Structure and degradation of circular RNAs regulate PKR activation in innate immunity
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA507020 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR8245150_1.fastq.gz | Fastqsanger.gz | |||
| SRR8245150_2.fastq.gz | Fastqsanger.gz | |||
| SRR8245160_1.fastq.gz | Fastqsanger.gz | |||
| SRR8245160_2.fastq.gz | Fastqsanger.gz | |||
| SRR8245151_1.fastq.gz | Fastqsanger.gz |
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Project description:RNase L–mediated rapid degradation of circRNAs is required for PKR activation in innate immune responses and linked to the autoimmune disease systemic lupus erythematosus.
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Project description:Exon(s) back-splicing-generated circular RNAs as a group can suppress the double-stranded RNA (dsRNA)-activated protein kinase R (PKR) in cells. We have sought to synthesize immunogenicity-free, short dsRNA-containing circular RNAs as potent PKR inhibitors. Here, we report that circular RNAs synthesized by permuted td introns from T4 bacteriophage and by Anabeana pre-tRNA group I intron induce an immune response. Mechanistically, autocatalytic splicing introduces extra ~74 nt and ~186 nt fragments (E1+E2+spacer) that form additional stable loop structures that likely provoke innate immune responses in circular RNA products. In contrast, circular RNAs produced by T4 RNA ligase without unwanted fragment exhibit little immunogenicity. Importantly, directly-ligated circular RNAs form short dsRNA-regions exhibit 103~106 fold higher efficiency than the reported chemical compounds C16 and 2-AP in suppressing PKR activation, highlighting the use of circular RNAs as novel inhibitors for PKR over-reaction diseases.
2021-12-23 | GSE183941 | GEO
Project description:Protein kinase RNA-activated (PKR) induces immune response by sensing viral double-stranded RNAs (dsRNAs). However, growing evidence suggests that PKR can also be activated by endogenously expressed dsRNAs. Here, we capture these dsRNAs by formaldehyde-mediated crosslinking and immunoprecipitation-sequencing and find that various noncoding RNAs interact with PKR. Surprisingly, the majority of the PKR-interacting RNA repertoire is occupied by mitochondrial RNAs (mtRNAs). MtRNAs can form intermolecular dsRNAs owing to bidirectional transcription of mitochondrial genome and regulate PKR and eIF2α phosphorylation to control cell signaling and translation. Moreover, PKR activation by mtRNAs is counteracted by PKR phosphatases, disruption of which causes apoptosis from PKR overactivation even in uninfected cells. Our work unveils dynamic regulation of PKR even without infection and establishes PKR as a sensor for nuclear and mitochondrial signaling cues in regulating cellular metabolism.
2018-07-18 | GSE108986 | GEO
Project description:Efforts to advance RNA aptamers as a novel therapeutic modality have been limited by their susceptibilty to degradation and immunogenicity. In a previous study, we demonstrated synthesized double-stranded circular RNAs (ds-cRNAs) with minimal immunogenicity targeted to dsRNA-activated Protein Kinase R (PKR). Here, we test the therapeutic potential of ds-cRNAs in a mouse model of imiquimod-induced psoriasis. We find that genetic supplementation of ds-cRNAs leads to inhibition of PKR, resulting in alleviation of downstream interferon alpha (IFNα)/dsRNA signals and attenuation of psoriasis phenotypes. Delivery of ds-cRNAs by lipid nanoparticles to the spleen attenuates PKR activity in examined splenocytes, resulting in reduced epidermal thickness. These findings suggest that ds-cRNAs represent a promising approach to mitigate excessive PKR activation for therapeutic purposes.
2024-03-04 | GSE248680 | GEO
Project description:Protein kinase R (PKR), an innate immune sensor for double-stranded RNA (dsRNA), is critical for antiviral defense, but its aberrant activation by cellular dsRNA is linked to various diseases. The dsRNA-binding protein PACT plays a crucial and controversial role in the PKR pathway. We demonstrate that PACT is an essential negative regulator of PKR against endogenous dsRNA ligands like inverted repeat Alu RNAs, which satisfy PKR’s selectivity for long dsRNA and robustly activate PKR in the absence of PACT. PACT employs an unusual inhibitory mechanism sensitive to dsRNA length and concentration, inhibiting PKR through low-affinity interactions most effective when both are bound to the same dsRNA molecule. Consequently, PACT’s inhibition of PKR is more robust when dsRNA is longer and less abundant, with minimal effect on abundant or short dsRNA. Thus, PACT functions as a rheostat, setting the PKR activation threshold for long endogenous dsRNA without altering its inherent activity.
2024-06-16 | GSE269684 | GEO
Project description:Circular RNAs (circRNAs) have emerged as a new type of endogenous non-coding RNA characterized by their covalently closed circular structure and a backspliced junction (BSJ) resulting from a back-splicing process. In the last years, several studies have shown that circRNAs are important modulators in the immune system, activation of inflammation, antibacterial and antiviral responses as well as autoimmune diseases, such as multiple sclerosis. In the last years, it has been proposed that a subset of circRNA form imperfect double-stranded structures that allow them to interact with PKR (double stranded RNA-activated protein kinase), an intracellular protein responsible for recognizing long double stranded RNAs, usually viral RNAs, in the cytoplasm and trigger an anti-viral response of the cell. Taking into account the implication of the innate immune response in multiple sclerosis pathophysiology and the discovery of circRNA deregulation in multiple sclerosis patients, we wanted to investigate the circRNA-PKR crosstalk and its implication in the pathology of multiple sclerosis. Using an in vitro model of the anti-viral response and RNA-seq to characterize circRNA expression, we observed that poly (I:C) (a double-stranded RNA) stimulation produces a global downregulation of circRNA, PKR phosphorylation and immune-related gene activation and that these effects are reversed when treatment is stopped. Additionally, we found that the overexpression of double stranded RNA-containing circRNAs is able to reduce the PKR phosphorylation and the activation of immune-related pathways induced by poly(I:C). Finally, we measured the PKR phosphorylation in PBMCs isolated from multiple sclerosis patients in relapse and remission phases, but we did not obtain conclusive results since there is a very big variability between samples. In conclusion, this work reinforces the evidence on the mechanism describing the interaction between PKR and circRNA containing double-stranded structures and their role in the immune response. Besides, our experiments highlight the importance of the structure in circRNA function. Lastly, it also sheds some light into the possible implication of the circRNA-PKR axis in multiple sclerosis although further research is needed to make a better characterization of this mechanism in order to understand its potential role in the regulation of the immune response of the disease.
2024-06-01 | GSE253443 | GEO
Project description:We optimized the SHAPE-MaP (selective 2'-hydroxyl acylation analyzed by primer extension and mutational profiling) assay to analyze structures of paired circular (circSHAPE-MaP) and linear (linearSHAPE-MaP) RNAs.
2019-03-20 | GSE126195 | GEO
Project description:In platelets, splicing and translation occur in the absence of a nucleus. However, the integrity and stability of mRNAs derived from megakaryocyte progenitor cells remain poorly quantified on a transcriptome-wide level. As circular RNAs (circRNAs) are resistant to degradation by exonucleases, their abundance relative to linear RNAs can be used as a surrogate marker for mRNA stability in the absence of transcription. Here we show that circRNAs are enriched in human platelets 17-188 fold relative to nucleated tissues, and 14-26 fold relative to samples digested with RNAseR to selectively remove linear RNA. We compare RNAseq read depths inside and outside circRNAs to provide in silico evidence of transcript circularity, show that exons within circRNAs are enriched ~13X in platelets relative to nucleated tissues, and identify 3162 genes significantly enriched for circRNAs including some where all RNAs appear to be derived from circular molecules. We also confirm that this is a feature of other anucleate cells through transcriptome sequencing of mature erythrocytes, demonstrate that circRNAs are not enriched in megakaryocytes, and that linear RNAs decay more rapidly than circRNAs in platelet preparations. Collectively, these results suggest that circulating platelets have lost on aveage over 90% of their progenitor mRNAs, and that translation in platelets occurrs against the backdrop of a highly degraded transcriptome. Finally, we find that transcripts classified as products of reverse transcriptase template switching are both enriched in platelets and resistant to decay, countering the recent suggestion that up to 50% of rearranged RNAs are artefacts. A single rRNA depleted total RNA sample was sequenced. This together with 25 publicly available rRNA depleted total RNA samples (including 3 from platelets) were analysed using PTESFinder v 1 (http://sourceforge.net/projects/ptesfinder-v1/) to identify back-splice junctions, characteristic of circRNA transcripts. The contribution of circRNA producing exons was analysed on a gene by gene basis as follows: All circRNA transcripts identified in any sample were first pooled to define exons which can contribute to circRNA generation using custom scripts (available on request). For each sample, expression estimates (RPKMI) across all circRNA producing exons were computed for each locus using the total size of exons (in bp) and the read counts mapping to them. Similarly, total size and exonic read counts for exons for which no circRNA were detected in any sample were used to compute expression estimates (RPKME) for non-circRNA producing exons for each locus. Abundance ratios (RPKMI/RPKME and RPKMI/RPKMI+RPKME) were calculated and compared between Platelets and human tissues using Wilcoxon signed-rank test. Please note that the '25sample_info_accn_no.txt' contains the accession numbers and tissue/cell type information for 25 samples analyzed together.
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Project description:Global analysis of gene expression in NIH3T3 cells over-expressing RNA-dependent serine/threonine protein kinase (PKR). Keywords: other
2004-09-22 | GSE682 | GEO