, Volume 116, Issue 3, pp 275–283 | Cite as

CENH3 interacts with the centromeric retrotransposon cereba and GC-rich satellites and locates to centromeric substructures in barley

  • Andreas Houben
  • Elizabeth Schroeder-Reiter
  • Kiyotaka Nagaki
  • Shuhei Nasuda
  • Gerhard Wanner
  • Minoru Murata
  • Takashi R. Endo
Research Article


The chromosomal location of centromere-specific histone H3 (CENH3) is the assembly site for the kinetochore complex of active centromeres. Chromatin immunoprecipitation data indicated that CENH3 interacts in barley with cereba, a centromeric retroelement (CR)-like element conserved among cereal centromeres and barley-specific GC-rich centromeric satellite sequences. Anti-CENH3 signals on extended chromatin fibers always colocalized with the centromeric sequences but did not encompass the entire area covered by such centromeric repeats. This indicates that the CENH3 protein is bound only to a fraction of the centromeric repeats. At mitotic metaphase, CENH3, histone H3, and serine 10 phosphorylated histone H3 predominated within distinct structural subdomains of the centromere, as demonstrated by immunogold labeling for high resolution scanning electron microscopy.


Chromatin Fiber Sister Chromatid Cohesion Centromeric Chromatin Primary Constriction Subtelomeric Repeat 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



The anti-OsCENH3 antibody was kindly provided by Drs. P. Talbert and S. Henikoff (Howard Hughes Medical Institute, USA) and the anti-histone H3 antibody was kindly provided by Dr. D. Demidov (IPK Gatersleben, Germany). We thank Ingo Schubert (IPK, Germany) for critical reading of the manuscript. AH was supported by a visiting fellowship of the Kyoto University (Japan). The authors gratefully acknowledge Katrin Kumke and Sabine Steiner for excellent technical assistance.


  1. Amor DJ, Kalitsis P, Sumer H, Choo KHA (2004) Building the centromere: from foundation proteins to 3D organization. Trends Cell Biol 14:359–368PubMedCrossRefGoogle Scholar
  2. Belostotsky DA, Ananiev EV (1990) Characterization of Relic DNA from barley genome. Theor Appl Genet 80:374–380CrossRefGoogle Scholar
  3. Blower MD, Sullivan BA, Karpen GH (2002) Conserved organization of centromeric chromatin in flies and humans. Dev Cell 2:319–330PubMedCrossRefGoogle Scholar
  4. Cheng Z, Dong F, Langdon T, Ouyang S, Buell CR, Gu M, Blattner FR, Jiang J (2002) Functional rice centromeres are marked by a satellite repeat and a centromere-specific retrotransposon. Plant Cell 14:1691–1704PubMedCrossRefGoogle Scholar
  5. Choo KHA (2001) Domain organization at the centromere and neocentromere. Dev Cell 1:165–177PubMedCrossRefGoogle Scholar
  6. Demidov D, Van Damme D, Geelen D, Blattner FR, Houben A (2005) Identification and dynamics of two classes of aurora-like kinases in Arabidopsis and other plants. Plant Cell 17:836–848PubMedCrossRefGoogle Scholar
  7. Galbraith DW, Harkins KR, Maddox JM, Ayres NM, Sharma DP, Firoozabady E (1983) Rapid flow cytometric analysis of the cell-cycle in intact plant-tissues. Science 220:1049–1051CrossRefGoogle Scholar
  8. Gernand D, Demidov D, Houben A (2003) The temporal and spatial pattern of histone H3 phosphorylation at serine 28 and serine 10 is similar in plants but differs between mono- and polycentric chromosomes. Cytogenet Genome Res 101:172–176PubMedCrossRefGoogle Scholar
  9. Grewal SI, Moazed D (2003) Heterochromatin and epigenetic control of gene expression. Science 301:798–802PubMedCrossRefGoogle Scholar
  10. Henikoff S, Dalal Y (2005) Centromeric chromatin: what makes it unique? Curr Opin Genet Dev 15:177–184PubMedCrossRefGoogle Scholar
  11. Heun P, Erhardt, S, Blower MD, Weiss S, Skora AD, Karpen G (2006) Mislocalization of the Drosophila centromere-specific histone CID promotes formation of functional ectopic kinetochores. Dev Cell 10:303–315PubMedCrossRefGoogle Scholar
  12. Houben A, Schubert I (2003) DNA and proteins of plant centromeres. Curr Opin Plant Biol 6:554–560PubMedCrossRefGoogle Scholar
  13. Hudakova S, Michalek W, Presting GG, ten Hoopen R, dos Santos K, Jasencakova Z, Schubert I (2001) Sequence organization of barley centromeres. Nucleic Acids Res 29:5029–5035PubMedCrossRefGoogle Scholar
  14. Jin W, Melo JR, Nagaki K, Talbert PB, Henikoff S, Dawe RK, Jiang J (2004) Maize centromeres: organization and functional adaptation in the genetic background of oat. Plant Cell 16:571–581PubMedCrossRefGoogle Scholar
  15. Jin WW, Lamb JC, Vega JM, Dawe RK, Birchler JA, Jiang J (2005) Molecular and functional dissection of the maize B chromosome centromere. Plant Cell 17:1412–1423PubMedCrossRefGoogle Scholar
  16. Karpen GH, Allshire RC (1997) The case for epigenetic effects on centromere identity and function. Trends Genet 13:489–496PubMedCrossRefGoogle Scholar
  17. Kato A, Lamb JC, Birchler JA (2004) Chromosome painting using repetitive DNA sequences as probes for somatic chromosome identification in maize. Proc Natl Acad Sci USA 101:13554–13559PubMedCrossRefGoogle Scholar
  18. Kumar A, Bennetzen JL (1999) Plant retrotransposons. Annu Rev Genet 33:479–532PubMedCrossRefGoogle Scholar
  19. Lam AL, Boivin CD, Bonney CF, Rudd MK, Sullivan BA (2006) Human centromeric chromatin is a dynamic chromosomal domain that can spread over noncentromeric DNA. Proc Natl Acad Sci USA 103:4186–4191PubMedCrossRefGoogle Scholar
  20. Langdon T, Seago C, Mende M, Leggett M, Thomas H, Forster JW, Jones RN, Jenkins G (2000) Retrotransposon evolution in diverse plant genomes. Genetics 156:313–325PubMedGoogle Scholar
  21. Lermontova I, Schubert V, Fuchs J, Klatte S, Macas J, Schubert I (2006) Loading of Arabidopsis centromeric histone CENH3 occurs mainly during G2 and requires the presence of the histone fold domain. Plant Cell 18:2443–2451PubMedCrossRefGoogle Scholar
  22. Maggert KA, Karpen GH (2001) The activation of a neocentromere in Drosophila requires proximity to an endogenous centromere. Genetics 158:1615–1628PubMedGoogle Scholar
  23. Malik HS, Henikoff S (2002) Conflict begets complexity: the evolution of centromeres. Curr Opin Genet Dev 12:711–718PubMedCrossRefGoogle Scholar
  24. Manzanero S, Arana P, Puertas MJ, Houben A (2000) The chromosomal distribution of phosphorylated histone H3 differs between plants and animals at meiosis. Chromosoma 109:308–317PubMedGoogle Scholar
  25. Nagaki K, Murata M (2005) Characterization of CENH3 and centromere-associated DNA sequences in sugarcane. Chromosome Res 13:195–203PubMedCrossRefGoogle Scholar
  26. Nagaki K, Talbert PB, Zhong CX, Dawe RK, Henikoff S, Jiang J (2003a) Chromatin immunoprecipitation reveals that the 180-bp satellite repeat is the key functional DNA element of Arabidopsis thaliana centromeres. Genetics 163:1221–1225PubMedGoogle Scholar
  27. Nagaki K, Song J, Stupar RM, Parokonny AS, Yuan Q, Ouyang S, Liu J, Hsiao J, Jones KM, Dawe RK, Buell CR, Jiang J (2003b) Molecular and cytological analyses of large tracks of centromeric DNA reveal the structure and evolutionary dynamics of maize centromeres. Genetics 163:759–770PubMedGoogle Scholar
  28. Nagaki K, Cheng Z, Ouyang S, Talbert PB, Kim M, Jones KM, Henikoff S, Buell CR, Jiang J (2004) Sequencing of a rice centromere uncovers active genes. Nat Genet 36:138–145PubMedCrossRefGoogle Scholar
  29. Nasuda S, Hudakova S, Schubert I, Houben A, Endo TR (2005) Stable barley chromosomes without centromeric repeats. Proc Natl Acad Sci USA 102:9842–9847PubMedCrossRefGoogle Scholar
  30. Presting GG, Malysheva L, Fuchs J, Schubert I (1998) A Ty3/gypsy retrotransposon-like sequence localizes to the centromeric regions of cereal chromosomes. Plant J 16:721–728PubMedCrossRefGoogle Scholar
  31. Schroeder-Reiter E, Houben A, Wanner G (2003) Immunogold labeling of chromosomes for scanning electron microscopy: a closer look at phosphorylated histone H3 in mitotic metaphase chromosomes of Hordeum vulgare. Chromosome Res 11:585–596PubMedCrossRefGoogle Scholar
  32. Schubert I, Shi F, Fuchs J, Endo TR (1998) An efficient screening for terminal deletions and translocations of barley chromosomes added to common wheat. Plant J 14:489–495CrossRefGoogle Scholar
  33. Schueler MG, Higgins AW, Rudd MK, Gustashaw K, Willard HF (2001) Genomic and genetic definition of a functional human centromere. Science 294:109–115PubMedCrossRefGoogle Scholar
  34. Shi J, Dawe RK (2006) Partitioning of the maize epigenome by the number of methyl groups on histone H3 lysines 9 and 27. Genetics 173:1571–1583PubMedCrossRefGoogle Scholar
  35. Shibata F, Murata M (2004) Differential localization of the centromere-specific proteins in the major centromeric satellite of Arabidopsis thaliana. J Cell Sci 117:2963–2970PubMedCrossRefGoogle Scholar
  36. Talbert PB, Masuelli R, Tyagi AP, Comai L, Henikoff S (2002) Centromeric localization and adaptive evolution of an Arabidopsis histone H3 variant. Plant Cell 14:1053–1066PubMedCrossRefGoogle Scholar
  37. Vafa O, Sullivan KF (1997) Chromatin containing CENP-A and alpha-satellite DNA is a major component of the inner kinetochore plate. Curr Biol 7:897–900PubMedCrossRefGoogle Scholar
  38. Vig BK (1994) Do specific nucleotide bases constitute the centromere? Mutat Res 309:1–10PubMedGoogle Scholar
  39. Wanner G, Formanek H (2000) A new chromosome model. J Struct Biol 132:147–161PubMedCrossRefGoogle Scholar
  40. Warburton PE, Cooke CA, Bourassa S, Vafa O, Sullivan BA, Stetten G, Gimelli G, Warburton D, Tyler-Smith C, Sullivan KF, Poirier GG, Earnshaw WC (1997) Immunolocalization of CENP-A suggests a distinct nucleosome structure at the inner kinetochore plate of active centromeres. Curr Biol 7:901–904PubMedCrossRefGoogle Scholar
  41. White TJ, Bruns T, Lee S, Taylor J (1990) Amplification and direct sequencing of fungal ribosomal RNA genes for Phylogenetics. In: PCR protocols: a guide to methods and applications. Academic, New York, pp 315–322Google Scholar
  42. Yan H, Jin W, Nagaki K, Tian S, Ouyang S, Buell CR, Talbert PB, Henikoff S, Jiang J (2005) Transcription and histone modifications in the recombination-free region spanning a rice centromere. Plant Cell 17:3227–3238PubMedCrossRefGoogle Scholar
  43. Zhong CX, Marshall JB, Topp C, Mroczek R, Kato A, Nagaki K, Birchler JA, Jiang J, Dawe RK (2002) Centromeric retroelements and satellites interact with maize kinetochore protein CENH3. Plant Cell 14:2825–2836PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2007

Authors and Affiliations

  • Andreas Houben
    • 1
  • Elizabeth Schroeder-Reiter
    • 2
  • Kiyotaka Nagaki
    • 3
  • Shuhei Nasuda
    • 4
  • Gerhard Wanner
    • 2
  • Minoru Murata
    • 3
  • Takashi R. Endo
    • 4
  1. 1.Leibniz Institute of Plant Genetics and Crop Plant Research (IPK)GaterslebenGermany
  2. 2.Department of Biology ILudwig-Maximillians-Universität MünchenMunichGermany
  3. 3.Research Institute for BioresourcesOkayama UniversityKurashikiJapan
  4. 4.Laboratory of Plant Genetics, Graduate School of AgricultureKyoto UniversityKyotoJapan

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