Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Structural Variant of a Tandem Repeat-containing Silencer Locus Reveals Distinct Pol II and Pol IV-dependent Pathways of siRNA Biogenesis

ABSTRACT: We used a two-component transgene system to study the RNA-dependent DNA methylation (RdDM) and transcriptional gene silencing (TGS) in Arabidopsis. By profiling the small RNA population in mutants defected in RdDM or RNA polymerase II-transcribed trigger for generating silencing siRNA, we investigated how repetitive loci such as tandem repeats were regulated transcriptionally through the action of RNA polymerase IV. Genome-wide small RNA profiling was done by Illumina TruSeq sample preparation followed by high-throughput sequencing with the Illumina HiSeq 2000 platform. The six samples represent mutants and their corresponding control lines.

ORGANISM(S): Arabidopsis thaliana

SUBMITTER: Blake Meyers

PROVIDER: E-GEOD-47852 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Distinct and concurrent pathways of Pol II- and Pol IV-dependent siRNA biogenesis at a repetitive trans-silencer locus in Arabidopsis thaliana.

Sasaki Taku T Lee Tzuu-fen TF Liao Wen-Wei WW Naumann Ulf U Liao Jo-Ling JL Eun Changho C Huang Ya-Yi YY Fu Jason L JL Chen Pao-Yang PY Meyers Blake C BC Matzke Antonius J M AJ Matzke Marjori M

The Plant journal : for cell and molecular biology 20140613 1

Short interfering RNAs (siRNAs) homologous to transcriptional regulatory regions can induce RNA-directed DNA methylation (RdDM) and transcriptional gene silencing (TGS) of target genes. In our system, siRNAs are produced by transcribing an inverted DNA repeat (IR) of enhancer sequences, yielding a hairpin RNA that is processed by several Dicer activities into siRNAs of 21-24 nt. Primarily 24-nt siRNAs trigger RdDM of the target enhancer in trans and TGS of a downstream GFP reporter gene. We anal ...[more]

PMID: 24798377

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Project description:In plants, RNA polymerase II (Pol II) transcription of inverted DNA repeats produces hairpin RNAs that are processed by several DICER-LIKE enzymes into siRNAs that are 21-24-nt in length. When targeted to transcriptional regulatory regions, the 24-nt size class can induce RNA-directed DNA methylation (RdDM) and transcriptional gene silencing (TGS). In a forward genetic screen to identify mutants defective in RdDM of a target enhancer leading to TGS of a downstream GFP reporter gene in Arabidopsis thaliana, we recovered a structurally mutated silencer locus, named SM-NM-^T35S, in which the 35S promoter driving transcription of an inverted repeat of target enhancer sequences had been specifically deleted. Although Pol II-dependent, hairpin-derived 21-24-nt siRNAs were no longer generated at the newly created SM-NM-^T35S locus, the GFP reporter gene was nevertheless still partially silenced. Silencing was associated with methylation in a short tandem repeat in the upstream target enhancer and with low levels of 24-nt tandem repeat siRNAs. Introducing an nrpd1 mutation into the SM-NM-^T35S line fully released GFP silencing and eliminated both the tandem repeat methylation and associated 24-nt siRNAs, demonstrating their dependence on Pol IV. Deletion of the 35S promoter thus revealed a Pol IV-dependent pathway of 24-nt siRNA biogenesis that was previously inhibited or masked by the Pol II-dependent pathway in wild-type plants. Both Pol II- and Pol IV-dependent siRNAs accrued predominantly from cytosine (C)-containing segments of the tandem repeat monomer, suggesting that the local base composition influenced siRNA accumulation. Preferential accumulation of siRNAs at C-containing sequences was also observed at an endogenous tandem repeat comprising discrete C-rich and AT-rich sections. Our studies illuminate the potential complexity of siRNA generation at repeat-containing loci and show that Pol IV can act in siRNA biogenesis in the absence of a conventional Pol II promoter. Examination of whole-genome DNA methylation status in transgenic T+S Arabidopsis plant

Dataset Information

Structural Variant of a Tandem Repeat-containing Silencer Locus Reveals Distinct Pol II and Pol IV-dependent Pathways of siRNA Biogenesis

Publications

Distinct and concurrent pathways of Pol II- and Pol IV-dependent siRNA biogenesis at a repetitive trans-silencer locus in Arabidopsis thaliana.

Similar Datasets

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