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Characterization of the two centromeric proteins CENP-C and MIS12 in Nicotiana species

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Abstract

Centromeres play an important role in chromosome transmission in eukaryotes and comprise specific DNA and proteins that form complexes called kinetochores. In tobacco, although a centromere-specific histone H3 (NtCENH3) and centromeric DNA sequence (Nt2-7) have been identified, no other kinetochore components have been determined. In this study, we isolated and characterized cDNAs encoding two centromeric proteins CENP-C and MIS12 from Nicotiana tabaccum. Two CENP-C homologues, NtCENP-C-1 and -2, isolated from N. tabaccum were similar to CENP-C from N. sylvestris and N. tomentosiformis, respectively. Similarly, two Mis12 homologues, NtMIS12-1 and -2, in N. tabaccum were shown to originate from N. sylvestris and N. tomentosiformis, respectively. Both respective homologues for CENP-C and Mis12 were expressed at the same level. This indicates that in a tetraploid species, N. tabaccum, two ancestral genes encoding the centromeric proteins participate equally in the functioning of centromeres.

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Abbreviations

AtCENP-C:

centromeric protein C of Arabidopsis thaliana

CAPS:

cleaved amplified polymorphic DNA sequence

DAPI:

4,6-diamino-2-phenylindole

EST:

expressed sequence tag

GFP:

green fluorescent protein

MIS12:

mini chromosome instability mutant 12

ORF:

open reading frame

PBS:

phosphate-buffered saline

PCR:

polymerase chain reaction

PHEMES:

PIPES (Piperazine-1,4-bis (2-ethanesulfonic acid)), HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid), EGTA (ethylene glycol tetraacetic acid), MgCl2, sorbitol

RACE:

rapid amplification of cDNA ends

RT-PCR:

reverse transcription polymerase chain reaction

T-DNA:

transfer DNA

References

  • Amor DJ, Kalitsis P, Sumer H, Choo KH (2004) Building the centromere: from foundation proteins to 3D organization. Trends Cell Biol 14:359–368

    Article  PubMed  CAS  Google Scholar 

  • Dawe RK, Reed LM, Yu HG, Muszynski MG, Hiatt EN (1999) A maize homolog of mammalian CENPC is a constitutive component of the inner kinetochore. Plant Cell 11:1227–1238

    Article  PubMed  CAS  Google Scholar 

  • Earnshaw WC, Rothfield N (1985) Identification of a family of human centromere proteins using autoimmune sera from patients with scleroderma. Chromosoma 91:313–321

    Article  PubMed  CAS  Google Scholar 

  • Fukagawa T, Brown WRA (1997) Efficient conditional mutation of the vertebrate CENP-C gene. Hum Mol Genet 6:2301–2308

    Article  PubMed  CAS  Google Scholar 

  • Fukagawa T, Mikami Y, Nishihashi A et al (2001) CENP-H, a constitutive centromere component, is required for centromere targeting of CENP-C in vertebrate cells. EMBO J 20:4603–4617

    Article  PubMed  CAS  Google Scholar 

  • Goshima G, Saitoh S, Yanagida M (1999) Proper metaphase spindle length is determined by centromere proteins Mis12 and Mis6 required for faithful chromosome segregation. Genes Dev 13:1664–1677

    Article  PubMed  CAS  Google Scholar 

  • Houben A, Schroeder-Reiter E, Nagaki K et al (2007) CENH3 interacts with the centromeric retrotransposon cereba and GC-rich satellites and locates to centromeric substructures in barley. Chromosoma 116:275–283

    Article  PubMed  CAS  Google Scholar 

  • Kalitsis P, Fowler KJ, Earle E, Hill J, Choo KH (1998) Targeted disruption of mouse centromere protein C gene leads to mitotic disarray and early embryo death. Proc Natl Acad Sci U S A 95:1136–1141

    Article  PubMed  CAS  Google Scholar 

  • Karimi M, Inze D, Depicker A (2002) Gateway vectors for Agrobacterium-mediated plant transformation. Trends Plant Sci 7:193–195

    Article  PubMed  CAS  Google Scholar 

  • Lee H, Zhang W, Langdon T et al (2005) Chromatin immunoprecipitation cloning reveals rapid evolutionary patterns of centromeric DNA in Oryza species. Proc Natl Acad Sci U S A 102:11793–11798

    Article  PubMed  CAS  Google Scholar 

  • Mathur J, Szabados L, Schaefer S et al (1998) Gene identification with sequenced T-DNA tags generated by transformation of Arabidopsis cell suspension. Plant J 13:707–716

    Article  PubMed  CAS  Google Scholar 

  • Nagaki K, Murata M (2005) Characterization of CENH3 and centromere-associated DNA sequences in sugarcane. Chromosome Res 13:195–203

    Article  PubMed  CAS  Google Scholar 

  • Nagaki K, Talbert PB, Zhong CX, Dawe RK, Henikoff S, Jiang J (2003) Chromatin Immunoprecipitation reveals that the 180-bp satellite repeat is the key functional DNA element of Arabidopsis thaliana centromeres. Genetics 163:1221–1225

    PubMed  CAS  Google Scholar 

  • Nagaki K, Cheng Z, Ouyang S et al (2004) Sequencing of a rice centromere uncovers active genes. Nat Genet 36:138–145

    Article  PubMed  CAS  Google Scholar 

  • Nagaki K, Kashihara K, Murata M (2009) A centromeric DNA sequence colocalized with a centromere-specific histone H3 in tobacco. Chromosoma 118:249–257

    Article  PubMed  CAS  Google Scholar 

  • Ogura Y, Shibata F, Sato H, Murata M (2004) Characterization of a CENP-C homolog in Arabidopsis thaliana. Genes Genet Syst 79:139–144

    Article  PubMed  CAS  Google Scholar 

  • Politi V, Perini G, Trazzi S, Pliss A, Raska I, Earnshaw WC (2002) CENP-C binds the alpha-satellite DNA in vivo at specific centromere domains. J Cell Sci 115:2317–2327

    PubMed  CAS  Google Scholar 

  • Sato H, Shibata F, Murata M (2005) Characterization of a Mis12 homologue in Arabidopsis thaliana. Chromosome Res 13:827–834

    Article  PubMed  CAS  Google Scholar 

  • Talbert PB, Bryson TD, Henikoff S (2004) Adaptive evolution of centromere proteins in plants and animals. J Biol 3:18

    Article  PubMed  Google Scholar 

  • Talbert PB, Masuelli R, Tyagi AP, Comai L, Henikoff S (2002) Centromeric localization and adaptive evolution of an Arabidopsis histone H3 variant. Plant Cell 14:1053–1066

    Article  PubMed  CAS  Google Scholar 

  • Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Res 25:4876–4882

    Article  PubMed  CAS  Google Scholar 

  • Zhong CX, Marshall JB, Topp C et al (2002) Centromeric retroelements and satellites interact with maize kinetochore protein CENH3. Plant Cell 14:2825–2836

    Article  PubMed  CAS  Google Scholar 

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Acknowledgements

This work was supported by a grant from the Ministry of Education, Culture, Sports, Science and Technology in Japan (#18770007). Seeds of N. sylvestris and N. tomentosiformis were kindly provided by Japan Tobacco Inc.

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Authors and Affiliations

  1. Research Institute for Bioresources, Okayama University, Kurashiki, Japan

    Kiyotaka Nagaki, Kazunari Kashihara & Minoru Murata

Authors
  1. Kiyotaka Nagaki
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  2. Kazunari Kashihara
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  3. Minoru Murata
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Corresponding author

Correspondence to Kiyotaka Nagaki.

Additional information

Responsible Editor: J. S. (Pat) Heslop-Harrison

Electronic supplementary material

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Supplementary Figure S1

Alignment of CENP-C cDNAs from Nicotiana species. CENP-C cDNAs isolated in this study (NtCENP-C-1 and -2, NsCENP-C, and NtoCENP-C) were aligned. Start and stop codons are marked by red and blue boxes, respectively. Primer positions and the SalI site used for CAPS analyses are indicated by arrows and a green box, respectively (GIFF 319 KB)

High resolution image (TIFF 17.4 MB)

Supplementary Figure S2

Alignment of CENP-Cs from Solanaceae species and A. thaliana. CENP-Cs identified in this study (NtCENP-C-1 and -2, NsCENP-C, and NtoCENP-C) were aligned with AtCENP-C (A. thaliana), SlCENP-C (Solanum lycopersicum), and StCENP-C (S. tuberosum). A red box indicates the peptide sequence used to raise the anti-NtCENP-C antibody. N- and C-terminal conserved regions are marked by green and blue boxes, respectively. The CENP-C box is underlined in purple (GIFF 267 KB)

High resolution image (TIFF 13.1 MB)

Supplementary Figure S3

Alignment of MIS12 cDNAs from Nicotiana species. MIS12 cDNAs isolated in this study (NtMIS12-1 and -2, NsMIS12, and NtoMIS12) were aligned. Start and stop codons are marked by red and blue boxes, respectively. Primer positions and the EcoRI site used for CAPS analyses are indicated by arrows and a green box, respectively (GIFF 299 KB)

High resolution image (TIFF 16.7 MB)

Supplementary Figure S4

Alignment of MIS12s from Nicotiana species and A. thaliana. MIS12s identified in this study (NtMIS12-1 and -2, NsMIS12-C, and NtoMIS12-C) were aligned with AtMIS12 (A. thaliana). Blocks 1 and 2 are marked by blue boxes. A green line indicates the coiled-coil motif (GIFF 158 KB)

High resolution image (TIFF 9.18 MB)

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Nagaki, K., Kashihara, K. & Murata, M. Characterization of the two centromeric proteins CENP-C and MIS12 in Nicotiana species. Chromosome Res 17, 719–726 (2009). https://doi.org/10.1007/s10577-009-9064-8

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  • DOI: https://doi.org/10.1007/s10577-009-9064-8

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