ADAR2 affects mRNA coding sequence edits but not gene expression or splicing in vivo
Ontology highlight
ABSTRACT:
ORGANISM(S): Mus musculus
SUBMITTER: Allissa Dillman
PROVIDER: E-GEOD-70588 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:Adenosine deaminases (ADARs) are RNA binding proteins that bind to double stranded RNA and convert adenosine to inosine. Editing creates multiple isoforms of neurotransmitter receptors, including AMPA-subtype Glutamate channels such as Gria2 that is edited to introduce a Q to R amino acid change. Adar2 knock out mice die of seizures shortly after birth, but if the Gria2 Q/R editing site is mutated to mimic the edited version then the animals are viable. We performed RNA-Seq on the frontal cortices of Adar2-/- Gria2R/R mice and their Adar2+/+ Gria2R/R littermates, quantifying overall gene expression, splicing, and A to I editing in a transcriptome-wide fashion. We found 56 editing sites whose level of editing was significantly diminished in the Adar2 deficient animals. The majority of Adar2 responsive editing sites were in the coding regions of genes. Interestingly, other than Adar2 expression, there were only two additional statistically significant differentially expressed genes, Flnb and Cdh13. There were also only three exons that showed statistically significant differences in expression levels between the two genotypes. This work illustrates that ADAR2 is important in site-specific changes of protein coding sequences but has only modest effects on gene expression or splicing in vivo.
2016-01-01 | GSE70588 | GEO
Project description:Background: Adenosine deaminases that act on RNA (ADARs) bind to double-stranded and structured RNAs and deaminate adenosines to inosines. This A to I editing is widespread and required for normal life and development. Besides mRNAs and repetitive elements, ADARs can target miRNA precursors. Editing of miRNA precursors can affect processing efficiency and alter target specificity. Interestingly, ADARs can also influence miRNA abundance independent of RNA-editing. In mouse embryos where editing levels are low, ADAR2 was found to be the major ADAR protein that affects miRNA abundance. Here we extend our analysis to adult mouse brains where high editing levels are observed. Results: Using Illumina deep sequencing we compare the abundances of mature miRNAs and editing events within them, between wild-type and ADAR2 knockout mice in the adult mouse brain. Reproducible changes in abundance of specific miRNAs are observed in ADAR2 deficient mice. Most of these quantitative changes seem unrelated to A to I editing events. However, many A to G transitions in cDNAs prepared from mature miRNA sequences, reflecting A to I editing events in the RNA, are observed with frequencies reaching up to 80%. About half of these editing events are primarily caused by ADAR2 while a few miRNAs show increased editing in the absence of ADAR2, suggesting preferential editing by ADAR1. Moreover, novel, previously unknown editing events were identified in several miRNAs. In general 64% of all editing events are located within the seed region of mature miRNAs. In one of these cases retargeting of the edited miRNA could be verified in reporter assays. Also, altered processing efficiency upon editing near a processing site could be experimentally verified. Conclusions: ADAR2 can significantly influence the abundance of certain miRNAs in the brain. Only in a few cases changes in miRNA abundance can be explained by miRNA editing. Thus, ADAR2 binding to miRNA precursors, without editing them, may influence their processing and thereby abundance. ADAR1 and ADAR2 have both overlapping and distinct specificities for editing of miRNA editing sites. Over 60% of editing occurs in the seed region possibly changing target specificities for many edited miRNAs. Examination of the effect of ADAR2 on mature miRNA abundance and sequence in adult mouse brain.
2014-10-02 | E-GEOD-61066 | biostudies-arrayexpress
Project description:Background: Adenosine deaminases that act on RNA (ADARs) bind to double-stranded and structured RNAs and deaminate adenosines to inosines. This A to I editing is widespread and required for normal life and development. Besides mRNAs and repetitive elements, ADARs can target miRNA precursors. Editing of miRNA precursors can affect processing efficiency and alter target specificity. Interestingly, ADARs can also influence miRNA abundance independent of RNA-editing. In mouse embryos where editing levels are low, ADAR2 was found to be the major ADAR protein that affects miRNA abundance. Here we extend our analysis to adult mouse brains where high editing levels are observed. Results: Using Illumina deep sequencing we compare the abundances of mature miRNAs and editing events within them, between wild-type and ADAR2 knockout mice in the adult mouse brain. Reproducible changes in abundance of specific miRNAs are observed in ADAR2 deficient mice. Most of these quantitative changes seem unrelated to A to I editing events. However, many A to G transitions in cDNAs prepared from mature miRNA sequences, reflecting A to I editing events in the RNA, are observed with frequencies reaching up to 80%. About half of these editing events are primarily caused by ADAR2 while a few miRNAs show increased editing in the absence of ADAR2, suggesting preferential editing by ADAR1. Moreover, novel, previously unknown editing events were identified in several miRNAs. In general 64% of all editing events are located within the seed region of mature miRNAs. In one of these cases retargeting of the edited miRNA could be verified in reporter assays. Also, altered processing efficiency upon editing near a processing site could be experimentally verified. Conclusions: ADAR2 can significantly influence the abundance of certain miRNAs in the brain. Only in a few cases changes in miRNA abundance can be explained by miRNA editing. Thus, ADAR2 binding to miRNA precursors, without editing them, may influence their processing and thereby abundance. ADAR1 and ADAR2 have both overlapping and distinct specificities for editing of miRNA editing sites. Over 60% of editing occurs in the seed region possibly changing target specificities for many edited miRNAs. Examination of the effect of ADAR2 on gene expression in mature mouse wildtype and ADAR2 knockout brain using AffymetrixM-BM-. GeneChipM-BM-. Whole Transcript (WT) Expression Arrays (Analysis by KFB Regensburg, Germany)
2014-10-02 | E-GEOD-61067 | biostudies-arrayexpress
Project description:We used transgenic mouse embryos that are deficient in the two enzymatically active RNA editing enzymes ADAR1 and ADAR2 to compare relative frequencies but also sequence composition of mature miRNAs in these genetically modified backgrounds to wild-type mice by Illumina next gen sequencing. Deficiency of ADAR2 leads to a reproducible change in abundance of specific miRNAs and their predicted targets. Changes in miRNA abundance seem unrelated to editing events. Additional deletion of ADAR1 has surprisingly little impact on the mature miRNA repertoire, indicating that miRNA expression is primarily dependent on ADAR2. A to G transitions reflecting A to I editing events can be detected at few sites and at low frequency during the early embryonic stage investigated. Again, most editing events are ADAR2 dependent with only few editing sites being specifically edited by ADAR1. Besides known editing events in miRNAs a few novel, previously unknown editing events were identified. Some editing events are located to the seed region of miRNAs opening the possibility that editing leads to their retargeting. GSM852140-8: sequencing of mature miRNAs of wt, ADAR2-/- and ADAR1-/-/ADAR2-/- female mouse embryos at E11.5 GSM863778-81: Gene expression was measured in wiltype, ADAR2-/- and ADAR1-/-/ADAR2-/- E11.5 whole female mouse embryos using Agilent Whole Mouse Genome Oligo Microarrays 8x60K.
2012-02-02 | E-GEOD-34626 | biostudies-arrayexpress
Project description:Adenosine deaminases, RNA specific (ADAR) are proteins that deaminate adenosine to inosine which is then recognized in translation as guanosine. To study the roles of ADAR proteins in RNA editing and gene regulation, we carried out DNA and RNA sequencing, RNA interference and RNA-immunoprecipitation in human B-cells. We also characterized the ADAR protein complex by mass spectrometry. The results uncovered over 60,000 sites where the adenosines (A) are edited to guanosine (G) and several thousand genes whose expression levels are influenced by ADAR. We also identified more than 100 proteins in the ADAR protein complex; these include splicing factors, heterogeneous ribonucleoproteins and several members of the dynactin protein family. Our findings show that in human B-cells, ADAR proteins are involved in two independent functions: A-to-G editing and gene expression regulation. In addition, we showed that other types of RNA-DNA sequence differences are not mediated by ADAR proteins, and thus there are co- or post-transcriptional mechanisms yet to be determined. Here we studied human B-cells where ADAR proteins (ADAR1 and ADAR2) are expressed but APOBECs are not. We identified the sequence differences between DNA and the corresponding RNA in B-cells from two individuals. Then, we carried out RNA interference, RNA-immunoprecipitation and next generation sequencing to determine the contribution of ADAR proteins in mediating A-to-G editing and other types of RNA-DNA sequence differences.
2013-09-20 | E-GEOD-38233 | biostudies-arrayexpress
Project description:Background: Adenosine deaminases that act on RNA (ADARs) bind to double-stranded and structured RNAs and deaminate adenosines to inosines. This A to I editing is widespread and required for normal life and development. Besides mRNAs and repetitive elements, ADARs can target miRNA precursors. Editing of miRNA precursors can affect processing efficiency and alter target specificity. Interestingly, ADARs can also influence miRNA abundance independent of RNA-editing. In mouse embryos where editing levels are low, ADAR2 was found to be the major ADAR protein that affects miRNA abundance. Here we extend our analysis to adult mouse brains where high editing levels are observed. Results: Using Illumina deep sequencing we compare the abundances of mature miRNAs and editing events within them, between wild-type and ADAR2 knockout mice in the adult mouse brain. Reproducible changes in abundance of specific miRNAs are observed in ADAR2 deficient mice. Most of these quantitative changes seem unrelated to A to I editing events. However, many A to G transitions in cDNAs prepared from mature miRNA sequences, reflecting A to I editing events in the RNA, are observed with frequencies reaching up to 80%. About half of these editing events are primarily caused by ADAR2 while a few miRNAs show increased editing in the absence of ADAR2, suggesting preferential editing by ADAR1. Moreover, novel, previously unknown editing events were identified in several miRNAs. In general 64% of all editing events are located within the seed region of mature miRNAs. In one of these cases retargeting of the edited miRNA could be verified in reporter assays. Also, altered processing efficiency upon editing near a processing site could be experimentally verified. Conclusions: ADAR2 can significantly influence the abundance of certain miRNAs in the brain. Only in a few cases changes in miRNA abundance can be explained by miRNA editing. Thus, ADAR2 binding to miRNA precursors, without editing them, may influence their processing and thereby abundance. ADAR1 and ADAR2 have both overlapping and distinct specificities for editing of miRNA editing sites. Over 60% of editing occurs in the seed region possibly changing target specificities for many edited miRNAs.
2014-10-02 | GSE61067 | GEO
Project description:Background: Adenosine deaminases that act on RNA (ADARs) bind to double-stranded and structured RNAs and deaminate adenosines to inosines. This A to I editing is widespread and required for normal life and development. Besides mRNAs and repetitive elements, ADARs can target miRNA precursors. Editing of miRNA precursors can affect processing efficiency and alter target specificity. Interestingly, ADARs can also influence miRNA abundance independent of RNA-editing. In mouse embryos where editing levels are low, ADAR2 was found to be the major ADAR protein that affects miRNA abundance. Here we extend our analysis to adult mouse brains where high editing levels are observed. Results: Using Illumina deep sequencing we compare the abundances of mature miRNAs and editing events within them, between wild-type and ADAR2 knockout mice in the adult mouse brain. Reproducible changes in abundance of specific miRNAs are observed in ADAR2 deficient mice. Most of these quantitative changes seem unrelated to A to I editing events. However, many A to G transitions in cDNAs prepared from mature miRNA sequences, reflecting A to I editing events in the RNA, are observed with frequencies reaching up to 80%. About half of these editing events are primarily caused by ADAR2 while a few miRNAs show increased editing in the absence of ADAR2, suggesting preferential editing by ADAR1. Moreover, novel, previously unknown editing events were identified in several miRNAs. In general 64% of all editing events are located within the seed region of mature miRNAs. In one of these cases retargeting of the edited miRNA could be verified in reporter assays. Also, altered processing efficiency upon editing near a processing site could be experimentally verified. Conclusions: ADAR2 can significantly influence the abundance of certain miRNAs in the brain. Only in a few cases changes in miRNA abundance can be explained by miRNA editing. Thus, ADAR2 binding to miRNA precursors, without editing them, may influence their processing and thereby abundance. ADAR1 and ADAR2 have both overlapping and distinct specificities for editing of miRNA editing sites. Over 60% of editing occurs in the seed region possibly changing target specificities for many edited miRNAs.
2014-10-02 | GSE61066 | GEO
Project description:Adenosine DeAminases acting on RNA (ADAR) catalyzes adenosine-to-inosine (A-to-I) conversion within RNA duplex structures. While A-to-I editing is often dynamically regulated in a spatial-temporal manner, the mechanisms underlying its tissue-selective restriction remain elusive. We have previously reported that transcripts of voltage-gated calcium channel CaV1.3 are subject to brain-selective A-to-I RNA editing by ADAR2. Here, we show that editing of CaV1.3 mRNA is dependent on a 40 bp RNA duplex formed between exon 41 and an evolutionarily conserved editing site complementary sequence (ECS) located within the preceding intron. Heterologous expression of a mouse minigene that contained the ECS, intermediate intronic sequence and exon 41 with ADAR2 yielded robust editing. Interestingly, editing of CaV1.3 was potently inhibited by serine/arginine-rich splicing factor 9 (SRSF9). Mechanistically, the inhibitory effect of SRSF9 required direct RNA interaction. Selective down-regulation of SRSF9 in neurons provides a basis for the neuron-specific editing of CaV1.3 transcripts.
2018-04-17 | GSE113193 | GEO
Project description:Adenosine to inosine (A-to-I) RNA editing, which is catalyzed by adenosine deaminases acting on RNA (ADAR) family of enzymes ADAR1 and ADAR2, has been shown to contribute to multiple cancers. However, other than chronic myeloid leukemia (CML) blast crisis, relatively little is known about its role in other types of hematological malignancies. Here, we found that ADAR2, but not ADAR1 and ADAR3, was specifically downregulated in the core binding factor (CBF) AML with t(8;21) or inv(16) translocations. In t(8;21) AML, RUNX1-driven transcription of ADAR2 was repressed by the RUNX1-ETO AE9a fusion protein in a dominant negative manner. Further functional studies confirmed that ADAR2 could suppress leukemogenesis specifically in t(8;21) and inv16 AML cells dependent on its RNA editing capability. Expression of two exemplary ADAR2-regulated RNA editing targets COPA and COG3 inhibited clonogenic growth of human t(8;21) AML cells. Our findings support a hitherto unappreciated mechanism leading to ADAR2 dysregulation in CBF AML and highlight the functional relevance of loss of ADAR2-mediated RNA editing to CBF AML.
2023-04-01 | GSE220488 | GEO
Project description:ADARs, short for adenosine deaminases acting on RNA, were first found by their characteristic activity of deaminating up to 50% of all adenosines in double-stranded RNA in vitro. C. elegans has two ADARs, only one is found in the fly, and mammals express three ADARs. Importantly, the spectrum of physiological roles of these enzymes in the different organisms remains largely unknown. ADARs are ~750-1150 amino acids in size, feature 2-3 N-terminal double-stranded RNA binding motifs, an enzymatic domain related to that of prokaryotic cytidine deaminases, and a ~190 residue extended C-terminal domain of unknown function. The mammalian ADARs are detectable in many if not all tissues, with relatively prominent expression in the central nervous system, predicting tissue- or cell type-specific tasks for the mammalian ADARs. Studies over the last decade collectively indicated that one function of ADARs is the editing of nuclear transcripts to introduce select amino acid substitutions in proteins. Arguably the best-studied example for this type of RNA editing (‘A-to-I editing’) in the mammal is performed by ADAR2 and occurs in primary transcripts for the GluA2 subunit (also known as GluR-B or GluR2 subunit) of AMPA receptors mediating fast excitatory neurotransmission. A particular glutamine codon (CAG) in the exonic sequence for the inner lining of the glutamate-activated AMPA receptor is converted by deamination of the central adenosine into an unusual arginine codon (CIG; the resultant inosine is seen as guanosine by the protein translation machinery). This substitution of the gene-encoded glutamine codon by the edited arginine codon is found in nearly 100% of all GluA2 cDNA derived from brain mRNA. RNA editing in this instance ensures that GluA2-containing AMPA receptors become impermeable to Ca2+. In addition to switching a key functional determinant in GluA2 to nearly 100%, ADAR2 has been implicated in generating numerous amino acid substitutions at lower levels in various mammalian neurotransmitter receptors and voltage-gated ion channels, primarily but not exclusively of nervous tissue. Most of these substitutions elicit distinct functional consequences, which led to the notion that site-selective RNA editing by ADAR2 may ensure the functional fine-tuning of the affected transmembrane proteins. Such fine-tuning should reflect the expansion of the protein sequence directed by the respective gene into sequence subpopulations, each differing in particular functional aspects, such as transmitter efficacy, desensitization, or voltage threshold. If this notion is correct, ADAR2 knockout should elicit phenotypes, which might range from subtle to severe, depending on the physiological role(s) of the ADAR2 targets in different tissues. ADAR2 knockout mice, however, die soon after birth from severe epileptic seizures, a consequence of the changed ion conductance properties of AMPA receptors containing unedited GluA2. Fortunately, mice lacking ADAR2 are robust and have normal life span if they carry GluA2 alleles genetically altered to contain the particular arginine codon for a Ca2+ impermeable ion channel pore of AMPA receptors, thus rescinding the need for ADAR2-mediated GluA2 transcript editing. These mice are thus eminently suitable to study the phenotypic consequences of failure to edit all ADAR2 targets excepting GluA2. We therefore tested cohorts of ADAR2-/-/GluA2(R)/(R) mice in 16 well-established paradigms for central and peripheral phenotypic abnormalities. We report that we found few differences to control mice (GluA2(R)/(R)) and hence, the lack of ADAR2 appears not to affect most of the physiological functions tested. Brain of four mutant animals of ADAR-/-/GluR-BR/R versus a pool of four reference (wildtype) mice; two technical replicates for each mutant mouse including a dye swap experiment - in total 8 chip hybridisations
2010-11-01 | E-GEOD-15383 | biostudies-arrayexpress