Heavy de novo methylation at symmetrical and non-symmetrical sites is a hallmark of RNA-directed DNA methylation

Pélissier, Thierry; Thalmeir, Sabine; Kempe, Dirk; Sänger, Heinz-L.; Wassenegger, Michael

doi:10.1093/nar/27.7.1625

Cited by 166 publications

(123 citation statements)

References 40 publications

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“…So we propose that silencing involved a cis-acting mechanism via methylation spreading in cis from the T-DNA. Similar observations were previously reported, showing that cytosine methylation can spread from the T-DNA or a foreign DNA to the host genome (Pelissier et al, 1999). It was also reported that the methylation pattern of a chromosomal region strongly influences gene activity of T-DNA by imposing DNA methylation at the borders of the integrated DNA (Pröls and Meyer, 1992).…”

Section: Discussionsupporting

confidence: 85%

Relationship between allelic state of T-DNA and DNA methylation of chromosomal integration region in transformed Arabidopsis thaliana plants

2005

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T-DNA insertions are currently used as a tool to introduce, or knock out, specific genes. The expression of the inserted gene is frequently haphazard and up to now, it was proposed that transgene expression depends on the site of insertion within the genome, as well as the number of copies of the transgene. In this paper, we show that the allelic state of a T-DNA insertion can be at the origin of epigenetic silencing. A T-DNA insertional mutant was characterized to explore the function of AtBP80a', a vacuolar sorting receptor previously associated with germination. Seeds homozygous for the T-DNA do not germinate, but this can be overcome by a cold treatment and maintained by the following generations. The nongerminating phenotype is only observed in homozygous seed produced by heterozygous plants indicating that it is correlated with the allelic state of the T-DNA in parental lines. Analysis of the region between the T-DNA insertion and the ATG codon of atbp80a' showed that cytosine methylation is highly enhanced in chromatin containing the T-DNA. Data presented here show that an unpaired DNA region during meiosis could be at the origin of a de novo cytosine methylation mechanism.

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Section: Discussionsupporting

confidence: 85%

Relationship between allelic state of T-DNA and DNA methylation of chromosomal integration region in transformed Arabidopsis thaliana plants

2005

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“…The results indicate that the diminished expression of GFP: Sensor protein was due to specific cleavage of mRNA and not due to inhibition of translation. The results also indicate that the expression of reporter genes was not silenced at the transcriptional level, eliminating the possibility of transcriptional silencing caused by PSTVd RNA-mediated DNA methylation of the transgenes (63,96).…”

Section: Vol 81 2007 Viroid Rna Serves As Substrate For Dicer-like mentioning

confidence: 80%

A Structured Viroid RNA Serves as a Substrate for Dicer-Like Cleavage To Produce Biologically Active Small RNAs but Is Resistant to RNA-Induced Silencing Complex-Mediated Degradation

Itaya¹,

Zhong²,

Bundschuh

et al. 2007

J Virol

185

177

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RNA silencing is a potent means of antiviral defense in plants and animals. A hallmark of this defense response is the production of 21-to 24-nucleotide viral small RNAs via mechanisms that remain to be fully understood. Many viruses encode suppressors of RNA silencing, and some viral RNAs function directly as silencing suppressors as counterdefense. The occurrence of viroid-specific small RNAs in infected plants suggests that viroids can trigger RNA silencing in a host, raising the question of how these noncoding and unencapsidated RNAs survive cellular RNA-silencing systems. We address this question by characterizing the production of small RNAs of Potato spindle tuber viroid (srPSTVds) and investigating how PSTVd responds to RNA silencing. Our molecular and biochemical studies provide evidence that srPSTVds were derived mostly from the secondary structure of viroid RNAs. Replication of PSTVd was resistant to RNA silencing, although the srPSTVds were biologically active in guiding RNA-induced silencing complex (RISC)-mediated cleavage, as shown with a sensor system. Further analyses showed that without possessing or triggering silencing suppressor activities, the PSTVd secondary structure played a critical role in resistance to RISC-mediated cleavage. These findings support the hypothesis that some infectious RNAs may have evolved specific secondary structures as an effective means to evade RNA silencing in addition to encoding silencing suppressor activities. Our results should have important implications in further studies on RNA-based mechanisms of host-pathogen interactions and the biological constraints that shape the evolution of infectious RNA structures.A major focus of current biology is to understand how a pathogen has evolved mechanisms to achieve a balance among several interrelated activities that are crucial to establish a full infection: evading or suppressing host defense, minimizing destructive interference of host metabolism, and maximizing utilization of host factors to support replication and systemic spread. A full understanding of these mechanisms is not only necessary to build a foundation for developing technologies to combat pathogen diseases but also can provide fundamental mechanistic insights into the regulation of basic cellular processes.Recent studies have discovered small RNA-mediated gene silencing as a powerful antiviral mechanism in plants and animals (6,22,25,47,49,50,72,77,84,(87)(88)(89)98). Furthermore, small RNA-mediated gene silencing plays essential roles in regulating a wide variety of growth and development processes (4-6, 11, 13, 17, 23, 28, 42). A key mediator of RNA silencing is several classes of 21-to 24-nucleotide (nt) small RNAs.MicroRNAs (miRNAs) are produced by cleavage of hairpin RNA precursors encoded by the genome of an organism. Short interfering RNAs (siRNAs) are generated by cleavage of double-stranded RNAs (dsRNAs) that may originate from several sources, including cellular genomes, viral replication intermediates, aberrant cellular RNAs, overexpresse...

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“…Methylation analysis of centromeric repeats and rDNA repeats was conducted as described by others (32). Bisulfite sequencing was performed as described (12,34). The following degenerate primers, which allowed for unconverted or converted cytosines, were used to amplify the top strand of the NOS-NPTII target: 5Ј primer 5Ј-YATGAGYG-GAGAATTAAGGGAGT-3Ј (Y ϭ C or T); 3Ј primer 5Ј-CCRAATARCCTCTCCACCCAA-3Ј (R ϭ G or A).…”

Section: Methodsmentioning

confidence: 99%

RNA-directed DNA methylation inArabidopsis

Aufsatz

Mette

Winden

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

241

230

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In plants, double-stranded RNA that is processed to short RNAs Ϸ21-24 nt in length can trigger two types of epigenetic gene silencing. Posttranscriptional gene silencing, which is related to RNA interference in animals and quelling in fungi, involves targeted elimination of homologous mRNA in the cytoplasm. RNAdirected DNA methylation involves de novo methylation of almost all cytosine residues within a region of RNA-DNA sequence identity. RNA-directed DNA methylation is presumed to be responsible for the methylation observed in protein coding regions of posttranscriptionally silenced genes. Moreover, a type of transcriptional gene silencing and de novo methylation of homologous promoters in trans can occur if a double-stranded RNA contains promoter sequences. Although RNA-directed DNA methylation has been described so far only in plants, there is increasing evidence that RNA can also target genome modifications in other organisms. To understand how RNA directs methylation to identical DNA sequences and how changes in chromatin configuration contribute to initiating or maintaining DNA methylation induced by RNA, a promoter double-stranded RNA-mediated transcriptional gene silencing system has been established in Arabidopsis. A genetic analysis of this system is helping to unravel the relationships among RNA signals, DNA methylation, and chromatin structure.T he term ''RNA silencing'' refers to epigenetic gene silencing effects that are initiated by double-stranded RNA (dsRNA) (1). Discovered independently in plants, fungi, and animals, RNA silencing phenomena are revealing new ways to repress gene expression and to subdue transposable elements and viruses that produce dsRNA during their replication cycle (2-8). A fundamental step in RNA silencing pathways is cleavage of dsRNA into short RNAs (9), which are believed to act as guides for enzyme complexes that either degrade or modify homologous nucleic acids.The most familiar type of RNA silencing occurs primarily in the cytoplasm and is termed posttranscriptional gene silencing (PTGS) in plants, quelling in Neurospora, and RNA interference (RNAi) in animals. PTGS͞RNAi involves a dsRNA that is processed by an RNase III-like enzyme called Dicer into short interfering (si) RNAs 21-22 nt in length. The antisense siRNAs associate with a ribonuclease complex and guide sequencespecific degradation of complementary mRNAs (5-8).A second form of RNA silencing involves sequence-specific changes at the genome level. RNA-directed DNA methylation (RdDM) (10), which has been described so far only in plants, leads to de novo methylation of almost all cytosine residues within the region of sequence identity between the triggering RNA and the target DNA. Similarly to PTGS͞RNAi, RdDM requires a dsRNA that is cleaved to short RNAs Ϸ21-24 nt in length (11). It is not yet certain whether the short RNAs or dsRNA guide methylation of homologous DNA sequences, although the length of short RNAs is consistent with the minimum DNA target size of RdDM (Ϸ30 bp) (12).RdDM is assumed to be the ...

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Heavy de novo methylation at symmetrical and non-symmetrical sites is a hallmark of RNA-directed DNA methylation

Cited by 166 publications

References 40 publications

Relationship between allelic state of T-DNA and DNA methylation of chromosomal integration region in transformed Arabidopsis thaliana plants

Relationship between allelic state of T-DNA and DNA methylation of chromosomal integration region in transformed Arabidopsis thaliana plants

A Structured Viroid RNA Serves as a Substrate for Dicer-Like Cleavage To Produce Biologically Active Small RNAs but Is Resistant to RNA-Induced Silencing Complex-Mediated Degradation

RNA-directed DNA methylation inArabidopsis

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