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Convergence of distinct RNA-silencing pathways on GW182/Tnrc6 [sgRNA screen]

ABSTRACT: sgRNA screens for factors influencing Trim71 silencing activity.

ORGANISM(S): Mus musculus

PROVIDER: GSE214708 | GEO | 2023/06/26

REPOSITORIES: GEO

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Convergence of distinct RNA-silencing pathways on GW182/Tnrc6

Project description:The RNA-binding protein Trim71/Lin41 is a phylogenetically conserved developmental regulator that functions in mammalian stem cell reprogramming, brain development and cancer. Trim71 recognizes target mRNAs through hairpin motifs and silences them through molecular mechanisms that await identification. Deposited data are mass spectrometry data from immunoprecipitation experiments with endogenously tagged Trim71, Ago2, and Tnrc6a. Experiments are carried out in mouse embryonic stem cells (mESCs) and uncover reciprocal interactions of Trim71, Ago2, and Tnrc6a. Trim71 protein interactions are largely independent of Ago2 levels, but strongly depend on the presence of RNA. A second set of experiments is mass spectrometry data from immunoprecipitation experiments in mESCs overexpressing a tagged peptide derived from Homo sapiens TNRC6B (‘FLAG-HA::T6BWT::Cherry’), that is known to block the Tnrc6-Ago2 interaction. We show that a wild-type version strongly binds Ago1 and Ago2, but not Trim71, while a mutant version binds neither Ago1, 2, nor Trim71.

2023-07-04 | PXD034537 | Pride

Convergence of distinct RNA-silencing pathways on GW182/Tnrc6 II

Project description:RNA-sequencing in mouse embryonic stem cells: Ctrl: GFP_5xT71BS (mES [129S6/SvEvTac], yGlob [GFP_5xT71BS]. Trim71-/-: GFP_5xT71BS (mES [129S6/SvEvTac], yGlob [GFP_5xT71BS] Trim71 -/-. Note pervasive gene expression changes in samples with TNRC6A overexpression in comparison with samples with CAS9 overexpression.

2024-02-13 | GSE231730 | GEO

Convergence of distinct RNA-silencing pathways on GW182/Tnrc6 [RNA-seq]

Project description:RNA-sequencing in mouse embryonic stem cells. Experiment 1 (Samples 1-9): Base cell line (WT): GFP_5xT71BS (mES [129S6/SvEvTac], yGlob [GFP_5xT71BS], stable transfection FLAG-Cas9::T2A::Blasticidin). Trim71 KO: GFP_5xT71BS Trim71-/-. Ago2 KO: GFP_5xT71BS Ago2-/-. Experiment 2 (Samples 10-21): parental cell line: GFP_5xT71BS. T6B_Wt: Overexpression of T6B_WT, T6B_Mut: Overexpression of T6B_MUT. Trim71_KO: GFP_5xT71BS, Trim71-/- transfected with empty plasmid. Ago2_KO: GFP_5xT71BS, Ago2-/- transfected with empty plasmid.

2023-06-26 | GSE205967 | GEO

Convergence of distinct RNA-silencing pathways on GW182/Tnrc6

2023-07-04 | PXD041242 | Pride

Convergence of distinct RNA-silencing pathways on GW182/Tnrc6 [RIP-seq]

Project description:RIP-sequencing in mouse embryonic stem cells. Experiment 1 (Samples 1-18): Base cell line (untagged control cells): GFP_5xT71BS (mES [129S6/SvEvTac], yGlob [GFP_5xT71BS], stable transfection FLAG-Cas9::T2A::Blasticidin). HA-Tnrc6a: endogenously HA-AVI-tagged Tnrc6a. HA-Tnrc6a_Trim71KO: HA-AVI-tagged Tnrc6a, Trim71-/-. Experiment 2 (Samples 19-42): Ctrl: base cell line GFP_5xT71BS. Tnrc6a: endogenously HA-AVI-tagged Tnrc6a. Tnrc6a_Trim71KO: HA-AVI-tagged Tnrc6a, Trim71-/-. Tnrc6a_Ago2KO: HA-AVI-tagged Tnrc6a, Ago2-/-.

2023-06-26 | GSE205966 | GEO

Convergence of distinct RNA-silencing pathways on GW182/Tnrc6 [RIP-seq II]

Project description:RIP-sequencing in mouse embryonic stem cells. Experiment 3 (Samples 2-24) & 4 (Samples 25-48): Base cell line (untagged control cells): GFP_5xT71BS (mES [129S6/SvEvTac], yGlob [GFP_5xT71BS], stable transfection FLAG-Cas9::T2A::Blasticidin). HA-Tnrc6a: endogenously HA-AVI-tagged Tnrc6a. HA-Tnrc6a_Trim71KO: HA-AVI-tagged Tnrc6a, Trim71-/-. Experiment 2 (Samples 19-42): Ctrl: base cell line GFP_5xT71BS. Tnrc6a: endogenously HA-AVI-tagged Tnrc6a. Tnrc6a_Trim71KO: HA-AVI-tagged Tnrc6a, Trim71-/-. Tnrc6a_Ago2KO: HA-AVI-tagged Tnrc6a, Ago2-/-.

2024-02-13 | GSE214755 | GEO

Convergence of distinct RNA-silencing pathways on GW182/Tnrc6

Project description:This SuperSeries is composed of the SubSeries listed below.

2023-06-26 | GSE205968 | GEO

CRISPR activation screen identifies BCL-2 proteins and B3GNT2 as drivers of cancer resistance to T cell-mediated cytotoxicity (sgRNA distribution)

Project description:SgRNA library distributions of CRISPR activation screens for tumor immune evasion

2022-02-24 | GSE197309 | GEO

Expression of TNRC6 (GW182) Proteins Is Not Necessary for Gene Silencing by Fully Complementary RNA Duplexes.

Project description:The trinucleotide repeat containing 6 (TNRC6) family of proteins are core components of RNA interference (RNAi) and consist of three paralogs (TNRC6A, TNRC6B, and TNRC6C). The TNRC6 paralogs associate with argonaute (AGO) protein, the core RNAi factor, and bridge its interactions with other proteins. We obtained TNRC6A and TNRC6B single and double knockout cell lines to investigate how the TNRC6 paralogs contribute to RNAi. We found that TNRC6 proteins are not required for gene silencing when duplex RNAs are fully complementary. TNRC6 expression was necessary for regulation by a microRNA. TNRC6A, but not TNRC6B, expression was necessary for transcriptional activation by a duplex RNA targeting a gene promoter. By contrast, AGO2 is required for all three gene expression pathways. TNRC6A can affect the Dicer localization in cytoplasm versus the nucleus, but none of the three TNRC6 paralogs was necessary for nuclear localization of AGO2. Our data suggest that the roles of the TNRC6 paralogs differ in some details and that TNRC6 is not required for clinical therapeutic silencing mechanisms that involve fully complementary duplex RNAs.

| S-EPMC6885777 | biostudies-literature

A screen for cis-regulatory elements for murine miR-142 using an sgRNA library prepared by Molecular Chipper technology

Project description:CRISPR-based loss-of-function screens have been proven powerful to identify genetic regulators in mammalian cells, but current approaches for single guide RNA (sgRNA) library construction are expensive and difficult to be adapted in most laboratories. Here, we present a Molecular Chipper technology for inexpensive and easily customizable sgRNA library generation, and a proof-of-principle screen that identifies novel cis-regulatory regions for miR-142 biogenesis. This method will be useful for functional interrogation of non-coding elements in mammalian genomes

2016-03-17 | GSE70011 | GEO

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