Genome-wide expression of mRNAs and microRNAs in response to ERI2 knockdown
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE100610 | GEO | 2020/06/01
REPOSITORIES: GEO
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Project description:MicroRNAs (miRNAs) are endogenous noncoding small RNAs with important roles in many biological pathways; their generation and activity are under precise regulation. Emerging evidence suggests that miRNA pathways are precisely modulated with controls at the level of transcription, processing, and stability, with miRNA deregulation linked with diseases and neurodegenerative disorders. In the Drosophila miRNA biogenesis pathway, long primary miRNA transcripts undergo sequential cleavage to release the embedded miRNAs. Mature miRNAs are then loaded into Argonaute1 (Ago1) within the RNA-induced silencing complex (RISC). Intriguingly, we found that Drosophila miR-34 displays multiple isoforms that differ at the 3′ end, suggesting a novel biogenesis mechanism involving 3′ end processing. To define the cellular factors responsible, we performed an RNA interference (RNAi) screen and identified a putative 3′→5′ exoribonuclease CG9247/nibbler essential for the generation of the smaller isoforms of miR-34. Nibbler (Nbr) interacts with Ago1 and processes miR-34 within RISC. Deep sequencing analysis revealed a larger set of multi-isoform miRNAs that are controlled by nibbler. These findings suggest that Nbr-mediated 3′ end processing represents a critical step in miRNA maturation that impacts miRNA diversity.
2011-11-17 | GSE31689 | GEO
Project description:MicroRNAs (miRNAs) are endogenous noncoding small RNAs with important roles in many biological pathways; their generation and activity are under precise regulation. Emerging evidence suggests that miRNA pathways are precisely modulated with controls at the level of transcription, processing, and stability, with miRNA deregulation linked with diseases and neurodegenerative disorders. In the Drosophila miRNA biogenesis pathway, long primary miRNA transcripts undergo sequential cleavage to release the embedded miRNAs. Mature miRNAs are then loaded into Argonaute1 (Ago1) within the RNA-induced silencing complex (RISC). Intriguingly, we found that Drosophila miR-34 displays multiple isoforms that differ at the 3M-bM-^@M-2 end, suggesting a novel biogenesis mechanism involving 3M-bM-^@M-2 end processing. To define the cellular factors responsible, we performed an RNA interference (RNAi) screen and identified a putative 3M-bM-^@M-2M-bM-^FM-^R5M-bM-^@M-2 exoribonuclease CG9247/nibbler essential for the generation of the smaller isoforms of miR-34. Nibbler (Nbr) interacts with Ago1 and processes miR-34 within RISC. Deep sequencing analysis revealed a larger set of multi-isoform miRNAs that are controlled by nibbler. These findings suggest that Nbr-mediated 3M-bM-^@M-2 end processing represents a critical step in miRNA maturation that impacts miRNA diversity. Total RNA was isolated using mirVana (Ambion) and size-fractionated by PAGE into 18-30nt. These were independently processed and sequenced using the Illumina GAII platform. In total, four libraries were analyzed.
2011-11-17 | E-GEOD-31689 | biostudies-arrayexpress
Project description:It has previously been assumed that the generally high stability of microRNAs (miRNAs) reflects their tight association with Argonaute (Ago) proteins, essential components of the RNA-induced silencing complex (RISC). However, recent data have suggested that the majority of mature miRNAs are not, in fact, Ago associated. Here, we demonstrate that endogenous human miRNAs vary widely, by >100-fold, in their level of RISC association and show that the level of Ago binding is a better indicator of inhibitory potential than is the total level of miRNA expression. While miRNAs of closely similar sequence showed comparable levels of RISC association in the same cell line, these varied between different cell types. Moreover, the level of RISC association could be modulated by overexpression of complementary target mRNAs. Together, these data indicate that the level of RISC association of a given endogenous miRNA is regulated by the available RNA targetome and predicts miRNA function.
2014-05-30 | GSE58127 | GEO
Project description:It has previously been assumed that the generally high stability of microRNAs (miRNAs) reflects their tight association with Argonaute (Ago) proteins, essential components of the RNA-induced silencing complex (RISC). However, recent data have suggested that the majority of mature miRNAs are not, in fact, Ago associated. Here, we demonstrate that endogenous human miRNAs vary widely, by >100-fold, in their level of RISC association and show that the level of Ago binding is a better indicator of inhibitory potential than is the total level of miRNA expression. While miRNAs of closely similar sequence showed comparable levels of RISC association in the same cell line, these varied between different cell types. Moreover, the level of RISC association could be modulated by overexpression of complementary target mRNAs. Together, these data indicate that the level of RISC association of a given endogenous miRNA is regulated by the available RNA targetome and predicts miRNA function. This series includes microRNA sequencing data for human cell lines 293, C8166, A549, SH-SY5Y, and an EBV-transformed LCL line. For each cell line, total small RNA isolated by TRIzol was sequenced for comparison to RISC-associated microRNAs as determined by sequencing small RNAs from an Argonaute immunoprecipitation.
2014-05-30 | E-GEOD-58127 | biostudies-arrayexpress
Project description:One of the major players controlling RNA decay is the cytoplasmic 5’ to 3’ exoribonuclease, which is conserved among eukaryotic organisms. In Arabidopsis, the 5’ to 3’ exoribonuclease XRN4 is involved in disease resistance, the response to ethylene, RNAi and miRNA-mediated RNA decay. Curiously, XRN4 appears to display selectivity among its substrates because certain 3’ cleavage products formed by miRNA-mediated decay, such as from ARF10 mRNA, accumulate in the xrn4 mutant, whereas others, such as from AGO1, do not. To examine the nature of this selectivity, transcripts that differentially accumulate in xrn4 were identified by combining PARE and Affymetrix arrays. Certain functional categories, such as stamen-associated proteins and hydrolases, were over-represented among transcripts decreased in xrn4, whereas transcripts encoding nuclear-encoded chloroplast-targeted proteins and nucleic acid-binding proteins were over-represented in transcripts increased in xrn4. To ascertain if RNA sequence influences the apparent XRN4 selectivity, a series of chimeric constructs were generated where the miRNA-complementary sites and different portions of the surrounding sequences from AGO1 and ARF10 were interchanged. Analysis of the resulting transgenic plants revealed that the presence of a 150 nucleotide sequence downstream of the ARF10 miRNA-complementary site conferred strong accumulation of the 3’ cleavage products in xrn4. In addition, sequence analysis of differentially accumulating transcripts led to the identification of 27 hexamer motifs that were over-represented in transcripts or miRNA-cleavage products accumulating in xrn4. Taken together, the data indicate that specific mRNA sequences, like those in ARF10, and mRNAs from select functional categories are attractive targets for XRN4-mediated decay. Mixed-stage inflorescences from Col-0 and xrn4-5 plants were harvested. Total RNA was extracted and hybridized to the ATH1 array to determine genes differentially expressed in xrn4-5 compared to Col-0. 2 replicates each.
2011-01-14 | E-GEOD-23027 | biostudies-arrayexpress
Project description:One of the major players controlling RNA decay is the cytoplasmic 5'-to-3' exoribonuclease, which is conserved among eukaryotic organisms. In Arabidopsis, the 5'-to-3' exoribonuclease XRN4 is involved in disease resistance, the response to ethylene, RNAi, and miRNA-mediated RNA decay. Curiously, XRN4 appears to display selectivity among its substrates because certain 3' cleavage products formed by miRNA-mediated decay, such as from ARF10 mRNA, accumulate in the xrn4 mutant, whereas others, such as from AGO1, do not. To examine the nature of this selectivity, transcripts that differentially accumulate in xrn4 were identified by combining PARE and Affymetrix arrays. Certain functional categories, such as stamen-associated proteins and hydrolases, were over-represented among transcripts decreased in xrn4, whereas transcripts encoding nuclear-encoded chloroplast-targeted proteins and nucleic acid-binding proteins were over-represented in transcripts increased in xrn4. To ascertain if RNA sequence influences the apparent XRN4 selectivity, a series of chimeric constructs was generated in which the miRNA-complementary sites and different portions of the surrounding sequences from AGO1 and ARF10 were interchanged. Analysis of the resulting transgenic plants revealed that the presence of a 150 nucleotide sequence downstream from the ARF10 miRNA-complementary site conferred strong accumulation of the 3' cleavage products in xrn4. In addition, sequence analysis of differentially accumulating transcripts led to the identification of 27 hexamer motifs that were over-represented in transcripts or miRNA-cleavage products accumulating in xrn4. Taken together, the data indicate that specific mRNA sequences, like those in ARF10, and mRNAs from select functional categories are attractive targets for XRN4-mediated decay.
2011-01-14 | GSE23027 | GEO
Project description:TRPM7 (transient receptor potential cation channel subfamily M member 7) is a chanzyme with channel and kinase domains essential for embryo development. Using gamete-specific Trpm7-null lines, we report that TRPM7-mediated Mg2+ influx is indispensable for reaching the blastocyst stage. TRPM7 was expressed dynamically from gametes to blastocysts, displaying stage-specific and distinct localizations on the plasma membrane, cytoplasm, and nucleus, and undergoes cleavage that produces C- terminal kinase fragments. TRPM7 underpinned Mg2+ homeostasis throughout this time, and excess Mg2+ but not Zn2+ or Ca2+ overcame the arrest of Trpm7-null embryos; expressing Trpm7 mRNA restored development, but mutant versions failed or were less effective. Transcriptomic analyses of embryos lacking Trpm7 revealed an abundance of oxidative stress-pathway genes, confirmed by mitochondrial dysfunction, and a reduction of transcription factor networks essential for proliferation; Mg2+ supplementation corrected these defects. Hence, TRPM7 underpins Mg2+ homeostasis in preimplantation embryos, prevents oxidative stress, and promotes gene expression patterns necessary for developmental progression and cell lineage specification.
2024-01-22 | GSE241487 | GEO
Project description:mRNA and microRNA expression was examined in global cellular fractions and in RNA-induced silencing complex (RISC)-immunoprecipitated cell fractions in cultured primary human astrocytes (ScienCell) and in cultured human U-87 MG astrocytoma cells (ATCC). ABSTRACT: Background: GW/P bodies are cytoplasmic ribonucleoprotein-rich foci that are involved in microRNA (miRNA)-mediated messenger RNA (mRNA) silencing and degradation. These mRNA regulatory functions within GW/P bodies are mediated by GW182 and its binding partner hAgo2 when bound to miRNA within the RNA-induced silencing complex (RISC). Although miRNAs and mRNAs are known to be localized to RISC in a variety of cells, to date no published study has examined the profile of specific miRNA and mRNA targeted to the RISC. Methodology/Principle Findings: In this study, RISC mRNA and miRNA components were profiled by microarray analysis of human U-87 astrocytoma cells and primary human astrocytes with total RNA extracted from the RISC as well as the global cellular fractions. The novel findings of this study were fourfold: (1) miRNAs are highly enriched in primary astrocyte RISC compared to U-87 astrocytoma RISC, (2) astrocytoma cells and primary astrocytes each contain unique RISC miRNA profiles as compared to their respective cellular miRNA profiles, (3) miR-195, 10b, 29b, 19b, 34a and 455-3p were upregulated and miR-181b was downregulated in U-87 astrocytoma RISC as compared to primary astrocyte RISC, and (4) RISC contain mostly downregulated mRNAs in primary astrocytes and U-87 astrocytoma cells. Conclusions/Significance: We show that in U-87 astrocytoma cells, miR-34a and miR-195 were upregulated in RISC suggesting an oncogenic role for these miRNAs. Three miR34a-targeted mRNAs and two miR-195-targeted mRNAs were downregulated. One miR-195-targeted mRNA was upregulated. Biological pathway analysis of RISC mRNA components suggests that the RISC plays a pivotal role in cancer, inflammatory disease, immunological disease, the cell cycle, cellular movement and numerous cell signaling pathways. This study points to the importance of the RISC and ultimately GW/P body composition and function and in miRNA and mRNA deregulation in astrocytoma cells and possibly for other brain tumors.
2011-05-26 | GSE21514 | GEO
Project description:3’ end methylation catalyzed by HUA ENHANCER1 (HEN1) is a crucial step of small RNA stabilization in plants, yet how unmethylated small RNAs undergo degradation remains largely unknown. Using a reverse genetic approach, we here show that ATRIMMER2 (ATRM2), a DEDDy-type 3’ to 5’ exoribonuclease, acts in the degradation of unmethylated miRNAs and miRNA*s in Arabidopsis. A loss-of-function mutations in ATRM2 partially suppress the morphological defects caused by HEN1 malfunction, with restored levels of a subset of miRNAs and receded expression of corresponding miRNA targets. Loss of ATRM2 has negligible effect on miRNA trimming, and further increase the fertility of hen1 heso1 urt1, a mutant with an almost complete abolishment of miRNA uridylation, indicating that ATRM2 may neither be involved in 3’ to 5’ trimming nor be the enzyme that specifically degrades uridylated miRNAs. Notably, the fold changes of miRNAs and their corresponding miRNA*s were significantly co-regulated in hen1 atrm2 as compared with those in hen1. Unexpectedly, we also observed a markedly increase of 3’ to 5’ trimming of several miRNA*s but not miRNAs in ATRM2 compromised backgrounds. These data suggest an action of ATRM2 on miRNA/miRNA* duplexes, and the existence of an unknown exoribonuclease for specific trimming of miRNA*. This asymmetric effect on miRNA/miRNA* is likely related to ARGONAUTE (AGO) proteins, which can distinguish miRNAs from miRNA*s. Finally, we show that ATRM2 colocalizes and physically interacts with ARGONAUTE1 (AGO1). Taken together, our results suggest that ATRM2 may be involved in the surveillance of unmethylated miRNA/miRNA* duplexes during the initiation step of RISC assembly.
2018-12-01 | GSE107070 | GEO
Project description:Messenger RNAs are regulated by a variety of degradation mechanisms in mammalian cells. In the canonical animal microRNA pathway, microRNAs in complex with Argonaute proteins bind to many mRNA targets with imperfect complementarity, leading to degradation of the mRNA through the regular decay machinery. The ancestral “slicer” endonuclease activity of Argonaute2 itself, which requires more extensive complementarity with the target RNA, is not used in this pathway, and has only been observed in two microRNA-guided cases. Nevertheless, the cleavage capacity of mammalian Ago2 is conserved and essential for viability. Here, we assess the endonucleolytic function of Ago2 and other nucleases by identifying cleavage products retaining 5`-phosphate groups in mouse ES cells on a transcriptome-wide scale. We detect a significant signature of Ago2-dependent cleavage events and validate several targets. Unexpectedly, a broader class of Ago2-independent cleavage sites is also observed, indicating participation of additional nucleases in this mode of mRNA regulation. Within this class, we identify a cohort of Drosha-dependent mRNA cleavage events, including one in the Dgcr8 mRNA, that functionally regulate mRNA levels in mES cells. Together, these results highlight the underappreciated role of endonucleolytic cleavage in controlling mRNA fates in mammals. Global 5`-phosphate-dependent RACE in WT, Ago2-KO and Drosha-excised mouse ES cells and human 293S cells
2010-06-25 | E-GEOD-21975 | biostudies-arrayexpress