DOI QR코드

DOI QR Code

Variation in the number of nucleoli and incomplete homogenization of 18S ribosomal DNA sequences in leaf cells of the cultivated Oriental ginseng (Panax ginseng Meyer)

  • Chelomina, Galina N. (Institute of Biology and Soil Science, Far Eastern Branch of the Russian Academy of Sciences) ;
  • Rozhkovan, Konstantin V. (Institute of Biology and Soil Science, Far Eastern Branch of the Russian Academy of Sciences) ;
  • Voronova, Anastasia N. (Institute of Biology and Soil Science, Far Eastern Branch of the Russian Academy of Sciences) ;
  • Burundukova, Olga L. (Institute of Biology and Soil Science, Far Eastern Branch of the Russian Academy of Sciences) ;
  • Muzarok, Tamara I. (Institute of Biology and Soil Science, Far Eastern Branch of the Russian Academy of Sciences) ;
  • Zhuravlev, Yuri N. (Institute of Biology and Soil Science, Far Eastern Branch of the Russian Academy of Sciences)
  • Received : 2015.02.06
  • Accepted : 2015.07.14
  • Published : 2016.04.15

Abstract

Background: Wild ginseng, Panax ginseng Meyer, is an endangered species of medicinal plants. In the present study, we analyzed variations within the ribosomal DNA (rDNA) cluster to gain insight into the genetic diversity of the Oriental ginseng, P. ginseng, at artificial plant cultivation. Methods: The roots of wild P. ginseng plants were sampled from a nonprotected natural population of the Russian Far East. The slides were prepared from leaf tissues using the squash technique for cytogenetic analysis. The 18S rDNA sequences were cloned and sequenced. The distribution of nucleotide diversity, recombination events, and interspecific phylogenies for the total 18S rDNA sequence data set was also examined. Results: In mesophyll cells, mononucleolar nuclei were estimated to be dominant (75.7%), while the remaining nuclei contained two to four nucleoli. Among the analyzed 18S rDNA clones, 20% were identical to the 18S rDNA sequence of P. ginseng from Japan, and other clones differed in one to six substitutions. The nucleotide polymorphism was more expressed at the positions 440-640 bp, and distributed in variable regions, expansion segments, and conservative elements of core structure. The phylogenetic analysis confirmed conspecificity of ginseng plants cultivated in different regions, with two fixed mutations between P. ginseng and other species. Conclusion: This study identified the evidences of the intragenomic nucleotide polymorphism in the 18S rDNA sequences of P. ginseng. These data suggest that, in cultivated plants, the observed genome instability may influence the synthesis of biologically active compounds, which are widely used in traditional medicine.

Keywords

References

  1. Zhuravlev YN, Kolyada AS. Araliaceae: ginseng and others. Vladivostok: Dalnauka; 1996 [in Russian].
  2. Arnheim N. Concerted evolution in multigene families. In: Nei M, Koehn R, editors. Evolution of genes and proteins. Sunderland: Sinauer; 1983. p. 38-61.
  3. Brown DD, Wensink PC, Jordan E. A comparison of the ribosomal DNA's of Xenopus laevis and Xenopus mulleri: the evolution of tandem genes. J Mol Biol 1972;63:57-73. https://doi.org/10.1016/0022-2836(72)90521-9
  4. Dover GA. Concerted evolution, molecular drive and natural selection. Curr Biol 1994;4:1165-6. https://doi.org/10.1016/S0960-9822(00)00265-7
  5. Nei M, Rooney AP. Concerted and birth-and-death evolution of multigene families. Annu Rev Genet 2005;39:121-52. https://doi.org/10.1146/annurev.genet.39.073003.112240
  6. Ohta T. Evolution of gene families. Gene 2000;259:45-52. https://doi.org/10.1016/S0378-1119(00)00428-5
  7. Ohta T. The mutational load of a multigene family with uniform members. Genet Res 1989;53:141-5. https://doi.org/10.1017/S0016672300028020
  8. Fenton B, Malloch G, Germa F. A study of variation in rDNA ITS regions shows that two haplotypes coexist within a single aphid genome. Genome 1998;41:337-45. https://doi.org/10.1139/g98-030
  9. Harris DJ, Crandall KA. Intra-genomic variation within ITS1 and ITS2 of freshwater crayfishes (Decapoda: Cambaridae): implications for phylogenetic and microsatellite studies. Mol Biol Evol 2000;17:284-91. https://doi.org/10.1093/oxfordjournals.molbev.a026308
  10. Parkin EJ, Butlin RK. Within- and between-individual sequence variation among ITS1 copies in the meadow grasshopper Chorthippus parallelus indicates frequent intrachromosomal gene conversion. Mol Biol Evol 2004;27:1595-601.
  11. Xu H, Wang Z, Ding X, Zhou K, Xu L. Differentiation of Dendrobium species used as “Huangcao Shihu” by rDNA ITS sequence analysis. Planta Med 2005;71:1-3.
  12. Matyasek R, Renny-Byfield S, Fulnecek J, Macas J, Grandbastien M-A, Nichols R, Leitch A, Kovarik A. Next generation sequencing analysis reveals a relationship between rDNA unit diversity and locus number in Nicotiana diploids. BMC Genomics 2012;13:722. https://doi.org/10.1186/1471-2164-13-722
  13. Kovarik A, Dadejova M, Lim YK, Chase MW, Clarkson JJ, Knapp S, Leitch AR. Evolution of rDNA in Nicotiana allopolyploids: a potential link between rDNA homogenization and epigenetics. Ann Bot 2008;101:815-23. https://doi.org/10.1093/aob/mcn019
  14. Keller I, Chintauan-Marquier IC, Veltsos P, Nichols RA. Ribosomal DNA in the grasshopper Podisma pedestris: escape from concerted evolution. Genetics 2006;174:863-74. https://doi.org/10.1534/genetics.106.061341
  15. Marquez LM, Miller DJ, Mackenzie JB, Van Oppen MJM. Pseudogenes contribute to extreme diversity of nuclear ribosomal DNA in the hard coral Acropora. Mol Biol Evol 2003;20:1077-86. https://doi.org/10.1093/molbev/msg122
  16. Muir G, Fleming CC, Schlotterer C. Three divergent rDNA clusters predate the species divergence in Quercus petraea (Matt.) Liebl. and Quercus robur L. Mol Biol Evol 2001;18:112-9. https://doi.org/10.1093/oxfordjournals.molbev.a003785
  17. Ruggiero MV, Procaccini G. The rDNA ITS region in the lessepsian marine angiosperm Halophila stipulacea (Forssk.) Aschers. (Hydrocharitaceae): intragenomic variability and putative pseudogenic sequences. J Mol Evol 2004;58:115-21. https://doi.org/10.1007/s00239-003-2536-0
  18. Campbell CS, Wojciechowski MF, Baldwin BG, Alice LA, Donoghue MJ. Persistent nuclear ribosomal DNA sequence polymorphism in the Amelanchier agamic complex (Rosaceae). Mol Biol Evol 1997;14:81-90. https://doi.org/10.1093/oxfordjournals.molbev.a025705
  19. Crease TJ, Lynch M. Ribosomal DNA variation in Daphnia pulex. Genetics 1991;141:1327-37.
  20. Krieger J, Fuerst PA. Characterization of nuclear 18S rRNA gene sequence diversity and expression in an individual lake sturgeon (Acipenser fulvescens). J Appl Ichthyol 2004;20:433-9. https://doi.org/10.1111/j.1439-0426.2004.00610.x
  21. Liu ZL, Zhang D, Wang ZQ, Ma XF, Wang XR. Intragenomic and interspecific 5S rDNA sequence variation in five Asian pines. Am J Bot 2003;90:17-24. https://doi.org/10.3732/ajb.90.1.17
  22. Pillet L, Fontaine D, Pawlowski J. Intra-genomic ribosomal RNA polymorphism and morphological variation in Elphidium macellum suggests inter-specific hybridization in foraminifera. PLoS ONE 2012;7:e32373. https://doi.org/10.1371/journal.pone.0032373
  23. Robles F, de la Herran R, Ludwig A, Ruiz-Rejon C, Ruiz-Rejon M, Garrido-Ramos M. Genomic organization and evolution of the 5S ribosomal DNA in the ancient fish sturgeon. Genome 2005;48:18-28. https://doi.org/10.1139/g04-077
  24. Wei N-WV, Wallace CC, Dai C-F, Pillay KRM, Chen CA. Analyses of the ribosomal internal transcribed spacers (ITS) and the 5.8S gene indicate that extremely high rDNA heterogeneity is a unique feature in the Scleractinian coral genus Acropora (Scleractinia; Acroporidae). Zool Stud 2006;45:404-18.
  25. Wendel JF. Genome evolution in polyploids. Plant Mol Biol 2000;42:225-49. https://doi.org/10.1023/A:1006392424384
  26. Bartova E, Harnicarova Horakova A, Uhlirova R, Raska I, Galiova G, Orlova D, Kozubek S. Structure and epigenetics of nucleoli in comparison with nonnucleolar compartments. J Histochem Cytochem 2010;58:391-403. https://doi.org/10.1369/jhc.2009.955435
  27. Catalano A, O'Day DH. Rad53 homologue forkhead-associated kinase A (FhkA) and $Ca^{2+}$-binding protein 4a (CBP4a) are nucleolar proteins that differentially redistribute during mitosis in Dictyostelium. Cell Div 2013;8:4. https://doi.org/10.1186/1747-1028-8-4
  28. Kobayashi T, Heck DJ, Nomura M, Horiuchi T. Expansion and contraction of ribosomal DNA repeats in Saccharomyces cerevisiae: requirement of replication fork blocking (Fob1) protein and the role of RNA polymerase I. Genes Dev 1998;12:3821-30. https://doi.org/10.1101/gad.12.24.3821
  29. Kobayashi T. A new role of the rDNA and nucleolus in the nucleus rDNA instability maintains genome integrity. Bioessays 2008;30:267-72. https://doi.org/10.1002/bies.20723
  30. Kobayashi T. Regulation of ribosomal RNA gene copy number and its role in modulating genome integrity and evolutionary adaptability in yeast. Cell Mol Life Sci 2011;68:1395-403. https://doi.org/10.1007/s00018-010-0613-2
  31. Olausson KH, Nister M, Londsrom MS. p53-Dependent and -independent nucleolar stress responses. Cell 2012;1:774-98. https://doi.org/10.3390/cells1040774
  32. Show P, Brown J. Nucleoli: composition, function, and dynamics. Plant Physiol 2012;158:44-51. https://doi.org/10.1104/pp.111.188052
  33. Khrolenko YA, Burundukova OL, Lauve LS, Muzarok TI, Makhan'kov VV, Zhuravlev YN. Characterization of the variability of nucleoli in the cells of Panax ginseng Meyer in vivo and in vitro. J Ginseng Res 2012;36:322-6. https://doi.org/10.5142/jgr.2012.36.3.322
  34. Muratova EN. Nucleolus staining methods for karyotype analysis of conifers. Bot J 1995;80:82-6.
  35. Kiselev KV, Bulgakov VP. Stability of the rolC gene and its expression in 15-year-old cell cultures of Panax ginseng. Appl Biochem Microbiol 2009;45:252-8.
  36. Fushimi H, Komatsu K, Isobe M, Namba T. 18S ribosomal RNA gene sequences of three Panax species and the corresponding ginseng drugs. Biol Pharm Bull 1996;19:1530-2. https://doi.org/10.1248/bpb.19.1530
  37. Krieger J, Hett AK, Fuerst PA, Birstein VJ, Ludwig A. Unusual intraindividual variation of the nuclear 18S rRNA gene is widespread within the Acipenseridae. J Hered 2006;97:218-25. https://doi.org/10.1093/jhered/esj035
  38. Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. MEGA 5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 2011;28:2731-9. https://doi.org/10.1093/molbev/msr121
  39. Librado P, Rozas J. DnaSP v5: a software for comprehensive analysis of DNA polymorphism data. Bioinformatics 2009;25:1451-2. https://doi.org/10.1093/bioinformatics/btp187
  40. Martin DP, Lemey P, Lott M, Moulton V, Posada D, Lefeuvre P. RDP3: a flexible and fast computer program for analyzing recombination. Bioinformatics 2010;26:2462-3. https://doi.org/10.1093/bioinformatics/btq467
  41. Posada D, Crandall KA. MODELTEST: testing the model of DNA substitution. Bioinformatics 1998;14:817-8. https://doi.org/10.1093/bioinformatics/14.9.817
  42. Armache JP, Jarasch A, Anger AM, Villa E, Becker T, Bhushan S, Jossinet F, Habeck M, Dindar G, Franckenberg S, et al. Cryo-EM structure and rRNA model of a translating eukaryotic 80S ribosome at 5.5-A resolution. Proc Natl Acad Sci USA 2010;107:19748-53. https://doi.org/10.1073/pnas.1009999107
  43. Krieger J, Fuerst PA. Evidence of multiple alleles of the nuclear 18S ribosomal RNA gene in sturgeon. Appl Ichthyol 2002;18:290-7. https://doi.org/10.1046/j.1439-0426.2002.00377.x
  44. Koren OG, Potenko VV, Zhuravlev YN. Inheritance and variation of allozymes in Panax ginseng C.A. Meyer (Araliaceae). Int J Plant Sci 2003;164:189-95. https://doi.org/10.1086/344758
  45. Kozyrenko MM, Artyukova EV, Lauve LS, Zhuravlev YN, Reunova GD. The genetic variability of Panax ginseng callus lines. BioTechnologia 2001;1:19-26.
  46. Zhuravlev YN, Reunova GD, Kozyrenko MM, Artyukova EV, Muzarok TI. Genetic variation of wild ginseng populations (RAPD analysis). Mol Biol 1998;32:910-4 [in Russian].
  47. Komatsu K, Zhu S, Fushimi H, Qui TK, Cai S, Kadota S. Phylogenetic analysis based on 18S rRNA gene and matK gene sequences of Panax vietnamensis and five related species. Planta Med 2001;67:461-5. https://doi.org/10.1055/s-2001-15821
  48. Grushvitskii IV. Ginseng: the aspects of biology. Leningrad: Nauka; 1961.
  49. Gurzenkov NN, Kolyada AS. Study of the karyotype of Panax ginseng C.A. Meyer (Araliaceae). In: Gursenkov NN, Moskaliuk TA, Chernyshev VD, editors. Biological researches of the Gornotaezhnaya station. Ussuriysk: Rossiiskaia akademiia nauk; 1996. p. 101-5.
  50. Raven PH. The bases of angiosperm phylogeny: cytology. Ann Missouri Bot Gard 1975;62:724-64. https://doi.org/10.2307/2395272
  51. Yi T, Lowry PP, Plunkett GM. Chromosomal evolution in Araliaceae and close relatives. Taxon 2004;19:987-1005.
  52. Bulgakov VP, Lauve LS, Tchernoded GK, Khodakovskaya MV, Zhuravlev YN. Chromosome variation in ginseng cells transformed with the rolC plant oncogene. Russ J Genet 2000;36:150-6.
  53. Kunakh VA. Genome variation of plant somatic cells, II: variation in nature. Biopolim Kletka 1995;11:5-40.
  54. Lauve LS, Burundukova OL, Muzarok TI, Zhuravlev YN. Chromosome numbers of Panax ginseng (Araliaceae). Bot J 2008;93:158-61.
  55. Kunakh VA, Mozhilevskaya LP, Adonin VI, Gubar SI. Productivity and genetic structure of Panax ginseng C.A. Meyer cell populations during the in vitro cultivation. BioTechnologia 2003;3:25-35.
  56. Dorofeev VY, Karnachuk RA, Pulkina SV, Komleva EV, Dubina VB, Medvedeva JV. Atragene speciosa Weinm culture in vitro: the cytogenetic analysis and formation of triterpenoid glycosides and flavonoids. Vestnik Tomskogo Univ 2009;3:37-41.
  57. Shakhbazov VG, Shestopalova NG. Some peculiarities of nucleoli in cells of onion seed. Dokl USSR 1971;196:58-64.
  58. Korshikov II , Tkachev YA. The nucleusenucleolus characteristics of seed progeny for Crimean pine (Pinus pallasiana D. Don) from wild population and planting of anthropogenic contaminated territories. Ind Bot 2011;11:157-61.
  59. Burundukova OL, Ivanov LA, Ivanova LA, Kiselev KV, Makhan'kov VV, Lauve LS, Khrolenko YA, Burkovskaya EV, Velivetskaya TA, Ignatiev AV, et al. Morphofunctional principles determining the changes in the adaptation strategy of ginseng (Panax ginseng C.A. Meyer) during its domestication. Dokl Academii Nauk 2012;446:584-97.
  60. Ngezahayo F, Wang XL, Yu XM, Jiang LL, Chu YJ, Shen BH, Yan ZK, Liu B. Habitat-induced reciprocal transformation in the root phenotype of Oriental ginseng is associated with alteration in DNA methylation. Chinese Sci Bull 2011;56:1685-90. https://doi.org/10.1007/s11434-011-4484-1
  61. Alvenson AJ, Kolnick L. Intragenomic nucleotide polymorphism among small subunit (18S) rDNA paralogs in the diatom genus Skeletonema (Bacillariophyta). J Phycol 2005;41:1248-57. https://doi.org/10.1111/j.1529-8817.2005.00136.x
  62. Chelomina GN, Rozhkovan KV, Ivanov SA, Bulgakov VP. Multiplicity of alleles of nuclear 18S rRNA gene of Amur sturgeons: genes and pseudogenes? Dokl Biochem Biophys 2008;420:115-8. https://doi.org/10.1134/S1607672908030058
  63. Chelomina GN, Rozhkovan KV, Rachek EI, Zhuravlev YN. Increased genetic diversity of 18S rDNA in genomes of $F_1$ hybrids of sturgeons Acipenser schrenckii $\times$ A. baerii and A. schrenckii $\times$ Huso dauricus. Dokl Biol Sci 2008;421:271-4. https://doi.org/10.1134/S0012496608040157
  64. Rittie L, Perbal B. Enzymes used in molecular biology: a useful guide. J Cell Commun Signal 2008;2:25-45. https://doi.org/10.1007/s12079-008-0026-2
  65. Lodish H, Berk A, Zipursky SL, Matsudaira P, Baltimore D, Darnell J. DNA replication, repair, and recombination. Molecular cell biology. New York: W.H. Freeman and Company; 1999. p. 454-81.
  66. Cronn R, Cedroni M, Haselkorn T, Grover C, Wendel JF. PCR-mediated recombination in amplification products derived from polyploid cotton. Theor Appl Genet 2002;104:482-9. https://doi.org/10.1007/s001220100741

Cited by

  1. Concerted evolution rapidly eliminates sequence variation in rDNA coding regions but not in intergenic spacers in Nicotiana tabacum allotetraploid vol.303, pp.8, 2017, https://doi.org/10.1007/s00606-017-1442-7
  2. Nucleolar Dominance in a Tetraploidy Hybrid Lineage Derived From Carassius auratus red var. ( [GRAPHIC OMISSION] ) × Megalobrama amblycephala ( [GRAPHIC OMISSION] ) vol.9, pp.None, 2016, https://doi.org/10.3389/fgene.2018.00386
  3. Protein metabolic changes and nucleolus organizer regions activity in the lymphocytes of neonatal calves during the development of respiratory diseases vol.12, pp.10, 2019, https://doi.org/10.14202/vetworld.2019.1657-1667
  4. Age-Dependent and Tissue-Specific Alterations in the rDNA Clusters of the Panax ginseng C. A. Meyer Cultivated Cell Lines vol.10, pp.10, 2020, https://doi.org/10.3390/biom10101410