Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Contribution of sequence and structure to mRNA-binding of Argonaute/miRNA complexes and degradation of miRNA targets

ABSTRACT: Relative contribution of sequence and structural features to the mRNA-binding of Argonaute/miRNA complexes and the degradation of miRNA targets How miRNAs recognize their target sites is a puzzle that many experimental and computational studies aimed to solve. Several features such as perfect pairing of the miRNA seed, additional pairing in the 3’ region of the miRNA, relative position in the 3’ UTR and the A/U content of the environment of the putative site have been found to be relevant. Here we have used a large number of previously published data sets to assess the power that various sequence and structure features have in distinguishing between putative sites that do and those that do not appear to be functional. We found that although different data sets can give widely different answers when it comes to ranking the relative importance of these features, the sites inferred from most transcriptomics experiments as well as from comparative genomics appear similar at this level. This suggests that miRNA target sites have been selected in evolution on their ability to trigger mRNA degradation. To understand at what step in the miRNA induced response individual features play a role, we further transfected human HEK293 cells with miRNAs and analyzed the association of Argonaute/miRNA complexes with target mRNAs and the degradation of these messages. We found that structural features are only important for Argonaute binding, while sequence features such as the A/U content of the 3’ UTR are important for mRNA degradation. Keywords: IP-array analysis

ORGANISM(S): Homo sapiens

PROVIDER: GSE14537 | GEO | 2009/01/27

SECONDARY ACCESSION(S): PRJNA111621

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Transcription profiling of human HEK cells transfected with miRNA to investigate the contribution of sequence and structure to mRNA-binding of Argonaute/miRNA complexes and degradation of miRNA targets

Project description:Relative contribution of sequence and structural features to the mRNA-binding of Argonaute/miRNA complexes and the degradation of miRNA targets; How miRNAs recognize their target sites is a puzzle that many experimental and computational studies aimed to solve. Several features such as perfect pairing of the miRNA seed, additional pairing in the 3â region of the miRNA, relative position in the 3â UTR and the A/U content of the environment of the putative site have been found to be relevant. Here we have used a large number of previously published data sets to assess the power that various sequence and structure features have in distinguishing between putative sites that do and those that do not appear to be functional. We found that although different data sets can give widely different answers when it comes to ranking the relative importance of these features, the sites inferred from most transcriptomics experiments as well as from comparative genomics appear similar at this level. This suggests that miRNA target sites have been selected in evolution on their ability to trigger mRNA degradation. To understand at what step in the miRNA induced response individual features play a role, we further transfected human HEK293 cells with miRNAs and analyzed the association of Argonaute/miRNA complexes with target mRNAs and the degradation of these messages. We found that structural features are only important for Argonaute binding, while sequence features such as the A/U content of the 3â UTR are important for mRNA degradation. Experiment Overall Design: Ago2 IPs of mock, miRNA-7, and miR-124 transfected cells were analyzed for 2 biological replicates. IPs of the biological replicates were analyzed with 2 technical replicates.

2009-01-31 | E-GEOD-14537 | biostudies-arrayexpress

miRNA-target chimeras identified with CLEAR-CLIP reveal miRNA 3' end pairing as a major determinant of Argonaute binding in vivo [mouse cortex CLEAR-CLIP]

Project description:microRNAs (miRNAs) act as sequence-specific guides for Argonaute (AGO) proteins, which mediate post-transcriptional silencing of target mRNAs. Despite their importance in many biological processes, rules governing AGO-miRNA targeting are only partially understood. We use a modified AGO HITS-CLIP strategy, termed CLEAR (Covalent Ligation of Endogenous Argonaute-bound RNAs) CLIP that enriches miRNAs ligated to their endogenous mRNA targets. CLEAR-CLIP mapped ~130,000 endogenous miRNA-target interactions in mouse brain and ~40,000 in human hepatoma cells. Motif and structural analysis define expanded pairing rules for over 200 mammalian miRNAs. Most interactions combine seed-based pairing with distinct, miRNA-specific patterns of auxiliary pairing. At some regulatory sites, this specificity confers distinct silencing functions to miRNA family members with shared seed sequences but divergent 3’ ends. This work provides a means for explicit biochemical identification of miRNA sites in vivo, leading to the discovery that miRNA 3’ end pairing is a general determinant of AGO binding specificity.

2015-11-15 | GSE73058 | GEO

miRNA-target chimeras identified with CLEAR-CLIP reveal miRNA 3' end pairing as a major determinant of Argonaute binding in vivo [human Huh7.5 AGO-CLIP & CLEAR-CLIP]

2015-11-15 | GSE73057 | GEO

MicroRNA 3'-compensatory pairing occurs through two binding modes, with affinity shaped by nucleotide identity and position [2]

Project description:MicroRNAs (miRNAs), in association with Argonaute (AGO) proteins, direct repression by pairing to sites within mRNAs. Compared to pairing preferences of the miRNA seed region (nucleotides 2–8), preferences of the miRNA 3′ region are poorly understood, due to the sparsity of measured affinities for the many pairing possibilities. We used RNA bind-n-seq with purified AGO2–miRNA complexes to measure relative affinities of >1,000 3′-pairing architectures for each miRNA. In some cases, optimal 3′ pairing increased affinity by >500-fold. Some miRNAs had two high-affinity 3′-pairing modes—one of which included additional nucleotides bridging seed and 3′ pairing to enable high-affinity pairing to miRNA nucleotide 11. The affinity of binding and the position of optimal pairing both tracked with the occurrence of G or oligo(G/C) nucleotides within the miRNA. These and other results advance understanding of miRNA targeting, providing insight into how optimal 3′ pairing is determined for each miRNA.

2022-03-31 | GSE196457 | GEO

MicroRNA 3'-compensatory pairing occurs through two binding modes, with affinity shaped by nucleotide identity and position [1]

2022-03-31 | GSE174715 | GEO

RAB protein controls Argonaute localization for proper miRNA function in C. elegans

Project description:MicroRNAs are critical regulators of gene expression in animals. To repress their target mRNAs, these small RNAs are first loaded onto Argonaute proteins to form the silencing complex called miRISC. The complex must then be transported to its target mRNA where it interacts mostly within the 3’UTR through base-pairing. After target repression, the constituents of the miRISC undergo degradation or recycling mediated through their accurate sorting and localization in the cell. While some reports have linked intracellular trafficking to miRNA activity, it is still unclear how these pathways coordinate to allow proper miRNA-mediated gene silencing and turnover. Through a forward genetic screen performed in the nematode Caenorhabditis elegans, we identified the RabGAP tbc-11 as an important factor for the miRNA pathway. We show that TBC-11 is likely a GAP for the small GTPase RAB-6 and that its regulation is required for miRNA function in animals. The absence of functional TBC-11 leads to the persistent activation of RAB-6, which increases the localization of miRNA-unloaded Argonaute ALG-1 on endomembranes. This inactive form of Argonaute accumulates on polysomes, leading to defective miRNA-mediated target mRNA repression. Together, our results establish the importance of intracellular trafficking for miRNA function and demonstrates the involvement of a small GTPase in proper Argonaute localization in vivo.

2021-03-16 | GSE141719 | GEO

Widespread microRNA degradation elements in target mRNAs can assist the encoded proteins

Project description:Binding of microRNAs (miRNA) to mRNAs normally results in post-transcriptional repression. However, extensive base-pairing between miRNAs and target RNAs can trigger miRNA degradation, a phenomenon called target RNA-directed miRNA degradation (TDMD). Here, we systematically analyzed Argonaute-CLASH (crosslinking, ligation, and sequencing of miRNA-target RNA hybrids) data, and identified numerous candidate TDMD triggers based on their ability to induce non-templated nucleotide addition at the miRNA 3 end. When overexpressed in HEK293T cells, nine highly conserved triggers induce degradation of corresponding miRNAs. Consistent with previous reports, both the TDMD base-pairing and surrounding sequences are important for TDMD. CRISPR knockout of endogenous trigger or ZSWIM8, a ubiquitin ligase essential for TDMD, reduce miRNA degradation. Furthermore, degradation of miR-221 and miR-222 by a trigger in BCL2L11, which encodes an apoptotic protein, enhances apoptosis. Therefore, we uncovered widespread TDMD triggers in target RNAs and demonstrated that they can functionally cooperate with the encoded proteins.

2024-06-25 | GSE178943 | GEO

Transcriptome-wide miR-155 binding map reveals widespread non-canonical microRNA targeting [CLIP-seq data]

Project description:microRNAs (miRNAs) are essential components of gene regulation, but identification of miRNA targets remains a major challenge. Most target prediction and discovery relies on perfect complementarity of the miRNA seed to the 3’ untranslated region (UTR). However, it is unclear to what extent miRNAs target sites without seed matches. Here, we performed a transcriptome-wide identification of the endogenous targets of a single miRNA—miR-155—in a genetically controlled manner. We found that approximately forty percent of miR-155-dependent Argonaute binding occurs at sites without perfect seed matches. The majority of these non-canonical sites feature extensive complementarity to the miRNA seed with one mismatch. These non-canonical sites confer regulation of gene expression albeit less potently than canonical sites. Thus, non-canonical miRNA binding sites are widespread, often contain seed-like motifs, and can regulate gene expression, generating a continuum of targeting and regulation. Argonaute (AGO) HITS-CLIP Libraries generated from wild type and miR-155 knockout activated T cells.

2012-11-09 | GSE41285 | GEO

The ZSWIM8 ubiquitin ligase mediates target-directed microRNA degradation [dataset 1]

Project description:MicroRNAs (miRNAs) associate with Argonaute (AGO) proteins to direct widespread post-transcriptional gene repression. Although association with AGO typically protects miRNAs from nucleases, extensive pairing to some unusual target RNAs can trigger miRNA degradation. Here we found that this target-directed miRNA degradation (TDMD) required the ZSWIM8 Cullin-RING E3 ubiquitin ligase. This and other findings suggested and supported a mechanistic model of TDMD in which target-directed proteolysis of AGO by the ubiquitin–proteasome pathway exposes the miRNA for degradation. Moreover, loss-of-function studies indicated that the ZSWIM8 Cullin-RING ligase accelerates degradation of numerous miRNAs in cells of mammals, flies, and nematodes, thereby specifying the half-lives of most short-lived miRNAs. These results elucidate the mechanism of TDMD and expand the inferred role of TDMD in shaping miRNA levels in bilaterian animals.

2020-12-18 | GSE163393 | GEO

The ZSWIM8 ubiquitin ligase mediates target-directed microRNA degradation [datatset 7]

2020-12-18 | GSE163392 | GEO