. , Steenkamp, E.T. , Coetzee, M.P. , Wingfield, M.J. and Wingfield, B.D. (2013) Concerted evolution in the ribosomal RNA cistron. PLoS One, 8, e59355. PubMed PMC

Navashin, M. (1934) Chromosomal alterations caused by hybridization and their bearing upon certain general genetic problems. Cytologia, 5, 169–203.

Neves, N. , Delgado, M. , Silva, M. , Caperta, A. , Morais‐Cecilio, L. and Viegas, W. (2005) Ribosomal DNA heterochromatin in plants. Cytogenet. Genome Res. 109, 104–111. PubMed

Neves, N. , Heslop‐Harrison, J.S. and Viegas, W. (1995) rRNA gene activity and control of expression mediated by methylation and imprinting during embryo development in wheat x rye hybrids. Theor. Appl. Genet. 91, 529–533. PubMed

Nicoloff, H. , Anastassovakristeva, M. , Rieger, R. and Kunzel, G. (1979) Nucleolar dominance as observed in barley translocation lines with specifically reconstructed SAT chromosomes. Theor. Appl. Genet. 55, 247–251. PubMed

Phillips, R.L. , Kleese, R.A. and Wang, S.S. (1971) Nucleolus organizer region of maize (Zea mays L.) ‐ chromosomal site of DNA complementary to ribosomal RNA. Chromosoma, 36, 79–88.

Pikaard, C.S. (2000) Nucleolar dominance: uniparental gene silencing on a multi‐megabase scale in genetic hybrids. Plant Mol. Biol. 43, 163–177. PubMed

Pontes, O. , Lawrence, R.J. , Silva, M. , Preuss, S. , Costa‐Nunes, P. , Earley, K. , Neves, N. , Viegas, W. and Pikaard, C.S. (2007) Postembryonic establishment of megabase‐scale gene silencing in nucleolar dominance. PLoS One, 2, e1157. PubMed PMC

Preuss, S.B. , Costa‐Nunes, P. , Tucker, S. et al (2008) Multimegabase silencing in nucleolar dominance involves siRNA‐directed DNA methylation and specific methylcytosine‐binding proteins. Mol. Cell, 32, 673–684. PubMed PMC

Rabanal, F.A. , Mandakova, T. , Soto‐Jimenez, L.M. et al (2017) Epistatic and allelic interactions control expression of ribosomal RNA gene clusters in Arabidopsis thaliana . Genome Biol. 18, 75. PubMed PMC

Robicheau, B.M. , Susko, E. , Harrigan, A.M. and Snyder, M. (2017) Ribosomal RNA genes contribute to the formation of pseudogenes and junk DNA in the human genome. Genome Biol. Evol. 9, 380–397. PubMed PMC

Rosato, M. , Kovarik, A. , Garilleti, R. and Rossello, J.A. (2016) Conserved organisation of 45S rDNA sites and rDNA gene copy number among major clades of early land plants. PLoS One, 11, e0162544. PubMed PMC

Rozas, J. , Ferrer‐Mata, A. , Sánchez‐DelBarrio, J.C. , Guirao‐Rico, S. , Librado, P. , Ramos‐Onsins, S.E. and Sánchez‐Gracia, A. (2017) DnaSP 6: DNA sequence polymorphism analysis of large data sets. Mol. Biol. Evol. 34, 3299–3302. PubMed

Schubert, I. and Künzel, G. (1990) Position‐dependent NOR activity in barley. Chromosoma, 99, 352–359.

Shaw, P. (2013) The plant nucleolus In Plant Genome Diversity, vol 2, (Leitch I.J., Greilhuber J., Dolezel J. and Wendel J.F., eds). Wien: Springer‐Verlag, pp. 65–76.

Sochorova, J. , Coriton, O. , Kuderova, A. , Lunerova, J. , Chevre, A.M. and Kovarik, A. (2017) Gene conversion events and variable degree of homogenization of rDNA loci in cultivars of Brassica napus . Ann. Bot. 119, 13–26. PubMed PMC

Symonova, R. (2019) Integrative rDNAomics‐Importance of the oldest repetitive fraction of the eukaryote genome. Genes, 10(5), 345. PubMed PMC

Szostak, J.W. and Wu, R. (1980) Unequal crossing over in the ribosomal DNA of Saccharomyces cerevisiae . Nature, 284, 426–430. PubMed

Takaiwa, F. , Oono, K. , Iida, Y. and Sugiura, M. (1985) The complete nucleotide sequence of a rice 25S rRNA gene. Gene, 37, 255–259. PubMed

Takaiwa, F. , Oono, K. and Sugiura, M. (1984) The complete nucleotide sequence of a rice 17S rRNA gene. Nucleic Acids Res. 12, 5441–5448. PubMed PMC

Unfried, I. and Gruendler, P. (1990) Nucleotide sequence of the 5.8S and 25S rRNA genes and of the internal transcribed spacers from Arabidopsis thaliana . Nucleic Acids Res. 18, 4011. PubMed PMC

Vieira, R. , Mellosampayo, T. and Viegas, W.S. (1990) 1R chromosome nucleolus organizer region activation by 5‐azacytidine in wheat x rye hybrids. Genome, 33, 707–712.

Volkov, R.A. , Komarova, N.Y. and Hemleben, V. (2007) Ribosomal DNA in plant hybrids: inheritance, rearrangement, expression. Syst. Biodivers. 5, 261–276.

Volkov, R.A. , Kostishin, S.S. and Panchuk, I.I. (1993) rDNA organization in species from subfamily Prunoideae. Mol. Biol. 27, 1356–1367.

Volkov, R.A. , Panchuk, I.I. , Borisjuk, N.V. , Hosiawa‐Baranska, M. , Maluszynska, J. and Hemleben, V. (2017) Evolutional dynamics of 45S and 5S ribosomal DNA in ancient allohexaploid Atropa belladonna . BMC Plant Biol. 17, 21. PubMed PMC

Wallace, H. and Birnstiel, M.L. (1966) Ribosomal cistrons and the nucleolar organizer. Biochim. Biophys. Acta, 114, 296–310. PubMed

Wang, W. , Ma, L. , Becher, H. , Garcia, S. , Kovarikova, A. , Leitch, I.J. , Leitch, A.R. and Kovarik, A. (2016) Astonishing 35S rDNA diversity in the gymnosperm species Cycas revoluta Thunb. Chromosoma, 125, 683–699. PubMed PMC

Weiss‐Schneeweiss, H. , Emadzade, K. , Jang, T.S. and Schneeweiss, G.M. (2013) Evolutionary consequences, constraints and potential of polyploidy in plants. Cytogenet. Genome Res. 140, 137–150. PubMed PMC

Wendel, J.F. , Lisch, D. , Hu, G. and Mason, A.S. (2018) The long and short of doubling down: polyploidy, epigenetics, and the temporal dynamics of genome fractionation. Curr. Opin. Genet. Dev. 49, 1–7. PubMed

Wendel, J.F. , Schnabel, A. and Seelanan, T. (1995) Bidirectional interlocus concerted evolution following allopolyploid speciation in cotton (Gossypium). Proc. Natl. Acad. Sci. USA, 92, 280–284. PubMed PMC

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