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Variation of 45S rDNA intergenic spacers in Arabidopsis thaliana

Abstract

Approximately seven hundred 45S rRNA genes (rDNA) in the Arabidopsis thaliana genome are organised in two 4 Mbp-long arrays of tandem repeats arranged in head-to-tail fashion separated by an intergenic spacer (IGS). These arrays make up 5 % of the A. thaliana genome. IGS are rapidly evolving sequences and frequent rearrangements inside the rDNA loci have generated considerable interspecific and even intra-individual variability which allows to distinguish among otherwise highly conserved rRNA genes. The IGS has not been comprehensively described despite its potential importance in regulation of rDNA transcription and replication. Here we describe the detailed sequence variation in the complete IGS of A. thaliana WT plants and provide the reference/consensus IGS sequence, as well as genomic DNA analysis. We further investigate mutants dysfunctional in chromatin assembly factor-1 (CAF-1) (fas1 and fas2 mutants), which are known to have a reduced number of rDNA copies, and plant lines with restored CAF-1 function (segregated from a fas1xfas2 genetic background) showing major rDNA rearrangements. The systematic rDNA loss in CAF-1 mutants leads to the decreased variability of the IGS and to the occurrence of distinct IGS variants. We present for the first time a comprehensive and representative set of complete IGS sequences, obtained by conventional cloning and by Pacific Biosciences sequencing. Our data expands the knowledge of the A. thaliana IGS sequence arrangement and variability, which has not been available in full and in detail until now. This is also the first study combining IGS sequencing data with RFLP analysis of genomic DNA.

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Acknowledgments

We thank Veronika Pavlištová for providing us with the revertant A. thaliana lines. Access to computing and storage facilities owned by parties and projects contributing to the National Grid Infrastructure MetaCentrum, provided under the programme “Projects of Large Research, Development, and Innovations Infrastructures” (CESNET LM2015042) is greatly appreciated. This work was supported by GACR 13-11563P, 16-04166Y and by the Ministry of Education, Youth and Sports of the Czech Republic-projects KONTAKT II no. LH15189 and CEITEC 2020 (LQ1601). Work in the laboratory of C.G. is supported by Grants BFU2012-34821 and BFU2015-68396-R.

Author contributions

J.F. and M.D. designed the study, K.H., M.D., I.M. and L.V. performed experiments, all authors performed analysis and interpretation of results, K.H., M.D. and J.F. wrote the manuscript.

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Correspondence to Martina Dvořáčková or Jiří Fajkus.

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Havlová, K., Dvořáčková, M., Peiro, R. et al. Variation of 45S rDNA intergenic spacers in Arabidopsis thaliana . Plant Mol Biol 92, 457–471 (2016). https://doi.org/10.1007/s11103-016-0524-1

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Keywords

  • Arabidopsis thaliana
  • Chromatin assembly factor
  • Nucleolus organizer region
  • 45S ribosomal DNA
  • Intergenic spacer
  • rDNA rearrangements