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Transcriptome-wide discovery of microRNA binding sites in human brain by Ago2 HITS-CLIP [Ago2-miRNA complexes]

ABSTRACT: Here, seeking to gain insight into the array of transcripts engaged with miRNAs in human brain, we performed HITS-CLIP to profile transcriptome-wide Ago2:RNA interactions in a panel of eleven post-mortem adult human brain samples harvested from adult motor cortex and cingulate gyrus, regions associated with movement and psychiatric disorders . High-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP) Eleven post-mortem adult human brain tissues were subjected to ultraviolet radiation to crosslink proteins with nucleic acids, and HITS-CLIP was performed as previously described by Chi et al Nature. 2009 Jul 23;460(7254):479-86 using an anti-Ago2 polyclonal antibody and some additional modifications.

ORGANISM(S): Homo sapiens

SUBMITTER: Ryan Boudreau

PROVIDER: E-GEOD-52083 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Transcriptome-wide discovery of microRNA binding sites in human brain.

Boudreau Ryan L RL Jiang Peng P Gilmore Brian L BL Spengler Ryan M RM Tirabassi Rebecca R Nelson Jay A JA Ross Christopher A CA Xing Yi Y Davidson Beverly L BL

Neuron 20140102 2

The orchestration of brain function requires complex gene regulatory networks that are modulated, in part, by microRNAs (miRNAs). These noncoding RNAs associate with argonaute (Ago) proteins in order to direct posttranscriptional gene suppression via base pairing with target transcripts. In order to better understand how miRNAs contribute to human-specialized brain processes and neurological phenotypes, identifying their targets is of paramount importance. Here, we address the latter by profilin ...[more]

PMID: 24389009

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Transcriptome-wide discovery of microRNA binding sites in human brain by Ago2 HITS-CLIP [Ago2-miRNA-target mRNA complexes]

Project description:Here, seeking to gain insight into the array of transcripts engaged with miRNAs in human brain, we performed HITS-CLIP to profile transcriptome-wide Ago2:RNA interactions in a panel of eleven post-mortem adult human brain samples harvested from adult motor cortex and cingulate gyrus, regions associated with movement and psychiatric disorders . High-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP) Eleven post-mortem adult human brain tissues were subjected to ultraviolet radiation to crosslink proteins with nucleic acids, and HITS-CLIP was performed as previously described by Chi et al Nature. 2009 Jul 23;460(7254):479-86 using an anti-Ago2 polyclonal antibody and some additional modifications.

2014-01-03 | E-GEOD-52082 | biostudies-arrayexpress

Elucidation of Transcriptome-Wide MicroRNA Binding Sites in Human Cardiac Tissues by Ago2 HITS-CLIP

Project description:We deciphered Ago2:RNA interactions in post-mortem human heart tissues using crosslinking immunoprecipitation coupled with high-throughput sequencing (HITS-CLIP) to generate the first transcriptome-wide map of miR targeting events in human myocardium, detecting 4000 cardiac Ago2 binding sites across >2200 target transcripts. Profiling AGO2-associated RNAs in cardiac tissues obtained from 6 donors with heart diseases of differing etiologies

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Transcriptome-wide discovery of microRNA binding sites in human brain by Ago2 HITS-CLIP [Ago2-miRNA complexes]

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Transcriptome-wide discovery of microRNA binding sites in human brain by Ago2 HITS-CLIP [Ago2-miRNA-target mRNA complexes]

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Remodeling of Ago2-mRNA interactions upon cellular stress reflects miRNA complementarity and correlates with altered translation rates (part 3)

Project description:When adapting to environmental stress, cells attenuate and reprogram their translational output. In part, these altered translation profiles are established through changes in the interactions between RNA-binding proteins and mRNAs. The Ago2/microRNA machinery has been shown to participate in stress-induced translational upregulation of a particular mRNA, CAT-1; however, a detailed, transcriptome-wide understanding of the involvement of Ago2 in the process has been lacking. Here, we profiled the overall changes in Ago2-mRNA interactions upon arsenite stress by CLIP-seq. Ago2 displayed a significant remodeling of its transcript occupancy, with the majority of 3` UTR and CDS sites exhibiting stronger interaction. Interestingly, target sites that were destined for release from Ago2 upon stress were depleted in miRNA complementarity signatures, suggesting an alternative mode of interaction. To compare the changes in Ago2 binding patterns across transcripts with changes in their translational states, we measured mRNA profiles on ribosome/polysome gradients by RNA-seq. Increased Ago2 occupancy correlated with stronger repression of translation for those mRNAs, as evidenced by a shift toward lighter gradient fractions upon stress, while release of Ago2 was associated with the limited number of transcripts that remained translated. Taken together, these data point to a role for Ago2 and the mammalian microRNAs in mediating the translational component of the stress response. In this sub-series, CLIP and RNAseq data from the hippuristanol treatment experiment are presented. Experiments on 293S cells +/- hippuristanol treatment, in 2 biological replicates. Here, for each treatment/replicate sample, an aliquot was used for RNA-seq, and the rest was split into three aliquots to perform 3 parallel CLIP-seq protocols with different antibodies. So, each RNA-seq dataset here corresponds to 3 CLIP-seq datasets.

2013-07-15 | E-GEOD-44378 | biostudies-arrayexpress

Remodeling of Ago2-mRNA interactions upon cellular stress reflects miRNA complementarity and correlates with altered translation rates (part 2)

Project description:When adapting to environmental stress, cells attenuate and reprogram their translational output. In part, these altered translation profiles are established through changes in the interactions between RNA-binding proteins and mRNAs. The Ago2/microRNA machinery has been shown to participate in stress-induced translational upregulation of a particular mRNA, CAT-1; however, a detailed, transcriptome-wide understanding of the involvement of Ago2 in the process has been lacking. Here, we profiled the overall changes in Ago2-mRNA interactions upon arsenite stress by CLIP-seq. Ago2 displayed a significant remodeling of its transcript occupancy, with the majority of 3` UTR and CDS sites exhibiting stronger interaction. Interestingly, target sites that were destined for release from Ago2 upon stress were depleted in miRNA complementarity signatures, suggesting an alternative mode of interaction. To compare the changes in Ago2 binding patterns across transcripts with changes in their translational states, we measured mRNA profiles on ribosome/polysome gradients by RNA-seq. Increased Ago2 occupancy correlated with stronger repression of translation for those mRNAs, as evidenced by a shift toward lighter gradient fractions upon stress, while release of Ago2 was associated with the limited number of transcripts that remained translated. Taken together, these data point to a role for Ago2 and the mammalian microRNAs in mediating the translational component of the stress response. In this sub-series, CLIP and RNAseq data from the emetine treatment experiment are presented. Experiments on 293S cells +/- emetine treatment, in 1 biological replicate. Here, for each treatment/replicate sample, an aliquot was used for RNA-seq, and the rest was split into three aliquots to perform 3 parallel CLIP-seq protocols with different antibodies. So, each RNA-seq dataset here corresponds to 3 CLIP-seq datasets.

2013-07-15 | E-GEOD-44377 | biostudies-arrayexpress

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2016-08-09 | GSE83410 | GEO

Remodeling of Ago2-mRNA interactions upon cellular stress reflects miRNA complementarity and correlates with altered translation rates (part 1)

Project description:When adapting to environmental stress, cells attenuate and reprogram their translational output. In part, these altered translation profiles are established through changes in the interactions between RNA-binding proteins and mRNAs. The Ago2/microRNA machinery has been shown to participate in stress-induced translational upregulation of a particular mRNA, CAT-1; however, a detailed, transcriptome-wide understanding of the involvement of Ago2 in the process has been lacking. Here, we profiled the overall changes in Ago2-mRNA interactions upon arsenite stress by CLIP-seq. Ago2 displayed a significant remodeling of its transcript occupancy, with the majority of 3` UTR and CDS sites exhibiting stronger interaction. Interestingly, target sites that were destined for release from Ago2 upon stress were depleted in miRNA complementarity signatures, suggesting an alternative mode of interaction. To compare the changes in Ago2 binding patterns across transcripts with changes in their translational states, we measured mRNA profiles on ribosome/polysome gradients by RNA-seq. Increased Ago2 occupancy correlated with stronger repression of translation for those mRNAs, as evidenced by a shift toward lighter gradient fractions upon stress, while release of Ago2 was associated with the limited number of transcripts that remained translated. Taken together, these data point to a role for Ago2 and the mammalian microRNAs in mediating the translational component of the stress response. In this sub-series, CLIP and RNAseq data from the arsenite treatment experiment are presented. Experiments on 293S cells +/- arsenite treatment, in 4 biological replicates. On each treatment/replicate biological sample, both a CLIP-seq protocol and an RNA-seq protocol were performed, so these datasets are “paired” in addition to the treatment pairing.

2013-07-15 | E-GEOD-44376 | biostudies-arrayexpress

A nuclear role for the Argonaute protein AGO2 in mammalian gametogenesis

Project description:The reduced sperm count observed in Ago2 cKO mice implies that AGO2 has non-redundant functions in the male germ line. Because AGO2 is a key protein in the RNA interference (RNAi) pathway, we first postulated that AGO2 loss disrupts normal transcriptional and translational dynamics of target mRNAs relevant to sperm maturation. To address this hypothesis, we took advantage of the apparently normal spermatogenesis in Ago2 cKO mice, which allowed us to purify matched meiotic and post-meiotic germ cells from Ago2 cKO and wild type controls. We examined changes in the transcriptome and proteome of these two spermatogenic stages in Ago2 cKO relative to control mice using RNA-seq and quantitative mass spectrometry (MS). To further examine if the changes in mRNA and protein levels detected in Ago2 cKO germ cells was due to a loss of regulation by the canonical AGO2-miRNA pathway, we characterized AGO2 protein interactors by AGO2 immunoprecipitation-mass spectrometry (IP-MS) in cytoplasmic and nuclear fractions of post-meiotic cells

2022-08-01 | PXD027935 | Pride

Homo sapiens

Project description:Human Ago2-HITS-CLIP

| PRJNA389304 | ENA