Plant and Cell Physiology Advance Access originally published online on April 5, 2007
Plant and Cell Physiology 2007 48(5):745-752; doi:10.1093/pcp/pcm043
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Regulation of Arabidopsis thaliana 5S rRNA Genes
1Unité Mixte de Recherche CNRS 6547 BIOMOVE, Université Blaise Pascal, 24 Avenue des Landais, F-63177 Aubière Cedex, France
2Leibniz- Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstrasse 3, D-06466 Gatersleben, Germany
*Corresponding author: E-mail, sylvette.tourmente{at}univ-bpclermont.fr; Fax, +33-4-73-40-77-77.
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Abstract |
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The Arabidopsis thaliana genome comprises around 1,000 copies of 5S rRNA genes encoding both major and minor 5S rRNAs. In mature wild-type leaves, the minor 5S rRNA genes are silent. Using different mutants of DNA methyltransferases (met1, cmt3 and met1 cmt3), components of the RNAi pathway (ago4) or post-translational histone modifier (hda6/sil1), we show that the corresponding proteins are needed to maintain proper methylation patterns at heterochromatic 5S rDNA repeats. Using reverse transcription–PCR and cytological analyses, we report that a decrease of 5S rDNA methylation at CG or CNG sites in these mutants leads to the release of 5S rRNA gene silencing which occurred without detectable changes of the 5S rDNA chromatin structure. In spite of severely reduced DNA methylation, the met1 cmt3 double mutant revealed no increase in minor 5S rRNA transcripts. Furthermore, the release of silencing of minor 5S rDNAs can be achieved without increased formation of euchromatic loops by 5S rDNA, and is independent from the global heterochromatin content. Additionally, fluorescence in situ hybridization with centromeric 180 bp repeats confirmed that these highly repetitive sequences, in spite of their elevated transcriptional activity in the DNA methyltransferase mutants (met1, cmt3 and met1 cmt3), remain within chromocenters of the mutant nuclei.
Keywords: Arabidopsis thaliana - 5S rDNA - DNA methylation - Transcription
Abbreviations: CC, chromocenter; DAPI, 4',6-diamidino-2-phenylindole; FISH, fluorescence in situ hybridization; RdDM, RNA-directed DNA methylation; RISC, RNA-induced silencing complex; RNAi, RNA interference; RT–PCR, reverse transcription–PCR; siRNA, small interfering RNA; WT, wild type.
3Present address: University of Geneva, Laboratory of Plant Genetics Sciences III, 30 Quai Ernest-Ansermet, CH-1211 Genève 4, Switzerland.
(Received February 20, 2007; Accepted April 4, 2007)
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