Circular RNAs in the Mammalian Brain Are Highly Abundant, Conserved, and Dynamically Expressed

doi:10.1016/J.MOLCEL.2015.03.027

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Circular RNAs in the Mammalian Brain Are Highly Abundant, Conserved, and Dynamically Expressed

Journal Article•DOI•

Circular RNAs in the Mammalian Brain Are Highly Abundant, Conserved, and Dynamically Expressed

Agnieszka Rybak-Wolf¹, Christin Stottmeister¹, Petar Glažar¹, Marvin Jens¹, Natalia S. Pino², Sebastian A. Giusti², Mor Hanan³, Mikaela Behm⁴, Osnat Bartok³, Reut Ashwal-Fluss³, Margareta Herzog¹, Luisa Schreyer¹, Panagiotis Papavasileiou¹, Andranik Ivanov¹, Marie Öhman⁴, Damian Refojo², Sebastian Kadener³, Nikolaus Rajewsky¹ - Show less +14 more•Institutions (4)

Max Delbrück Center for Molecular Medicine¹, Max Planck Society², Hebrew University of Jerusalem³, Stockholm University⁴

04 Jun 2015-Molecular Cell (Cell Press)-Vol. 58, Iss: 5, pp 870-885

TL;DR: A circRNA brain expression atlas and evidence for important circRNA functions and values as biomarkers are provided and discovered and analyzed thousands of neuronal human and mouse circRNAs.

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About: This article is published in Molecular Cell.The article was published on 2015-06-04 and is currently open access. It has received 1759 citations till now. The article focuses on the topics: Circular RNA & Non-coding RNA.

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Journal Article•DOI•

The biogenesis, biology and characterization of circular RNAs.

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Lasse Sommer Kristensen¹, Maria S. Andersen¹, Lotte V. W. Stagsted¹, Karoline K. Ebbesen¹, Thomas B. Hansen¹, Jørgen Kjems¹ - Show less +2 more•Institutions (1)

Aarhus University¹

08 Aug 2019-Nature Reviews Genetics

TL;DR: Advances in high-throughput RNA sequencing and circRNA-specific computational tools have driven the development of state-of-the-art approaches for their identification, and novel approaches to functional characterization are emerging.

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Abstract: Circular RNAs (circRNAs) are covalently closed, endogenous biomolecules in eukaryotes with tissue-specific and cell-specific expression patterns, whose biogenesis is regulated by specific cis-acting elements and trans-acting factors. Some circRNAs are abundant and evolutionarily conserved, and many circRNAs exert important biological functions by acting as microRNA or protein inhibitors ('sponges'), by regulating protein function or by being translated themselves. Furthermore, circRNAs have been implicated in diseases such as diabetes mellitus, neurological disorders, cardiovascular diseases and cancer. Although the circular nature of these transcripts makes their detection, quantification and functional characterization challenging, recent advances in high-throughput RNA sequencing and circRNA-specific computational tools have driven the development of state-of-the-art approaches for their identification, and novel approaches to functional characterization are emerging.

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2,372 citations

Journal Article•DOI•

Circular RNA profiling reveals an abundant circHIPK3 that regulates cell growth by sponging multiple miRNAs

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Qiupeng Zheng¹, Chunyang Bao¹, Weijie Guo², Weijie Guo¹, Shuyi Li¹, Shuyi Li², Jie Chen¹, Bing Chen¹, Yanting Luo¹, Dongbin Lyu¹, Yan Li¹, Guohai Shi¹, Linhui Liang¹, Jianren Gu², Xianghuo He¹, Shenglin Huang¹ - Show less +12 more•Institutions (2)

Fudan University¹, Shanghai Jiao Tong University²

06 Apr 2016-Nature Communications

TL;DR: The results provide evidence that circular RNA produced from precursor mRNA may have a regulatory role in human cells and characterize one abundant circRNA derived from Exon2 of the HIPK3 gene, termed circHIPK3.

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Abstract: Circular RNAs (circRNAs) represent a class of widespread and diverse endogenous RNAs that may regulate gene expression in eukaryotes. However, the regulation and function of human circRNAs remain largely unknown. Here we generate ribosomal-depleted RNA sequencing data from six normal tissues and seven cancers, and detect at least 27,000 circRNA candidates. Many of these circRNAs are differently expressed between the normal and cancerous tissues. We further characterize one abundant circRNA derived from Exon2 of the HIPK3 gene, termed circHIPK3. The silencing of circHIPK3 but not HIPK3 mRNA significantly inhibits human cell growth. Via a luciferase screening assay, circHIPK3 is observed to sponge to 9 miRNAs with 18 potential binding sites. Specifically, we show that circHIPK3 directly binds to miR-124 and inhibits miR-124 activity. Our results provide evidence that circular RNA produced from precursor mRNA may have a regulatory role in human cells.

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1,537 citations

Journal Article•DOI•

Circ-ZNF609 Is a Circular RNA that Can Be Translated and Functions in Myogenesis

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Ivano Legnini¹, Gaia Di Timoteo¹, Francesca Rossi¹, Mariangela Morlando¹, Francesca Briganti¹, Olga Sthandier¹, Alessandro Fatica¹, Tiziana Santini², Adrian Andronache², Mark Wade², Pietro Laneve², Nikolaus Rajewsky³, Irene Bozzoni², Irene Bozzoni¹ - Show less +10 more•Institutions (3)

Sapienza University of Rome¹, Istituto Italiano di Tecnologia², Max Delbrück Center for Molecular Medicine³

06 Apr 2017-Molecular Cell

TL;DR: One of them, circ-ZNF609, resulted in specifically controlling myoblast proliferation and is associated with heavy polysomes, providing an example of a protein-coding circRNA in eukaryotes.

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1,512 citations

Cites background or methods from "Circular RNAs in the Mammalian Brai..."

...Since circ-ZNF609 is highly expressed in neural tissues (Rybak-Wolf et al., 2015), we applied an established neuronal differentiation protocol (Wichterle and Peljto, 2008), and we checked for the expression of the FLAG-tagged circZNF609 RNA by northern blot in WT cells and in the F/D clone....
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...Moreover, theDNA that encodes circRNAs is more conserved than the DNA of flanking exons (Rybak-Wolf et al., 2015)....
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...It has been shown that circRNAexpression is oftenhighly conservedacross species and particularly abundant in mammalian neuronal tissues (Rybak-Wolf et al., 2015)....
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Journal Article•DOI•

Endogenous microRNA sponges: evidence and controversy.

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Daniel W. Thomson¹, Marcel E. Dinger¹, Marcel E. Dinger²•Institutions (2)

Garvan Institute of Medical Research¹, University of New South Wales²

01 May 2016-Nature Reviews Genetics

TL;DR: The evidence for and against the ceRNA hypothesis are critically evaluated to assess the impact of endogenous miRNA-sponge interactions and to propose an alternative function for messenger RNAs.

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Abstract: The competitive endogenous RNA (ceRNA) hypothesis proposes that transcripts with shared microRNA (miRNA) binding sites compete for post-transcriptional control. This hypothesis has gained substantial attention as a unifying function for long non-coding RNAs, pseudogene transcripts and circular RNAs, as well as an alternative function for messenger RNAs. Empirical evidence supporting the hypothesis is accumulating but not without attracting scepticism. Recent studies that model transcriptome-wide binding-site abundance suggest that physiological changes in expression of most individual transcripts will not compromise miRNA activity. In this Review, we critically evaluate the evidence for and against the ceRNA hypothesis to assess the impact of endogenous miRNA-sponge interactions.

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1,463 citations

Journal Article•DOI•

The Biogenesis, Functions, and Challenges of Circular RNAs.

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Xiang Li¹, Li Yang², Li Yang³, Ling-Ling Chen¹, Ling-Ling Chen³ - Show less +1 more•Institutions (3)

Chinese Academy of Sciences¹, CAS-MPG Partner Institute for Computational Biology², ShanghaiTech University³

02 Aug 2018-Molecular Cell

TL;DR: The recent progress on circRNA biogenesis and function is surveyed and technical obstacles in circRNA studies are discussed.

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1,270 citations

Cites background from "Circular RNAs in the Mammalian Brai..."

...…circRNAs Can Act as miRNA Sponges It has been proposed that the competing endogenous RNAs (ceRNAs) can act as sponges for miRNAs through their binding sites and that changes in ceRNA abundance from individual genes can modulate the activity of miRNAs on other target genes (Salmena et al., 2011)....
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References

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Journal Article•DOI•

An integrated encyclopedia of DNA elements in the human genome

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Principal investigators¹, Nhgri groups², Data production leads³, Lead analysts³•Institutions (3)

Wellcome Trust¹, University of Washington², Pennsylvania State University³

06 Sep 2012-Nature

TL;DR: The Encyclopedia of DNA Elements project provides new insights into the organization and regulation of the authors' genes and genome, and is an expansive resource of functional annotations for biomedical research.

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Abstract: The human genome encodes the blueprint of life, but the function of the vast majority of its nearly three billion bases is unknown. The Encyclopedia of DNA Elements (ENCODE) project has systematically mapped regions of transcription, transcription factor association, chromatin structure and histone modification. These data enabled us to assign biochemical functions for 80% of the genome, in particular outside of the well-studied protein-coding regions. Many discovered candidate regulatory elements are physically associated with one another and with expressed genes, providing new insights into the mechanisms of gene regulation. The newly identified elements also show a statistical correspondence to sequence variants linked to human disease, and can thereby guide interpretation of this variation. Overall, the project provides new insights into the organization and regulation of our genes and genome, and is an expansive resource of functional annotations for biomedical research.

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13,548 citations

Journal Article•DOI•

Circular RNAs are a large class of animal RNAs with regulatory potency

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Sebastian Memczak¹, Marvin Jens¹, Antigoni Elefsinioti¹, Francesca Torti¹, Janna Krueger¹, Agnieszka Rybak¹, Luisa Maier¹, Sebastian D. Mackowiak¹, Lea H. Gregersen¹, Mathias Munschauer¹, Alexander Loewer¹, Ulrike Ziebold¹, Markus Landthaler¹, Christine Kocks¹, Ferdinand le Noble¹, Nikolaus Rajewsky¹ - Show less +12 more•Institutions (1)

Max Delbrück Center for Molecular Medicine¹

21 Mar 2013-Nature

TL;DR: It is found that a human circRNA, antisense to the cerebellar degeneration-related protein 1 transcript (CDR1as), is densely bound by microRNA (miRNA) effector complexes and harbours 63 conserved binding sites for the ancient miRNA miR-7.

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Abstract: Circular RNAs (circRNAs) in animals are an enigmatic class of RNA with unknown function. To explore circRNAs systematically, we sequenced and computationally analysed human, mouse and nematode RNA. We detected thousands of well-expressed, stable circRNAs, often showing tissue/developmental-stage-specific expression. Sequence analysis indicated important regulatory functions for circRNAs. We found that a human circRNA, antisense to the cerebellar degeneration-related protein 1 transcript (CDR1as), is densely bound by microRNA (miRNA) effector complexes and harbours 63 conserved binding sites for the ancient miRNA miR-7. Further analyses indicated that CDR1as functions to bind miR-7 in neuronal tissues. Human CDR1as expression in zebrafish impaired midbrain development, similar to knocking down miR-7, suggesting that CDR1as is a miRNA antagonist with a miRNA-binding capacity ten times higher than any other known transcript. Together, our data provide evidence that circRNAs form a large class of post-transcriptional regulators. Numerous circRNAs form by head-to-tail splicing of exons, suggesting previously unrecognized regulatory potential of coding sequences.

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5,922 citations

"Circular RNAs in the Mammalian Brai..." refers background or methods in this paper

...(legend continued on next page) 6 Molecular Cell 58, 1–16, June 4, 2015 ª2015 Elsevier Inc. described expression pattern (Hansen et al., 2013; Memczak et al., 2013). circRNAs Are Highly Enriched in Synapses Many mRNAs are translated in the synapse in an activity-dependent fashion, which plays a…...
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..., sequencing ribosomal-depleted RNA, subsequent computational analyses, which uncover head-to-tail junctions (Gla zar et al., 2014; Memczak et al., 2013), and experimental validation....
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...…have recently emerged as a large class of animal RNAs (Hansen et al., 2013; Jeck and Sharpless, 2014; Jeck et al., 2013; Lasda and Parker, 2014; Memczak et al., 2013; Salzman et al., 2012, 2013) with complex tissue- and stage-specific expression patterns. circRNAs are much more stable than…...
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...Head-to-tail junctions were detected as previously described (Gla zar et al., 2014; Memczak et al., 2013)....
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...…this is a first systematic analysis of circRNA expression profiles in mammalian brains, and an Figure 1. circRNAs Are Highly Abundant in Mammalian Brain (A) circRNAs were detected as previously described (Memczak et al., 2013), an expression in deep sequencing data (‘‘Quantification module’’)....
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Journal Article•DOI•

Natural RNA circles function as efficient microRNA sponges

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Thomas B. Hansen¹, Trine I. Jensen¹, Bettina Hjelm Clausen, Jesper B. Bramsen¹, Bente Finsen, Christian Kroun Damgaard¹, Jørgen Kjems¹ - Show less +3 more•Institutions (1)

Aarhus University¹

21 Mar 2013-Nature

TL;DR: This study serves as the first functional analysis of a naturally expressed circular RNA, ciRS-7, which contains more than 70 selectively conserved miRNA target sites, and is highly and widely associated with Argonaute proteins in a miR-7-dependent manner.

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Abstract: MicroRNAs (miRNAs) are important post-transcriptional regulators of gene expression that act by direct base pairing to target sites within untranslated regions of messenger RNAs. Recently, miRNA activity has been shown to be affected by the presence of miRNA sponge transcripts, the so-called competing endogenous RNA in humans and target mimicry in plants. We previously identified a highly expressed circular RNA (circRNA) in human and mouse brain. Here we show that this circRNA acts as a miR-7 sponge; we term this circular transcript ciRS-7 (circular RNA sponge for miR-7). ciRS-7 contains more than 70 selectively conserved miRNA target sites, and it is highly and widely associated with Argonaute (AGO) proteins in a miR-7-dependent manner. Although the circRNA is completely resistant to miRNA-mediated target destabilization, it strongly suppresses miR-7 activity, resulting in increased levels of miR-7 targets. In the mouse brain, we observe overlapping co-expression of ciRS-7 and miR-7, particularly in neocortical and hippocampal neurons, suggesting a high degree of endogenous interaction. We further show that the testis-specific circRNA, sex-determining region Y (Sry), serves as a miR-138 sponge, suggesting that miRNA sponge effects achieved by circRNA formation are a general phenomenon. This study serves as the first, to our knowledge, functional analysis of a naturally expressed circRNA.

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5,885 citations

"Circular RNAs in the Mammalian Brai..." refers background in this paper

...Circular RNAs (circRNAs) have recently emerged as a large class of animal RNAs (Hansen et al., 2013; Jeck and Sharpless, 2014; Jeck et al., 2013; Lasda and Parker, 2014; Memczak et al., 2013; Salzman et al., 2012, 2013) with complex tissue- and stage-specific expression patterns. circRNAs are much…...
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...(legend continued on next page) 6 Molecular Cell 58, 1–16, June 4, 2015 ª2015 Elsevier Inc. described expression pattern (Hansen et al., 2013; Memczak et al., 2013). circRNAs Are Highly Enriched in Synapses Many mRNAs are translated in the synapse in an activity-dependent fashion, which plays a…...
[...]
...Circular RNAs (circRNAs) have recently emerged as a large class of animal RNAs (Hansen et al., 2013; Jeck and Sharpless, 2014; Jeck et al., 2013; Lasda and Parker, 2014; Memczak et al., 2013; Salzman et al., 2012, 2013) with complex tissue- and stage-specific expression patterns....
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...The human circRNA CDR1as has recently been identified as a negative regulator (‘‘sponge’’) of the microRNA miR-7 (Han- sen et al., 2013; Memczak et al., 2013), demonstrating the regulatory potential of circRNAs. circRNAs are thought to be predominantly produced by backsplicing reactions that…...
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...The human circRNA CDR1as has recently been identified as a negative regulator (‘‘sponge’’) of the microRNA miR-7 (Hansen et al., 2013; Memczak et al., 2013), demonstrating the regulatory potential of circRNAs....
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Journal Article•DOI•

Combinatorial microRNA target predictions.

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Azra Krek¹, Dominic Grün¹, Matthew N. Poy², Rachel Wolf¹, Lauren Rosenberg¹, Eric J Epstein², Philip MacMenamin¹, Isabelle da Piedade¹, Kristin C. Gunsalus¹, Markus Stoffel², Nikolaus Rajewsky¹ - Show less +7 more•Institutions (2)

New York University¹, Rockefeller University²

03 Apr 2005-Nature Genetics

TL;DR: PicTar, a computational method for identifying common targets of micro RNAs, is presented and widespread coordinate control executed by microRNAs is suggested, thus providing evidence for coordinate microRNA control in mammals.

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Abstract: MicroRNAs are small noncoding RNAs that recognize and bind to partially complementary sites in the 3' untranslated regions of target genes in animals and, by unknown mechanisms, regulate protein production of the target transcript. Different combinations of microRNAs are expressed in different cell types and may coordinately regulate cell-specific target genes. Here, we present PicTar, a computational method for identifying common targets of microRNAs. Statistical tests using genome-wide alignments of eight vertebrate genomes, PicTar's ability to specifically recover published microRNA targets, and experimental validation of seven predicted targets suggest that PicTar has an excellent success rate in predicting targets for single microRNAs and for combinations of microRNAs. We find that vertebrate microRNAs target, on average, roughly 200 transcripts each. Furthermore, our results suggest widespread coordinate control executed by microRNAs. In particular, we experimentally validate common regulation of Mtpn by miR-375, miR-124 and let-7b and thus provide evidence for coordinate microRNA control in mammals.

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4,660 citations

"Circular RNAs in the Mammalian Brai..." refers background or result in this paper

...2–9) of 58 miRNAs that are conserved among vertebrates (Krek et al., 2005) (Figure 6C)....
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...However, the ratio between them (signal-to-noise ratio, compare Krek et al., 2005) does not differ from flanking CDS (Figure 6E)....
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...Here, 30 UTRs serve as a positive control and show 3-fold enrichment for conserved miRNA binding, consistent with previous studies (Krek et al., 2005)....
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Journal Article•DOI•

Circular RNAs are abundant, conserved, and associated with ALU repeats

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William R. Jeck¹, Jessica A. Sorrentino¹, Kai Wang², Michael K. Slevin¹, Christin E. Burd¹, Jinze Liu², William F. Marzluff¹, Norman E. Sharpless - Show less +4 more•Institutions (2)

University of North Carolina at Chapel Hill¹, University of Kentucky²

01 Feb 2013-RNA

TL;DR: High-throughput sequencing of libraries prepared from ribosome-depleted RNA with or without digestion with the RNA exonuclease showed that ecircRNAs are abundant, stable, conserved and nonrandom products of RNA splicing that could be involved in control of gene expression.

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Abstract: Circular RNAs composed of exonic sequence have been described in a small number of genes. Thought to result from splicing errors, circular RNA species possess no known function. To delineate the universe of endogenous circular RNAs, we performed high-throughput sequencing (RNA-seq) of libraries prepared from ribosome-depleted RNA with or without digestion with the RNA exonuclease, RNase R. We identified >25,000 distinct RNA species in human fibroblasts that contained non-colinear exons (a "backsplice") and were reproducibly enriched by exonuclease degradation of linear RNA. These RNAs were validated as circular RNA (ecircRNA), rather than linear RNA, and were more stable than associated linear mRNAs in vivo. In some cases, the abundance of circular molecules exceeded that of associated linear mRNA by >10-fold. By conservative estimate, we identified ecircRNAs from 14.4% of actively transcribed genes in human fibroblasts. Application of this method to murine testis RNA identified 69 ecircRNAs in precisely orthologous locations to human circular RNAs. Of note, paralogous kinases HIPK2 and HIPK3 produce abundant ecircRNA from their second exon in both humans and mice. Though HIPK3 circular RNAs contain an AUG translation start, it and other ecircRNAs were not bound to ribosomes. Circular RNAs could be degraded by siRNAs and, therefore, may act as competing endogenous RNAs. Bioinformatic analysis revealed shared features of circularized exons, including long bordering introns that contained complementary ALU repeats. These data show that ecircRNAs are abundant, stable, conserved and nonrandom products of RNA splicing that could be involved in control of gene expression.

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3,310 citations

"Circular RNAs in the Mammalian Brai..." refers background in this paper

...One possible reason might be conserved expression of circRNAs, which have a high chance to be flanked by long introns (Jeck et al., 2013; Ivanov et al., 2015)....
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...Circular RNAs (circRNAs) have recently emerged as a large class of animal RNAs (Hansen et al., 2013; Jeck and Sharpless, 2014; Jeck et al., 2013; Lasda and Parker, 2014; Memczak et al., 2013; Salzman et al., 2012, 2013) with complex tissue- and stage-specific expression patterns. circRNAs are much…...
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