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1.
Cell ; 185(12): 2016-2034, 2022 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-35584701

RESUMO

Most circular RNAs are produced from the back-splicing of exons of precursor mRNAs. Recent technological advances have in part overcome problems with their circular conformation and sequence overlap with linear cognate mRNAs, allowing a better understanding of their cellular roles. Depending on their localization and specific interactions with DNA, RNA, and proteins, circular RNAs can modulate transcription and splicing, regulate stability and translation of cytoplasmic mRNAs, interfere with signaling pathways, and serve as templates for translation in different biological and pathophysiological contexts. Emerging applications of RNA circles to interfere with cellular processes, modulate immune responses, and direct translation into proteins shed new light on biomedical research. In this review, we discuss approaches used in circular RNA studies and the current understanding of their regulatory roles and potential applications.


Assuntos
RNA Circular , RNA , Proteínas/metabolismo , RNA/metabolismo , Precursores de RNA/metabolismo , Splicing de RNA , RNA Mensageiro/metabolismo
2.
Cell ; 177(4): 865-880.e21, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-31031002

RESUMO

Circular RNAs (circRNAs) produced from back-splicing of exons of pre-mRNAs are widely expressed, but current understanding of their functions is limited. These RNAs are stable in general and are thought to have unique structural conformations distinct from their linear RNA cognates. Here, we show that endogenous circRNAs tend to form 16-26 bp imperfect RNA duplexes and act as inhibitors of double-stranded RNA (dsRNA)-activated protein kinase (PKR) related to innate immunity. Upon poly(I:C) stimulation or viral infection, circRNAs are globally degraded by RNase L, a process required for PKR activation in early cellular innate immune responses. Augmented PKR phosphorylation and circRNA reduction are found in peripheral blood mononuclear cells (PBMCs) derived from patients with autoimmune disease systemic lupus erythematosus (SLE). Importantly, overexpression of the dsRNA-containing circRNA in PBMCs or T cells derived from SLE can alleviate the aberrant PKR activation cascade, thus providing a connection between circRNAs and SLE.


Assuntos
RNA Circular/metabolismo , RNA Circular/fisiologia , eIF-2 Quinase/metabolismo , Adolescente , Adulto , Doenças Autoimunes/genética , Linhagem Celular , Endorribonucleases/metabolismo , Feminino , Humanos , Imunidade Inata/genética , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/metabolismo , Lúpus Eritematoso Sistêmico/genética , Pessoa de Meia-Idade , Fosforilação , RNA/metabolismo , Splicing de RNA/genética , Estabilidade de RNA/fisiologia , RNA Circular/genética , RNA de Cadeia Dupla/metabolismo , Viroses/metabolismo , eIF-2 Quinase/imunologia
3.
Cell ; 185(13): 2390, 2022 Jun 23.
Artigo em Inglês | MEDLINE | ID: mdl-35750036
4.
Mol Cell ; 84(19): 3596-3609, 2024 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-39366349

RESUMO

Circular RNA is a group of covalently closed, single-stranded transcripts with unique biogenesis, stability, and conformation that play distinct roles in modulating cellular functions and also possess a great potential for developing circular RNA-based therapies. Importantly, due to its circular conformation, circular RNA generates distinct intramolecular base pairing that is different from the linear transcript. In this perspective, we review how circular RNA conformation can affect its turnover and modes of action, as well as what factors can modulate circular RNA conformation. We also discuss how understanding circular RNA conformation can facilitate learning about their functions as well as the remaining technological issues to further address their conformation. These efforts will ultimately inform the design of circular RNA-based platforms for biomedical applications.


Assuntos
Conformação de Ácido Nucleico , RNA Circular , RNA Circular/genética , RNA Circular/metabolismo , RNA Circular/química , Humanos , Animais , RNA/metabolismo , RNA/genética , RNA/química , Estabilidade de RNA , Pareamento de Bases , Relação Estrutura-Atividade
5.
Mol Cell ; 82(2): 420-434.e6, 2022 01 20.
Artigo em Inglês | MEDLINE | ID: mdl-34951963

RESUMO

Exon back-splicing-generated circular RNAs, as a group, can suppress double-stranded RNA (dsRNA)-activated protein kinase R (PKR) in cells. We have sought to synthesize immunogenicity-free, short dsRNA-containing RNA circles as PKR inhibitors. Here, we report that RNA circles synthesized by permuted self-splicing thymidylate synthase (td) introns from T4 bacteriophage or by Anabaena pre-tRNA group I intron could induce an immune response. Autocatalytic splicing introduces ∼74 nt td or ∼186 nt Anabaena extraneous fragments that can distort the folding status of original circular RNAs or form structures themselves to provoke innate immune responses. In contrast, synthesized RNA circles produced by T4 RNA ligase without extraneous fragments exhibit minimized immunogenicity. Importantly, directly ligated circular RNAs that form short dsRNA regions efficiently suppress PKR activation 103- to 106-fold higher than reported chemical compounds C16 and 2-AP, highlighting the future use of circular RNAs as potent inhibitors for diseases related to PKR overreaction.


Assuntos
Inibidores de Proteínas Quinases/farmacologia , RNA Circular/farmacologia , eIF-2 Quinase/antagonistas & inibidores , Células A549 , Bacteriófago T4/enzimologia , Bacteriófago T4/genética , Células HEK293 , Células HeLa , Humanos , Imunidade Inata/efeitos dos fármacos , Íntrons , Conformação de Ácido Nucleico , Inibidores de Proteínas Quinases/imunologia , RNA Ligase (ATP)/genética , RNA Ligase (ATP)/metabolismo , Precursores de RNA/genética , Precursores de RNA/metabolismo , RNA Circular/genética , RNA Circular/imunologia , Timidilato Sintase/genética , Timidilato Sintase/metabolismo , Proteínas Virais/genética , Proteínas Virais/metabolismo , eIF-2 Quinase/metabolismo
6.
Mol Cell ; 81(20): 4111-4113, 2021 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-34686312

RESUMO

Chen et al. (2021) have identified many internal ribosome entry site-like elements that can potentially drive circRNA translation. Dozens of such element-containing circRNAs-encoded peptides are validated, among which a circFGFR1-encoded protein acts as an antagonist of FGFR1.


Assuntos
Sítios Internos de Entrada Ribossomal , RNA Circular , Regulação da Expressão Gênica
7.
Mol Cell ; 69(1): 126-135.e6, 2018 01 04.
Artigo em Inglês | MEDLINE | ID: mdl-29304330

RESUMO

N6-methyladenosine (m6A) and adenosine-to-inosine (A-to-I) editing are two of the most abundant RNA modifications, both at adenosines. Yet, the interaction of these two types of adenosine modifications is largely unknown. Here we show a global A-to-I difference between m6A-positive and m6A-negative RNA populations. Both the presence and extent of A-to-I sites in m6A-negative RNA transcripts suggest a negative correlation between m6A and A-to-I. Suppression of m6A-catalyzing enzymes results in global A-to-I RNA editing changes. Further depletion of m6A modification increases the association of m6A-depleted transcripts with adenosine deaminase acting on RNA (ADAR) enzymes, resulting in upregulated A-to-I editing on the same m6A-depleted transcripts. Collectively, the effect of m6A on A-to-I suggests a previously underappreciated interplay between two distinct and abundant RNA modifications, highlighting a complex epitranscriptomic landscape.


Assuntos
Adenosina/análogos & derivados , Adenosina/química , Inosina/química , Edição de RNA/genética , RNA/genética , Adenosina Desaminase/genética , Adenosina Desaminase/metabolismo , Linhagem Celular Tumoral , Regulação da Expressão Gênica/genética , Células HEK293 , Células HeLa , Humanos , Metiltransferases/genética , Metiltransferases/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo
8.
Mol Cell ; 67(2): 214-227.e7, 2017 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-28625552

RESUMO

Circular RNAs (circRNAs) generated via back-splicing are enhanced by flanking complementary sequences. Expression levels of circRNAs vary under different conditions, suggesting participation of protein factors in their biogenesis. Using genome-wide siRNA screening that targets all human unique genes and an efficient circRNA expression reporter, we identify double-stranded RNA-binding domain containing immune factors NF90/NF110 as key regulators in circRNA biogenesis. NF90/NF110 promote circRNA production in the nucleus by associating with intronic RNA pairs juxtaposing the circRNA-forming exon(s); they also interact with mature circRNAs in the cytoplasm. Upon viral infection, circRNA expression is decreased, in part owing to the nuclear export of NF90/NF110 to the cytoplasm. Meanwhile, NF90/NF110 released from circRNP complexes bind to viral mRNAs as part of their functions in antiviral immune response. Our results therefore implicate a coordinated regulation of circRNA biogenesis and function by NF90/NF110 in viral infection.


Assuntos
Núcleo Celular/metabolismo , Proteínas do Fator Nuclear 90/metabolismo , Ligação Proteica , RNA Mensageiro/metabolismo , RNA Viral/metabolismo , RNA/biossíntese , Viroses/metabolismo , Transporte Ativo do Núcleo Celular , Núcleo Celular/efeitos dos fármacos , Perfilação da Expressão Gênica , Células HEK293 , Células HeLa , Interações Hospedeiro-Patógeno , Humanos , Proteínas do Fator Nuclear 90/genética , Proteínas do Fator Nuclear 90/imunologia , Poli I-C/farmacologia , RNA/química , RNA/genética , Interferência de RNA , Processamento Pós-Transcricional do RNA , Splicing de RNA , Estabilidade de RNA , RNA Circular , RNA Mensageiro/genética , RNA Viral/genética , Transfecção , Viroses/genética , Viroses/imunologia
9.
Nat Methods ; 18(1): 51-59, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-33288960

RESUMO

Circular RNAs (circRNAs) produced from back-spliced exons are widely expressed, but individual circRNA functions remain poorly understood owing to the lack of adequate methods for distinguishing circRNAs from cognate messenger RNAs with overlapping exons. Here, we report that CRISPR-RfxCas13d can effectively discriminate circRNAs from mRNAs by using guide RNAs targeting sequences spanning back-splicing junction (BSJ) sites featured in RNA circles. Using a lentiviral library that targets sequences across BSJ sites of highly expressed human circRNAs, we show that a group of circRNAs are important for cell growth mostly in a cell-type-specific manner and that a common oncogenic circRNA, circFAM120A, promotes cell proliferation by preventing the mRNA for family with sequence similarity 120A (FAM120A) from binding the translation inhibitor IGF2BP2. Further application of RfxCas13d-BSJ-gRNA screening has uncovered circMan1a2, which has regulatory potential in mouse embryo preimplantation development. Together, these results establish CRISPR-RfxCas13d as a useful tool for the discovery and functional study of circRNAs at both individual and large-scale levels.


Assuntos
Sistemas CRISPR-Cas , Neoplasias do Colo/patologia , Regulação Neoplásica da Expressão Gênica , RNA Circular/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Processamento Alternativo , Animais , Apoptose , Proliferação de Células , Neoplasias do Colo/genética , Neoplasias do Colo/metabolismo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
10.
Nat Chem Biol ; 17(7): 755-766, 2021 07.
Artigo em Inglês | MEDLINE | ID: mdl-34172967

RESUMO

RNA molecules fold into complex structures that enable their diverse functions in cells. Recent revolutionary innovations in transcriptome-wide RNA structural probing of living cells have ushered in a new era in understanding RNA functions. Here, we summarize the latest technological advances for probing RNA secondary structures and discuss striking discoveries that have linked RNA regulation and biological processes through interrogation of RNA structures. In particular, we highlight how different long noncoding RNAs form into distinct secondary structures that determine their modes of interactions with protein partners to realize their unique functions. These dynamic structures mediate RNA regulatory functions through altering interactions with proteins and other RNAs. We also outline current methodological hurdles and speculate about future directions for development of the next generation of RNA structure-probing technologies of higher sensitivity and resolution, which could then be applied in increasingly physiologically relevant studies.


Assuntos
RNA/química , Animais , Humanos , Conformação de Ácido Nucleico , RNA/metabolismo
11.
Methods ; 196: 47-55, 2021 12.
Artigo em Inglês | MEDLINE | ID: mdl-33571668

RESUMO

Circular RNAs are produced from back-splicing of exons of precursor mRNAs (pre-mRNAs). The sequences of exons in circular RNAs are identical to their linear cognate mRNAs, but the circular format may confer constraints on their folding and conformation, leading to potentially different functions from their linear RNA cognates. Here, we describe experimental and computational steps that optimize the selective 2'-hydroxyl acylation analyzed by primer extension and mutational profiling (SHAPE-MaP) to probe circular RNA secondary structure at single-nucleotide resolution in living cells.


Assuntos
RNA Circular , RNA , Conformação de Ácido Nucleico , RNA/química , Precursores de RNA , Splicing de RNA/genética , RNA Circular/genética
12.
IUBMB Life ; 68(11): 887-893, 2016 11.
Artigo em Inglês | MEDLINE | ID: mdl-27699981

RESUMO

RNA is essential for all kingdoms of life and exerts important functions beyond transferring genetic information from DNA to protein. With the advent of the state-of-the-art deep sequencing technology, a large portion of noncoding transcripts in eukaryotic genomes has been broadly identified. Among them, long noncoding RNAs (lncRNAs) have been emerged as a new class of RNA molecules that have regulatory potential in a variety of physiological and pathological processes. Here we summarize recent research progresses that have been made by scientists in China on lncRNAs, including their biogenesis, functional implication and the underlying mechanism of action at the current stage. © 2016 IUBMB Life, 68(11):887-893, 2016.


Assuntos
Pesquisa Biomédica/normas , RNA Longo não Codificante/fisiologia , Animais , Bibliometria , Carcinogênese/genética , Carcinogênese/metabolismo , Carcinogênese/patologia , China , Regulação Neoplásica da Expressão Gênica , Humanos , Melhoria de Qualidade , Interferência de RNA
13.
Nat Biotechnol ; 2024 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-38653797

RESUMO

Efforts to advance RNA aptamers as a new therapeutic modality have been limited by their susceptibility to degradation and immunogenicity. In a previous study, we demonstrated synthesized short double-stranded region-containing 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-α and 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.

14.
Genome Biol ; 23(1): 16, 2022 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-35012611

RESUMO

Many circular RNAs (circRNAs) are produced from back-splicing of exons of precursor mRNAs and are generally co-expressed with cognate linear RNAs. Methods for circRNA-specific knockout are lacking, largely due to sequence overlaps between forms. Here, we use base editors (BEs) for circRNA depletion. By targeting splice sites involved in both back-splicing and canonical splicing, BEs can repress circular and linear RNAs. Targeting sites predominantly for circRNA biogenesis, BEs could efficiently repress the production of circular but not linear RNAs. As hundreds of exons are predominantly back-spliced to produce circRNAs, this provides an efficient method to deplete circRNAs for functional study.


Assuntos
Splicing de RNA , RNA Circular , Éxons , RNA/genética , Precursores de RNA
15.
Genome Biol ; 22(1): 221, 2021 08 10.
Artigo em Inglês | MEDLINE | ID: mdl-34376223

RESUMO

Single-cell RNA-seq (scRNA-seq) profiles gene expression with high resolution. Here, we develop a stepwise computational method-called SCAPTURE to identify, evaluate, and quantify cleavage and polyadenylation sites (PASs) from 3' tag-based scRNA-seq. SCAPTURE detects PASs de novo in single cells with high sensitivity and accuracy, enabling detection of previously unannotated PASs. Quantified alternative PAS transcripts refine cell identity analysis beyond gene expression, enriching information extracted from scRNA-seq data. Using SCAPTURE, we show changes of PAS usage in PBMCs from infected versus healthy individuals at single-cell resolution.


Assuntos
Aprendizado Profundo , Poliadenilação , RNA-Seq , Análise de Célula Única , COVID-19/diagnóstico , Humanos , SARS-CoV-2 , Sensibilidade e Especificidade , Análise de Sequência de RNA , Transcriptoma
16.
Artigo em Inglês | MEDLINE | ID: mdl-32019863

RESUMO

RNA processing is critical for eukaryotic mRNA maturation and function. It appears there is no exception for other types of RNAs. Long noncoding RNAs (lncRNAs) represent a subclass of noncoding RNAs, have sizes of >200 nucleotides (nt), and participate in various aspects of gene regulation. Although many lncRNAs are capped, polyadenylated, and spliced just like mRNAs, others are derived from primary transcripts of RNA polymerase II and stabilized by forming circular structures or by ending with small nucleolar RNA-protein complexes. Here we summarize the recent progress in linking the processing and function of these unconventionally processed lncRNAs; we also discuss how directional RNA movement is achieved using the radial flux movement of nascent precursor ribosomal RNA (pre-rRNA) in the human nucleolus as an example.

17.
Genomics Proteomics Bioinformatics ; 17(5): 511-521, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31904419

RESUMO

Sequences of circular RNAs (circRNAs) produced from back-splicing of exon(s) completely overlap with those from cognate linear RNAs transcribed from the same gene loci with the exception of their back-splicing junction (BSJ) sites. Therefore, examination of global circRNA expression from RNA-seq datasets generally relies on the detection of RNA-seq fragments spanning BSJ sites, which is different from the quantification of linear RNA expression by normalized RNA-seq fragments mapped to whole gene bodies. Thus, direct comparison of circular and linear RNA expression from the same gene loci in a genome-wide manner has remained challenging. Here, we update the previously-reported CIRCexplorer pipeline to version 3 for circular and linear RNA expression analysis from ribosomal-RNA depleted RNA-seq (CIRCexplorer3-CLEAR). A new quantitation parameter, fragments per billion mapped bases (FPB), is applied to evaluate circular and linear RNA expression individually by fragments mapped to circRNA-specific BSJ sites or to linear RNA-specific splicing junction (SJ) sites. Comparison of circular and linear RNA expression levels is directly achieved by dividing FPBcirc by FPBlinear to generate a CIRCscore, which indicates the relative circRNA expression level using linear RNA expression level as the background. Highly-expressed circRNAs with low cognate linear RNA expression background can be readily identified by CIRCexplorer3-CLEAR for further investigation. CIRCexplorer3-CLEAR is publically available at https://github.com/YangLab/CLEAR.


Assuntos
RNA Circular/metabolismo , RNA/metabolismo , Interface Usuário-Computador , Linhagem Celular Tumoral , Humanos , RNA/química , Splicing de RNA , RNA Circular/química , Análise de Sequência de RNA , Transcriptoma
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