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Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Genome-wide maps of AGO1-bound small RNA in Arabidopsis leaves

ABSTRACT: We report flg22 regulate the accumulation of AGO1-bound small RNA in arabidopsis. We find that a number of miRNAs are up- or down-regulated by flg22, a well-studied PAMP. Examination of AGO1-bound small RNAs with or without flg22 treatment.

ORGANISM(S): Arabidopsis thaliana

SUBMITTER: Jianmin Zhou

PROVIDER: E-GEOD-20448 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Identification of microRNAs involved in pathogen-associated molecular pattern-triggered plant innate immunity.

Li Yan Y Zhang QingQing Q Zhang Jiangguang J Wu Liang L Qi Yijun Y Zhou Jian-Min JM

Plant physiology 20100217 4

Pathogen-associated molecular patterns (PAMPs) trigger plant defenses when perceived by surface-localized immune receptors. PAMP-triggered immunity (PTI) plays a vital role in the resistance of plants to numerous potential pathogens. MicroRNA (miRNA) biogenesis is known to be important for PTI, but miRNA species involved in this process have not been fully explored. Here we show that the Arabidopsis (Arabidopsis thaliana) miRNA effector protein, Argonaute1 (AGO1), is required for a number of PTI ...[more]

PMID: 20164210

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Profiling of Argonaute (AGO) 2-associated sRNAs after infection of bacterial pathogen Pseudomonas syringae

Project description:Transgenic Arabidopsis plants (AGO2::HA:AGO2) were treated with either mock (10 mM MgCl2) or Pseudomonas syringae pv. tomato (Pst) expressing avrRpt2 (R2) at a concentration of 2 x 107 cfu/ml for 14 hours. sRNAs associated with AGO1 and AGO2 were co-immunoprecipitated using antibodies against either AGO1 (AGO1-IP), or HA (hemagglutinin) (AGO2-IP). As controls, we also gel-purified the 18-28 nt fraction of the total RNAs from an AGO2 mutant (ago2-1). The co-immunoprecipitated or gel-purified RNAs were cloned and sequenced by Illumina deep sequencing. Examination of AGO-associated sRNAs in pathogen-treated or control plants

2011-05-10 | E-GEOD-26161 | biostudies-arrayexpress

Virus infection triggers widespread silencing of host genes by a distinct class of endogenous siRNAs in Arabidopsis

Project description:Virus infection triggers widespread silencing of host genes by a distinct class of endogenous siRNAs in Arabidopsis This study constructed and sequenced two independent small RNA libraries from the upper uninoculated leaves of (i) WT Arabidopsis plants 14 d after mock inoculation and of (ii) rdr1 rdr6 plants 14 d after infection with Fny CMV-Δ2b, and one library each from the upper uninoculated leaves of (iii) WT, (iv) rdr1, and (v) rdr6 plants 14 d after infection with TuMV-GFP. Coimmunoprecipitation with FLAG- and HA-specific antibodies was used to obtain (vi,vii) AGO1 and (viii,ix) AGO2 complexes, respectively, from the FLAG-AGO1/ago1-36 and HA-AGO2 plants 14 d after infection with Fny CMV-Δ2b for extracting total loaded small RNAs for the construction and sequencing of small RNA libraries.

2014-08-30 | E-GEOD-60929 | biostudies-arrayexpress

Small RNA loading licenses Argonaute for assembly into a transcriptional silencing complex

Project description:We analyzed the small RNAs bound to the fission yeast RITS and ARC complexes, and to Ago1 in wild-type and ARC mutant cells, to gain insight into the molecular role of ARC. We found that ARC contains longer heterochromatic siRNAs than RITS, and that ARC subunit Arb1 is required for loading small RNAs onto Ago1. Sequencing of 18-28 nt small RNAs from Tas3, Arb1 and Ago1 purifications.

2015-03-01 | E-GEOD-65223 | biostudies-arrayexpress

Profiling of small RNA populations in rice total extract and purified AGO1 complexes

Project description:microRNAs (miRNAs) are a class of small silencing RNAs that have regulatory roles in gene expression. miRNAs interact with Argonaute (AGO) proteins to form effector complexes that can cleave target mRNAs or repress their translation. Rice encodes four AGO1 homologs (AGO1a, AGO1b, AGO1c, and AGO1d). We used an RNAi approach to knock down the four AGO1s. The RNAi lines displayed pleiotropic developmental phenotypes and had increased accumulation of miRNA targets, suggesting the involvement of AGO1s in the miRNA pathway. Three of the AGO1s (AGO1a, AGO1b, and AGO1c complexes) were purified and further characterized. We showed that the three AGO1s all have a strong preference for binding small RNAs (sRNAs) with 5’ U and have Slicer activity. We catalogued the sRNAs in each AGO1 complex by deep sequencing, and found all three AGO1s predominantly bound known miRNAs. Most of the miRNAs were evenly distributed in the three AGO1 complexes, suggesting a redundant role for the AGO1s in the function of these miRNAs. Intriguingly, we also found a subset of miRNAs were specifically incorporated into or excluded from one of the AGO1s, suggesting that there is also functional specialization among the rice AGO1s. Four samples, total extract and three AGO1 complexes were analyzed

2009-11-23 | E-GEOD-18250 | biostudies-arrayexpress

Incorporation of transposable element small RNAs into the Arabidopsis AGO1 protein

Project description:Background: Transposable element 24 nucleotide small RNAs are not efficiently incorporated into the AGO1 protein, which is involved in endogenous RNAi and gene regulation through the microRNA and tasiRNA pathways. Results: The AGO1 protein incorporates large quantities of transposable element siRNAs when transposable elements are epigenetically activated and transcribed. The incorporation of transposable element siRNAs is at the expense of the most abundant microRNAs. These transposable element siRNAs can act as tasiRNAs, regulating genes that they have partial complementarity to. Conclusion: Transposable element small RNAs are more dynamic than previously thought. They can be incorporated into AGO1 and regulate genes. Three biological replicates of small RNA sequencing from two genotypes

2014-04-07 | E-GEOD-45990 | biostudies-arrayexpress

Identification of small RNAs in rice AGO1 complexes and their targets

Project description:This SuperSeries is composed of the following subset Series: GSE18248: Sequencing of rice degradome GSE18250: Profiling of small RNA populations in rice total extract and purified AGO1 complexes Refer to individual Series

2009-11-23 | E-GEOD-18251 | biostudies-arrayexpress

Argonaute Surveillance and Dicer-Independent priRNAs TriggerRNAi and Heterochromatin Formation

Project description:The assembly of fission yeast pericentromeric heterochromatin and generation of small interfering RNAs (siRNAs) from noncoding centromeric transcripts are mutually dependent processes. How this interdependent positive feedback loop is first triggered is a fundamental unanswered question. Here we show that two distinct Argonaute (Ago1)-dependent pathways mediate small RNA generation. RNA-dependent RNA polymerase complex (RDRC) and Dicer act on specific noncoding RNAs to generate siRNAs by a mechanism that requires the slicer activity of Ago1 but is independent of pre-existing heterochromatin. In the absence of RDRC or Dicer, a distinct class of small RNAs, called primal small RNAs (priRNAs), associate with Ago1. priRNAs are degradation products of abundant transcripts, which bind to Ago1 and target antisense transcripts that result from bidirectional transcription of DNA repeats. Our results suggest that a transcriptome surveillance mechanism based on the random association of RNA degradation products with Argonaute triggers siRNA amplification and heterochromatin assembly within DNA repeats. small RNA profiling in wild type S. pombe cells and in 12 mutant cells

2010-02-23 | E-GEOD-19734 | biostudies-arrayexpress

MCMV infection of cultured mouse cells induces expression of miR-7a.

Project description:One goal of viral infection is to reprogram the host cell to optimize viral replication. As part of this process, viral miRNAs may compete for components of the miRNA/siRNA pathway as well as regulate cellular targets. Mouse Cytomegalovirus has been described to generate large numbers of viral miRNAs during lytic infection and was therefore used to analyze the impact of viral miRNAs on the host cell small RNA system as well as to check for sorting of viral small RNAs into specific Ago-proteins. Deep sequencing analysis of MCMV infected cells revealed that viral miRNAs represent only app. 13% of all detected miRNAs. All previously described MCMV miRNAs with the exception of miR-m88-1* were confirmed and for the MCMV miR-m01-1 hairpin an additional miRNA, designated miR-m01-1-3p, was found. Its presence was confirmed by qPCR and Northern Blot. Deep sequencing after RISC IP with antibodies specific for either Ago1 or Ago2 showed that all MCMV miRNAs are loaded into both RISC complexes. The ratio of MCMV to mouse miRNAs was not increased after immunoprecipitation of Ago-proteins. Viral miRNAs therefore do not overwhelm the host miRNA processing system nor are they preferentially incorporated into RISC. We found that 3 mouse miRNAs showed an altered expression due to MCMV infection. Down-regulation of miR-27a, as previously described, could be confirmed. In addition, miR-26a was down-regulated and an up-regulation of miR-7a dependent on viral protein expression could be observed. Examination of small RNA expression in uninfected vs. infected cells, immunoprecipitation + sequencing of Ago1 and Ago2 loaded small RNAs in infected cells

2012-06-30 | E-GEOD-36919 | biostudies-arrayexpress

Hairpin RNA induces secondary small interfering RNA synthesis and silencing in trans in fission yeast

Project description:We provide evidence of the generation of Rdp1-mediated secondary siRNAs in vivo in fission yeast. Secondly, we show that the presence of Ago1-associated siRNAs does not guarantee robust silencing. We obtained sequences from gel purified small RNAs of wt and rdp1delta fission yeast cells and FLAG-ago1 associated small RNAs.

2010-02-15 | E-GEOD-20330 | biostudies-arrayexpress

Ago1 vs Ago2-IP small RNA deep-sequencing with age in Drosophila

Project description:Drosophila miRNAs show distinct change in isoform distribution pattern with age. Some miRNAs show accumulation of the short isoforms, while other miRNAs show the accumulation of the long isoforms with age. The increase of the long isoforms of some miRNAs reflects increased 2'-O-methylated miRNA isoforms with age. The increase in 2'-O-methylated miRNA isoforms reflected increased Ago2-loading, but not Ago1-loading of specific miRNA isoforms with age. This raised a question on whether there is global shift in small RNA loading pattern between Ago1 and Ago2 with age. To investigate change in small RNA loading pattern between Ago1 and Ago2 with age, we performed small RNA deep-sequencing of Ago1 vs Ago2-IP small RNAs at 3d and 30d in Drosophila. This analysis revealed global increase of miRNA loading into Ago2, but not into Ago1 with age. 3d and 30d FLAG-HA-Ago2 male flies were collected. Ago1 and Ago2 were immunoprecipitated by anti-Ago1 and anti-FLAG M2 beads respectively. RNA was purified from the beads, P32-labeled, and small RNA fraction was gel-purififed. Small RNA libraries were prepared using Illumina's TruSeq small RNA sample preparation kit (#RS-200-0012, Illumina, Inc. San Diego, CA), following the manufacturer's protocol. The libraries were multiplexed and sequenced on HiSeq2000 platform (Illumina).

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