Abstract
The nuclear ribosomal DNA (nrDNA) sequences are often used for phylogenetic analysis among organisms. Because DNA cytosine methylation and nucleolar dominancy are two common epigenetic mechanisms of nrDNA, we hypothesized that internal transcribed spacer 1 (ITS1), 5.8S rRNA and ITS2 of nrDNA sequences could be used as epigenetic biomarkers. Thus, this research was undertaken to study level and pattern of site-specific cytosine methylation of ITS1, 5.8S and ITS2 in nine tissues and/or developmental stage of pepper Capsicum annuum L. cultivar Demre Sivrisi. Tissues studied consisted of young and old roots at 30 and 90 days after sowing (das), mature dry seeds and seeds at 26 days of post anthesis (dpa), flowering buds at 1 day before flowering, pericarps at 3, 15 and 65 dpa. Levels and patterns of DNA cytosine methylation were identified at single base resolution using bisulfite conversion sequencing. Results of this study revealed that DNA cytosine level and pattern of ITS1, 5.8S and ITS2 were different in most tissues and/or developmental stages studied. In addition, methylation levels of CG, CHG and CHH contexts were also significantly different among the regions. Based on the findings of this study, it was concluded that high level of methylation of nrDNA sequences was relatively higher as observed in transposable element and promoter. On the other hand, its tissue-specific gene expression was effective as that of gene body and promoter methylation. Overall findings revealed that methylation levels of nrDNA could be used as biomarkers for tissue identification or age estimation in plants.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Baldwin BG, Sanderson MJ, Porter JM, Wojciechowski MF, Campbell CS, Donoghue MJ (1995) The ITS region of nuclear ribosomal DNA: a valuable source of evidence on Angiosperm phylogeny. Ann Mo Bot Gard 82:247–277. https://doi.org/10.2307/2399880
Banos S, Lentendu G, Kopf A et al (2018) A comprehensive fungi-specific 18S rRNA gene sequence primer toolkit suited for diverse research issues and sequencing platforms. BMC Microbiol 18:190. https://doi.org/10.1186/s12866-018-1331-4
Bernatavichute YV, Zhang X, Cokus S, Pellegrini M, Jacobsen SE (2008) Genome-wide association of histone h3 lysine nine methylation with CHG DNA methylation in Arabidopsis thaliana. PLoS ONE 3:e3156. https://doi.org/10.1371/journal.pone.0003156
Bhattacharyya M, De S, Chakrabarti S (2020) Origin and evolution of DNA methyltransferases (DNMT) along the tree of life: a multi-genome survey. bioRxiv. https://doi.org/10.1101/2020.04.09.033167
Bouyer D, Kramdi A, Kassam M, Heese M, Schnittger A, Roudier F, Colot V (2017) DNA methylation dynamics during early plant life. Genome Biol 18:179. https://doi.org/10.1186/s13059-017-1313-0
Chatterjee R, Vinson C (2012) CpG methylation recruits sequence specific transcription factors essential for tissue specific gene expression. Biochim Biophys Acta 1819:763–770. https://doi.org/10.1016/j.bbagrm.2012.02.014
Coleman AW (2003) ITS2 is a double-edged tool for eukaryote evolutionary comparisons. Trends Genet 19:370–375. https://doi.org/10.1016/S0168-9525(03)00118-5
Coleman AW (2015) Nuclear rRNA transcript processing versus internal transcribed spacer secondary structure. Trends Genet 31:157–163. https://doi.org/10.1016/j.tig.2015.01.002
de Assis R, Baba VY, Cintra LA, Goncalves LSA, Rodrigues R, Vanzela A (2020) Genome relationships and LTR-retrotransposon diversity in three cultivated Capsicum L. (Solanaceae) species. Res Sq. https://doi.org/10.21203/rs.2.16380/v1
Diesch J, Bywater MJ, Sanij E et al (2019) Changes in long-range rDNA-genomic interactions associate with altered RNA polymerase II gene programs during malignant transformation. Commun Biol 2:39. https://doi.org/10.1038/s42003-019-0284-y
Fan HH, Wei J, Li TC, Li ZP, Guo N, Cai YP, Lin Y (2013) DNA methylation alterations of upland cotton (Gossypium hirsutum) in response to cold stress. Acta Physiol Plant 35:2445–2453. https://doi.org/10.1007/s11738-013-1278-x
Garcia S, Khaitova CL, Kovarik A (2012) Expression of 5S rRNA genes linked to 35S rDNA in plants, their epigenetic modification and regulatory element divergence. BMC Plant Biol 12:95. https://doi.org/10.1186/1471-2229-12-95
Garcia S, Kovarik A, Leitch AR, Garnatje T (2017) Cytogenetic features of rRNA genes across land plants: analysis of the plant rDNA database. Plant J 89:1020–1030. https://doi.org/10.1111/tpj.13442
Goll MG, Bestor TH (2005) Eukaryotic cytosine methyltransferases. Annu Rev Biochem 74:481–514. https://doi.org/10.1146/annurev.biochem.74.010904.153721
Gong Z, Xue C, Zhang M, Guo R, Zhou Y, Shi G (2013) Physical localization and DNA methylation of 45S rRNA gene loci in Jatropha curcas L. PLoS ONE 8:e84284. https://doi.org/10.1371/journal.pone.0084284
Grabiele M, Debat HJ, Moscone EA, Ducasse DA (2012) 25S–18S rDNA IGS of Capsicum: molecular structure and comparison. Plant Syst Evol 298:313–321. https://doi.org/10.1007/s00606-011-0546-8
Gruntman E, Qi Y, Slotkin RK, Roeder T, Martienssen RA, Sachidanandam R (2008) Kismeth: analyzer of plant methylation states through bisulfite sequencing. BMC Bioinformatics 9:371. https://doi.org/10.1186/1471-2105-9-371
Hebestreit K, Dugas M, Klein HU (2013) Detection of significantly differentially methylated regions in targeted bisulfite sequencing data. Bioinformatics 29:1647–1653. https://doi.org/10.1093/bioinformatics/btt263
Hu N, Sui Y, Cai Y, Fan H, Yi L (2014) Effects of lanthanum on POD expression and DNA methylation of purple pepper under salt stress. J Rare Earth 32:467–475. https://doi.org/10.1016/S1002-0721(14)60095-8
Ince AG, Karaca M, Onus AN (2010a) CAPS-microsatellites: use of CAPS method to convert non-polymorphic microsatellites into useful markers. Mol Breeding 25:491–499. https://doi.org/10.1007/s11032-009-9347-y
Ince AG, Karaca M, Onus AN (2010b) Differential expression patterns of genes containing microsatellites in Capsicum annuum L. Mol Breed 25:645–658. https://doi.org/10.1007/s11032-009-9362-z
Ince AG, Yildiz F, Karaca M (2011) The MAGi DNA extraction method for fresh tissues and dry seeds. J Med Plants Res 5:5458–5464. https://academicjournals.org/journal/JMPR/article-full-text-pdf/7ECFB2716789
Ince AG, Karaca M, Elmasulu SY (2014) New microsatellite and CAPS-microsatellite markers for clarifying taxonomic and phylogenetic relationships within Origanum L. Mol Breed 34:643–654. https://doi.org/10.1007/s11032-014-0064-9
Karaca M, Ince AG, Elmasulu S, Onus AN, Turgut K (2005) Coisolation of genomic and organelle DNAs from 15 genera and 31 species of plants. Anal Biochem 343:353–355. https://doi.org/10.1016/j.ab.2005.03.021
Karaca M, Ince AG, Gocer E, Aydın A (2016) Exonic and intronic DNA methylation differences in a fiber specific gene of pima cotton (Gossypium barbadense L.). J Sci Eng Res 3:478–486. http://jsaer.com/download/vol-3-iss-4-2016/JSAER2016-03-04-478-486.pdf
Karaca M, Ince AG, Reddy UK (2020) Interspecifc grafting between Gossypium hirsutum, G. barbadense and G. herbaceum lines. Sci Rep 10:18649. https://doi.org/10.1038/s41598-020-75679-1
Kaur A, Grewal A, Sharma P (2018) Comparative analysis of DNA methylation changes in two contrasting wheat genotypes under water deficit. Biol Plant 62:471–478. https://doi.org/10.1007/s10535-018-0786-3
Kehie M, Kumaria S, Devi KS, Tandon P (2016) Genetic diversity and molecular evolution of Naga King Chili inferred from internal transcribed spacer sequence of nuclear ribosomal DNA. Meta Gene 7:56–63. https://doi.org/10.1016/j.mgene.2015.11.006
Komarova NY, Grabe T, Huigen DJ, Hemleben V, Volkov RA (2004) Organization, differential expression and methylation of rDNA in artificial Solanum allopolyploids. Plant Mol Biol 56:439–463. https://doi.org/10.1007/s11103-004-4678-x
Kwon JK, Kim BD (2009) Localization of 5S and 25S rRNA genes on somatic and meiotic chromosomes in Capsicum species of chili pepper. Mol Cells 27:205–209. https://doi.org/10.1007/s10059-009-0025-z
Law JA, Jacobsen SE (2010) Establishing, maintaining and modifying DNA methylation patterns in plants and animals. Nat Rev Genet 11:204–220. https://doi.org/10.1038/nrg2719
Matyasek R, Krumpolcova A, Lunerova J, Mikulaskova E, Rossello JA, Kovarik A (2019) Unique epigenetic features of ribosomal RNA genes (rDNA) in early diverging plants (Bryophytes). Front Plant Sci 10:1066. https://doi.org/10.3389/fpls.2019.01066
Mayer C, Schmitz KM, Li J, Grummt I, Santoro R (2006) Intergenic transcripts regulate the epigenetic state of rRNA genes. Mol Cell 22:351–361. https://doi.org/10.1016/j.molcel.2006.03.028
Ogneva ZV, Dubrovina AS, Kiselev KV (2016) Age-associated alterations in DNA methylation and expression of methyltransferase and demethylase genes in Arabidopsis thaliana. Biol Plant 60:628–634. https://doi.org/10.1007/s10535-016-0638-y
Osabe K, Clement JD, Bedon F, Pettolino FA, Ziolkowski L, Llewellyn DJ, Finnegan EJ, Wilson IW (2014) Genetic and DNA methylation changes in cotton (Gossypium) genotypes and tissues. PLoS ONE 9:e86049. https://doi.org/10.1371/journal.pone.0086049
Ostrowska-Mazurek A, Kasprzak P, Kubala S, Zaborowska M, Sobieszczuk-Nowicka E (2020) Epigenetic landmarks of leaf senescence and crop improvement. Int J Mol Sci 21:5125. https://doi.org/10.3390/ijms21145125
Purkayastha J, Alam SI, Gogoi HK, Singh L, Veer V (2012) Molecular characterization of “Bhut Jolokia” the hottest chili. J Biosci 37:757–768. https://doi.org/10.1007/s12038-012-9249-8
Ramchiary N, Kehie M, Brahma V, Kumaria S, Tandon P (2014) Application of genetics and genomics towards Capsicum translational research. Plant Biotechnol Rep 8:101–123. https://doi.org/10.1007/s11816-013-0306-z
Saez-Vasquez J, Caparros-Ruiz D, Barneche F, Echeverr M (2004) Characterization of a crucifer plant pre-rRNA processing complex. Biochem Soc Trans 32:578–580. https://doi.org/10.1042/BST0320578
Salozhin SV, Prokhorchuk EB, Georgiev GP (2005) Methylation of DNA—one of the major epigenetic markers. Biochem 70:525–532. https://doi.org/10.1007/s10541-005-0146-8
Scaldaferro MA, da Cruz MV, Cecchini NM, Moscone EA (2016) FISH and AgNor mapping of the 45S and 5S rRNA genes in wild and cultivated species of Capsicum (Solananceae). Genome 59:95–113. https://doi.org/10.1139/gen-2015-0099
Schoch CL, Seifertb KA, Huhndorf S et al (2012) Nuclear ribosomal internal transcribed spacer (ITS) region as a universal DNA barcode marker for Fungi. PNAS 109:6241–6246. https://doi.org/10.1073/pnas.1117018109
Schosserer M, Minois N, Angerer TB et al (2015) Methylation of ribosomal RNA by NSUN5 is a conserved mechanism modulating organismal lifespan. Nat Commun 6:6158. https://doi.org/10.1038/ncomms7158
Sekhon RS, Peterson T, Chopra S (2007) Epigenetic modifications of distinct sequences of the p1 regulatory gene specify tissue-specific expression patterns in maize. Genetics 175:1059–1070. https://doi.org/10.1534/genetics.106.066134
Shayevitch R, Askayo D, Keydar I, Ast G (2018) The importance of DNA methylation of exons on alternative splicing. RNA 24:1351–1362. https://www.rnajournal.org/cgi/doi/10.1261/rna.064865.117
Sochorova J, Garcia S, Galvez F et al (2018) Evolutionary trends in animal ribosomal DNA loci: introduction to a new online database. Chromosoma 127:141–150. https://doi.org/10.1007/s00412-017-0651-8
Song Q, Chen ZJ (2015) Epigenetic and developmental regulation in plant polyploids. Curr Opin Plant Biol 24:101–109. https://doi.org/10.1016/j.pbi.2015.02.007
Sun YL, Choi IL, Lee YB, Choi KY, Hong SK, Kang HM (2014) Molecular diversity and phylogentic analysis of Capsicum annuum varieties using the nrDNA ITS region. Sci Hortic 165:336–343. https://doi.org/10.1016/j.scienta.2013.11.009
Thomas H (2013) Senescence, ageing and death of the whole plant. New Phytol 197:696–711. https://doi.org/10.1111/nph.12047
To TK, Saze H, Kakutani T (2015) DNA Methylation within transcribed regions. Plant Physiol 168:1219–1225. https://doi.org/10.1104/pp.15.00543
Vining KJ, Pomraning KR, Wilhelm LJ, Priest HD, Pellegrini M, Mockler TC, Freitag M, Strauss SH (2012) Dynamic DNA cytosine methylation in the Populus trichocarpa genome: tissue-level variation and relationship to gene expression. BMC Genomics 13:27. https://doi.org/10.1186/1471-2164-13-27
Wang M, Lemos B (2020) Ribosomal DNA harbors an evolutionarily conserved clock of biological aging. Genome Res 29:325–333. https://doi.org/10.1101/gr.241745.118
Wendte JM, Zhang Y, Ji L, Shi X, Hazarika RR, Shahryary Y, Johannes F, Schmitz RJ (2019) Epimutations are associated with CHROMOMETHYLASE 3-induced de novo DNA methylation. Elife 8:e47891. https://doi.org/10.7554/eLife.47891
Wieckowski Y, Schiefelbein J (2012) Nuclear ribosome biogenesis mediated by the DIM1A rRNA dimethylase is required for organized root growth and epidermal patterning in Arabidopsis. Plant Cell 24:2839–2856. https://doi.org/10.1105/tpc.112.101022
Xiao K, Chen J, He Q, Wang Y, Shen H, Sun L (2020) DNA methylation is involved in the regulation of pepper fruit ripening and interacts with phytohormones. J Exp Bot 71:1928–1942. https://doi.org/10.1093/jxb/eraa003
Xu XW, Li T, Li Y, Wang HM (2015) Variation of DNA cytosine methylation patterns among parent lines and reciprocal hybrids in hot pepper. Chem Eng Trans 46:1345–1350. https://doi.org/10.3303/CET1546225
Yong W-S, Hsu F-M, Chen P-Y (2016) Profiling genome-wide DNA methylation. Epigenetics Chromatin 9:26. https://doi.org/10.1186/s13072-016-0075-3
Youn-Kyu P, Kim B-D, Kim B-S, Armstrong KC, Kim N-S (1999) Karyotyping of the chromosomes and physical mapping of the 5S rRNA and 18S–26S rRNA gene families in five different species in Capsicum. Genes Genet Syst 74:149–157. https://doi.org/10.1266/ggs.74.149
Yousefzadeh H, Colagar A, Tabari M, Sattarian A, Assadi M (2012) Utility of ITS region sequence and structure for molecular identification of Tilia species from Hyrcanian forestsm Iran. Plant Syst Evol 298:947–961. https://doi.org/10.1007/s00606-012-0604-x
Zakrzewski F, Schubert V, Viehoever P, Minoche AE, Dohm JC, Himmelbauer H, Weisshaar B, Schmidt T (2014) The CHH motif in sugar beet satellite DNA: a modulator for cytosine methylation. Plant J 78:937–995. https://doi.org/10.1111/tpj.12519
Zhang X, Fang H, Zhang W, Zhong B, Li Y, Wang X (2019) Ribosomal DNA methylation as stable biomarkers for detection of cancer in plasma. bioRxiv. https://doi.org/10.1101/651497
Zhou HC, Waminal N, Kim HH (2019) In silico mining and FISH mapping of a chromosome-specific satellite DNA in Capsicum annuum L. Genes Genomics 41:1001–1006. https://doi.org/10.1007/s13258-019-00832-8
Funding
This study was financially supported in part by a grant from Scientific and Technological Research Council of Turkey (Project No: 113O935) and the Scientific Research Projects Coordination Unit of Akdeniz University.
Author information
Authors and Affiliations
Contributions
All authors contributed to the study conception and design. Material preparation, data collection and analysis were performed by AGI and MK. Two authors read and approved the final manuscript.
Corresponding author
Ethics declarations
Conflict of interest
The authors have no conflicts of interest to declare that are relevant to the content of this article.
Additional information
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Rights and permissions
About this article
Cite this article
Ince, A.G., Karaca, M. Tissue and/or developmental stage specific methylation of nrDNA in Capsicum annuum. J Plant Res 134, 841–855 (2021). https://doi.org/10.1007/s10265-021-01287-3
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10265-021-01287-3