Natural variation in DNA methylation in ribosomal RNA genes of Arabidopsis thaliana

Eric J Richards,Hye Woo

doi:10.1186/1471-2229-8-92

Abstract

BackgroundDNA methylation is an important biochemical mark that silences repetitive sequences, such as transposons, and reinforces epigenetic gene expression states. An important class of repetitive genes under epigenetic control in eukaryotic genomes encodes ribosomal RNA (rRNA) transcripts. The ribosomal genes coding for the 45S rRNA precursor of the three largest eukaryotic ribosomal RNAs (18S, 5.8S, and 25–28S) are found in nucleolus organizer regions (NORs), comprised of hundreds to thousands of repeats, only some of which are expressed in any given cell. An epigenetic switch, mediated by DNA methylation and histone modification, turns rRNA genes on and off. However, little is known about the mechanisms that specify and maintain the patterns of NOR DNA methylation.ResultsHere, we explored the extent of naturally-occurring variation in NOR DNA methylation among accessions of the flowering plant Arabidopsis thaliana. DNA methylation in coding regions of rRNA genes was positively correlated with copy number of 45S rRNA gene and DNA methylation in the intergenic spacer regions. We investigated the inheritance of NOR DNA methylation patterns in natural accessions with hypomethylated NORs in inter-strain crosses and defined three different categories of inheritance in F1 hybrids. In addition, subsequent analysis of F2 segregation for NOR DNA methylation patterns uncovered different patterns of inheritance. We also revealed that NOR DNA methylation in the Arabidopsis accession Bor-4 is influenced by the vim1-1 (variant in methylation 1-1) mutation, but the primary effect is specified by the NORs themselves.ConclusionOur results indicate that the NORs themselves are the most significant determinants of natural variation in NOR DNA methylation. However, the inheritance of NOR DNA methylation suggests the operation of a diverse set of mechanisms, including inheritance of parental methylation patterns, reconfiguration of parental NOR DNA methylation, and the involvement of trans-acting modifiers.

Highlights

DNA methylation is an important biochemical mark that silences repetitive sequences, such as transposons, and reinforces epigenetic gene expression states
Our initial study of natural variation of nucleolus organizer region (NOR) DNA methylation in Arabidopsis focused on a small number of accessions [26], but here we extended our investigation to 88 Arabidopsis natural accessions to evaluate the degree of natural variation in NOR DNA methylation (Fig. 1C, Additional File 2)
The partitioning of hybridization signal among the three fractions defined general patterns, but some accessions deviated from the consensus patterns revealing that natural variation exists for qualitative differences in DNA methylation distribution [see Additional File 4], Our results indicate that ribosomal RNA (rRNA) gene copy number is a major predictor of NOR DNA methylation among Arabidopsis natural accessions (Fig. 2)

Summary

Introduction

DNA methylation is an important biochemical mark that silences repetitive sequences, such as transposons, and reinforces epigenetic gene expression states. DNA methylation is important for gene regulation, genomic imprinting, heterochromatin assembly, and protection of the genome against migrating transposable elements [2,3]. Both forward and reverse genetic approaches have identified important components of DNA methylation systems in eukaryotes,. The analysis of genetic variation among Arabidopsis accessions has identified components important for various aspects of the plant's biology, including the circadian clock [12], flowering time [13,14,15], pathogen resistance [16], and uncovered natural variation for drought responses [17] and freezing tolerance [18]. Exploring Arabidopsis accessions revealed that DNA methylation within the coding sequences of genes is highly polymorphic among Arabidopsis natural accessions and that these polymorphisms in genic methylation behave as heritable variation in inter-strain crosses [20]

Methods

Results

Discussion

Conclusion