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1.
Postepy Biochem ; 65(3): 173-182, 2019 10 01.
Artigo em Polonês | MEDLINE | ID: mdl-31643164

RESUMO

Endoribonuclease III Dicer plays a crucial role in the biogenesis of small regulatory RNAs, such as microRNAs (miRNAs) and small inter­fering RNAs (siRNAs). However, this is not the only role that Dicer plays in cells. For example, it has been shown that Dicer is involved in processing of diverse classes of RNA, including tRNA and snoRNA, cleavage of repeat-element-derived RNAs, and maintenance of genome integrity. Dicer has also been found to participate in the chromosome fragmentation during apoptosis or in the inflammatory processes. More­over, a recent discovery of Dicer-binding passive sites in mRNAs and long non-coding RNAs, and its putative nucleic acid chaperone activity, has pointed out a novel regulatory role of the enzyme. Here we focus on human Dicer and review its structure and function including recent findings on miRNA-independent roles and their impact on cell biology.


Assuntos
Ribonuclease III/química , Ribonuclease III/metabolismo , Fragmentação do DNA , Humanos , Chaperonas Moleculares/química , Chaperonas Moleculares/metabolismo , Processamento Pós-Transcricional do RNA , RNA Longo não Codificante/química , RNA Longo não Codificante/metabolismo , RNA Mensageiro/química , RNA Mensageiro/metabolismo , Pequeno RNA não Traduzido/biossíntese , Pequeno RNA não Traduzido/metabolismo
2.
Nihon Yakurigaku Zasshi ; 154(3): 108-113, 2019.
Artigo em Japonês | MEDLINE | ID: mdl-31527359

RESUMO

Similar to calcium (Ca2+) and chloride (Cl-) ion channels/transporters, potassium (K+) channels have been recognized as a crucial cancer treatment target. Recent studies have provided convincing evidences of positive correlation between elevated expression levels of Ca2+-activated K+ (KCa) channels and cancer proliferation, metastasis, and poor patient prognosis. In cancer cells, KCa1.1 and KCa3.1 KCa channels are co-localized with Ca2+-permeable Orai/TRP channels to provide a positive-feedback loop for Ca2+ entry. They are responsible for the promotion of cell growth and metastasis in the different types of cancer, and are therefore potential therapeutic targets and biomarkers for cancer. We determined the epigenetic and post-transcriptional dysregulation of KCa3.1 by class I histone deacetylase inhibitors in breast and prostate cancer cells. We further determined the transcriptional repression and protein degradation of KCa1.1 by vitamin D receptor agonists and androgen receptor antagonists, which are expected as potential therapeutic drugs for triple-negative breast cancer. The anti-inflammatory cytokine, interleukin-10 (IL-10) is an immunosuppressive factor involved in tumorigenesis, and plays a crucial role in escape from tumor immune surveillance. We determined KCa3.1 activators are a possible therapeutic option to suppress the tumor-promoting activities of IL-10. These results may provide new insights into cancer treatment focused on Ca2+-activated K+ channels.


Assuntos
Neoplasias da Mama/patologia , Inibidores de Histona Desacetilases/farmacologia , Canais de Potássio Cálcio-Ativados/metabolismo , Neoplasias da Próstata/patologia , Antagonistas de Receptores de Andrógenos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células , Epigênese Genética , Feminino , Humanos , Vigilância Imunológica , Interleucina-10/metabolismo , Masculino , Proteólise , Processamento Pós-Transcricional do RNA , Receptores de Calcitriol/agonistas
3.
Medicine (Baltimore) ; 98(26): e15858, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31261494

RESUMO

This analysis aims to describe the outcomes of two nonambulatory patients with Duchenne muscular dystrophy (DMD) who participated in two clinical studies. The two consecutive trials of eteplirsen (studies 201 and 202) were conducted in patients with DMD (N = 12) and confirmed genetic mutations amenable to exon 51 skipping.In study 201, 12 patients were randomized to receive once-weekly, double-blind intravenous infusions of eteplirsen 30 or 50 mg/kg or placebo for 24 weeks; patients then received open-label eteplirsen during weeks 25 through 28. All 12 patients continued onto open-label extension study 202 and received long-term treatment with eteplirsen. We compared cardiac, pulmonary, and upper limb function and dystrophin production in the nonambulatory twin patients versus the 10 ambulatory patients through 240 combined treatment weeks.Ten study patients remained ambulatory through both studies, while the identical twin patients both experienced early, rapid loss of ambulation. The twin patients had greater disease severity at baseline (6-minute walk test [6MWT], 330 and 256 m) versus the other patients (n = 10; 6MWT range, 341-418 m). They maintained cardiac and upper limb function through combined week 240, with outcomes similar to those of the patients who remained ambulatory. Dystrophin production was confirmed following eteplirsen treatment.Despite the loss of ambulation, other markers of disease progression remained relatively stable in the eteplirsen-treated twin patients and were similar to those of the ambulatory patients.


Assuntos
Morfolinos/uso terapêutico , Distrofia Muscular de Duchenne/tratamento farmacológico , Criança , Progressão da Doença , Doenças em Gêmeos , Método Duplo-Cego , Distrofina/genética , Distrofina/metabolismo , Humanos , Masculino , Morfolinos/efeitos adversos , Distrofia Muscular de Duchenne/genética , Distrofia Muscular de Duchenne/fisiopatologia , Processamento Pós-Transcricional do RNA/efeitos dos fármacos , Índice de Gravidade de Doença , Resultado do Tratamento , Teste de Caminhada , Caminhada
4.
Sheng Wu Gong Cheng Xue Bao ; 35(5): 775-783, 2019 May 25.
Artigo em Chinês | MEDLINE | ID: mdl-31222996

RESUMO

Messenger RNA (mRNA) can be modified by more than 100 chemical modifications. Among these modifications, N6-methyladenosine (m6A) is one of the most prevalent modifications. During the processes of cells differentiation, embryo development or stress, m6A can be modified on key mRNAs and regulate the progress of cells through modulating mRNA metabolism and translation. Other mRNA modifications, including N1-methyladenosine (m¹A), 5-methylcytosine (m5C) and pseudouridine, together with m6A form the epitranscriptome of mRNA that accurately modulate the mRNA translation. Here we review the types and characteristic of mRNA epigenetic modifications, especially the recent progresses of the function of m6A, we also expect the main research direction of m6A epigenetic modification in the future.


Assuntos
Adenosina/análogos & derivados , Epigênese Genética , Regulação da Expressão Gênica , RNA Mensageiro , Adenosina/genética , Adenosina/metabolismo , Diferenciação Celular/genética , Desenvolvimento Embrionário/genética , Processamento Pós-Transcricional do RNA , RNA Mensageiro/metabolismo
5.
Nat Commun ; 10(1): 2544, 2019 06 11.
Artigo em Inglês | MEDLINE | ID: mdl-31186424

RESUMO

Cas13d, the type VI-D CRISPR-Cas effector, is an RNA-guided ribonuclease that has been repurposed to edit RNA in a programmable manner. Here we report the detailed structural and functional analysis of the uncultured Ruminococcus sp. Cas13d (UrCas13d)-crRNA complex. Two hydrated Mg2+ ions aid in stabilizing the conformation of the crRNA repeat region. Sequestration of divalent metal ions does not alter pre-crRNA processing, but abolishes target cleavage by UrCas13d. Notably, the pre-crRNA processing is executed by the HEPN-2 domain. Furthermore, both the structure and sequence of the nucleotides U(-8)-C(-1) within the repeat region are indispensable for target cleavage, and are specifically recognized by UrCas13d. Moreover, correct base pairings within two separate spacer regions (an internal and a 3'-end region) are essential for target cleavage. These findings provide a framework for the development of Cas13d into a tool for a wide range of applications.


Assuntos
Proteínas de Bactérias/metabolismo , Proteínas Associadas a CRISPR/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas , Ribonucleases/metabolismo , Ruminococcus/genética , Sequência de Aminoácidos , Proteínas de Bactérias/química , Proteínas de Bactérias/genética , Proteínas Associadas a CRISPR/química , Proteínas Associadas a CRISPR/metabolismo , Sistemas CRISPR-Cas , Conformação de Ácido Nucleico , Domínios Proteicos , Processamento Pós-Transcricional do RNA , RNA Bacteriano/genética , RNA Bacteriano/metabolismo , RNA Guia/genética , Ribonucleases/química , Ribonucleases/genética , Ruminococcus/enzimologia
6.
Cell Mol Life Sci ; 76(19): 3745-3752, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31165201

RESUMO

RNA-binding proteins (RBPs) and microRNAs (miRNAs) are the most important regulators of mRNA stability and translation in eukaryotic cells; however, the complex interplay between these systems is only now coming to light. RBPs and miRNAs regulate a unique set of targets in either a positive or negative manner and their regulation is mainly opposed to each other on overlapping targets. In some cases, the levels of RBPs or miRNAs regulate the cellular levels of one another and decreased levels of either results in changes in translation of their targets. There is growing evidence that these regulatory circuits are crucial in the development and progression of cancer; however, the rules underlying synergism and antagonism between miRNAs and RNA-binding proteins remain unclear. Synthetic biology seeks to develop artificial systems to better understand their natural counterparts and to develop new, useful technologies for manipulation of gene expression at the RNA level. The recent development of artificial RNA-binding proteins promises to enable a much greater understanding of the importance of the functional interactions between RNA-binding proteins and miRNAs, as well as enabling their manipulation for therapeutic purposes.


Assuntos
MicroRNAs/metabolismo , Neoplasias/genética , Proteínas de Ligação a RNA/metabolismo , Regulação Neoplásica da Expressão Gênica , Doenças Genéticas Inatas/terapia , Humanos , Neoplasias/metabolismo , Neoplasias/terapia , Engenharia de Proteínas , Processamento Pós-Transcricional do RNA , Proteínas de Ligação a RNA/genética
7.
Nat Genet ; 51(6): 973-980, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31133750

RESUMO

We address the challenge of detecting the contribution of noncoding mutations to disease with a deep-learning-based framework that predicts the specific regulatory effects and the deleterious impact of genetic variants. Applying this framework to 1,790 autism spectrum disorder (ASD) simplex families reveals a role in disease for noncoding mutations-ASD probands harbor both transcriptional- and post-transcriptional-regulation-disrupting de novo mutations of significantly higher functional impact than those in unaffected siblings. Further analysis suggests involvement of noncoding mutations in synaptic transmission and neuronal development and, taken together with previous studies, reveals a convergent genetic landscape of coding and noncoding mutations in ASD. We demonstrate that sequences carrying prioritized mutations identified in probands possess allele-specific regulatory activity, and we highlight a link between noncoding mutations and heterogeneity in the IQ of ASD probands. Our predictive genomics framework illuminates the role of noncoding mutations in ASD and prioritizes mutations with high impact for further study, and is broadly applicable to complex human diseases.


Assuntos
Transtorno do Espectro Autista/genética , Aprendizado Profundo , Predisposição Genética para Doença , Genoma Humano , Genômica , Mutação , RNA não Traduzido , Algoritmos , Alelos , Transtorno do Espectro Autista/diagnóstico , Biologia Computacional/métodos , Expressão Gênica , Regulação da Expressão Gênica , Genes Reporter , Estudos de Associação Genética , Genômica/métodos , Humanos , Fenótipo , Processamento Pós-Transcricional do RNA , Transcrição Genética
8.
Gastroenterology ; 157(3): 731-743, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31103627

RESUMO

BACKGROUND & AIMS: Paneth cells secrete antimicrobial proteins including lysozyme via secretory autophagy as part of the mucosal protective response. The ELAV like RNA-binding protein 1 (ELAVL1, also called HuR) regulates stability and translation of messenger RNAs (mRNAs) and many aspects of mucosal physiology. We studied the posttranscriptional mechanisms by which HuR regulates Paneth cell function. METHODS: Intestinal mucosal tissues were collected from mice with intestinal epithelium (IE)-specific disruption of HuR (IE-HuR-/-), HuRfl/fl-Cre- mice (controls), and patients with inflammatory bowel diseases and analyzed by histology and immunohistochemistry. Paneth cell functions were determined by lysozyme-immunostaining assays. We isolated primary enterocytes from IE-HuR-/- and control mice and derived intestinal organoids. HuR and the chaperone CNPY3 were overexpressed from transgenes in intestinal epithelial cells (IECs) or knocked down with small interfering RNAs. We performed RNA pulldown assays to investigate interactions between HuR and its target mRNAs. RESULTS: Intestinal tissues from IE-HuR-/- mice had reduced numbers of Paneth cells, and Paneth cells had fewer lysozyme granules per cell, compared with tissues from control mice, but there were no effects on Goblet cells or enterocytes. Intestinal mucosa from patients with inflammatory bowel diseases had reduced levels of HuR and fewer Paneth cells. IE-HuR-/- mice did not have the apical distribution of TLR2 in the intestinal mucosa as observed in control mice. IECs from IE-HuR-/- mice expressed lower levels of CNPY3. Intestinal organoids from IE-HuR-/- mice were smaller and contained fewer buds compared with those generated from controls, and had fewer lysozyme-positive cells. In IECs, knockdown of HuR decreased levels of the autophagy proteins LC3-I and LC3-II, compared with control cells, and prevented rapamycin-induced autophagy. We found HuR to interact directly with the Cnpy3 mRNA coding region and increase levels of CNPY3 by increasing the stability and translation of Cnpy3 mRNA. CNPY3 bound TLR2, and cells with knockdown of CNPY3 or HuR lost membrane localization of TLR2, but increased cytoplasmic levels of TLR2. CONCLUSIONS: In studies of mice, IECs, and human tissues, we found HuR to increase expression of CNPY3 at the posttranscriptional level. CNPY3 is required for membrane localization of TLR2 and Paneth cell function.


Assuntos
Membrana Celular/metabolismo , Proteína Semelhante a ELAV 1/metabolismo , Intestino Delgado/metabolismo , Chaperonas Moleculares/metabolismo , Celulas de Paneth/metabolismo , Processamento Pós-Transcricional do RNA , Receptor 2 Toll-Like/metabolismo , Animais , Estudos de Casos e Controles , Células Cultivadas , Proteína Semelhante a ELAV 1/deficiência , Proteína Semelhante a ELAV 1/genética , Humanos , Doenças Inflamatórias Intestinais/genética , Doenças Inflamatórias Intestinais/metabolismo , Doenças Inflamatórias Intestinais/patologia , Intestino Delgado/patologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Chaperonas Moleculares/genética , Celulas de Paneth/patologia , Transporte Proteico , Transdução de Sinais , Regulação para Cima
9.
Mol Cell ; 74(4): 640-650, 2019 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-31100245

RESUMO

Cellular RNAs are naturally decorated with a variety of chemical modifications. The structural diversity of the modified nucleosides provides regulatory potential to sort groups of RNAs for organized metabolism and functions, thus affecting gene expression. Recent years have witnessed a burst of interest in and understanding of RNA modification biology, thanks to the emerging transcriptome-wide sequencing methods for mapping modified sites, highly sensitive mass spectrometry for precise modification detection and quantification, and extensive characterization of the modification "effectors," including enzymes ("writers" and "erasers") that alter the modification level and binding proteins ("readers") that recognize the chemical marks. However, challenges remain due to the vast heterogeneity in expression abundance of different RNA species, further complicated by divergent cell-type-specific and tissue-specific expression and localization of the effectors as well as modifications. In this review, we highlight recent progress in understanding the function of N6-methyladenosine (m6A), the most abundant internal mark on eukaryotic mRNA, in light of the specific biological contexts of m6A effectors. We emphasize the importance of context for RNA modification regulation and function.


Assuntos
Adenosina/análogos & derivados , Metilação , RNA Mensageiro/genética , RNA/genética , Adenosina/genética , Células Eucarióticas/metabolismo , Regulação da Expressão Gênica/genética , Especificidade de Órgãos/genética , Processamento Pós-Transcricional do RNA/genética , Transcriptoma
10.
BMC Bioinformatics ; 20(1): 223, 2019 May 02.
Artigo em Inglês | MEDLINE | ID: mdl-31046660

RESUMO

BACKGROUND: Over one hundred different types of post-transcriptional RNA modifications have been identified in human. Researchers discovered that RNA modifications can regulate various biological processes, and RNA methylation, especially N6-methyladenosine, has become one of the most researched topics in epigenetics. RESULTS: To date, the study of epitranscriptome layer gene regulation is mostly focused on the function of mediator proteins of RNA methylation, i.e., the readers, writers and erasers. There is limited investigation of the functional relevance of individual m6A RNA methylation site. To address this, we annotated human m6A sites in large-scale based on the guilt-by-association principle from an RNA co-methylation network. It is constructed based on public human MeRIP-Seq datasets profiling the m6A epitranscriptome under 32 independent experimental conditions. By systematically examining the network characteristics obtained from the RNA methylation profiles, a total of 339,158 putative gene ontology functions associated with 1446 human m6A sites were identified. These are biological functions that may be regulated at epitranscriptome layer via reversible m6A RNA methylation. The results were further validated on a soft benchmark by comparing to a random predictor. CONCLUSIONS: An online web server m6Acomet was constructed to support direct query for the predicted biological functions of m6A sites as well as the sites exhibiting co-methylated patterns at the epitranscriptome layer. The m6Acomet web server is freely available at: www.xjtlu.edu.cn/biologicalsciences/m6acomet .


Assuntos
Adenosina/análogos & derivados , Processamento Pós-Transcricional do RNA , Adenosina/metabolismo , Epigênese Genética , Humanos , Metilação , Software
11.
Nat Commun ; 10(1): 2065, 2019 05 06.
Artigo em Inglês | MEDLINE | ID: mdl-31061416

RESUMO

N6-Methyladenosine (m6A) modification has been implicated in the progression of several cancers. We reveal that during epithelial-mesenchymal transition (EMT), one important step for cancer cell metastasis, m6A modification of mRNAs increases in cancer cells. Deletion of methyltransferase-like 3 (METTL3) down-regulates m6A, impairs the migration, invasion and EMT of cancer cells both in vitro and in vivo. m6A-sequencing and functional studies confirm that Snail, a key transcription factor of EMT, is involved in m6A-regulated EMT. m6A in Snail CDS, but not 3'UTR, triggers polysome-mediated translation of Snail mRNA in cancer cells. Loss and gain functional studies confirm that YTHDF1 mediates m6A-increased translation of Snail mRNA. Moreover, the upregulation of METTL3 and YTHDF1 act as adverse prognosis factors for overall survival (OS) rate of liver cancer patients. Our study highlights the critical roles of m6A on regulation of EMT in cancer cells and translation of Snail during this process.


Assuntos
Adenosina/análogos & derivados , Transição Epitelial-Mesenquimal/genética , Neoplasias Hepáticas/genética , RNA/metabolismo , Fatores de Transcrição da Família Snail/genética , Adenosina/metabolismo , Animais , Linhagem Celular Tumoral , Regulação para Baixo , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Estimativa de Kaplan-Meier , Fígado/patologia , Neoplasias Hepáticas/mortalidade , Neoplasias Hepáticas/patologia , Metilação , Metiltransferases/genética , Metiltransferases/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Processamento Pós-Transcricional do RNA , Proteínas de Ligação a RNA/metabolismo , Análise Serial de Tecidos , Regulação para Cima , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Nat Cell Biol ; 21(5): 552-559, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31048770

RESUMO

The deposition of chemical modifications into RNA is a crucial regulator of temporal and spatial gene expression programs during development. Accordingly, altered RNA modification patterns are widely linked to developmental diseases. Recently, the dysregulation of RNA modification pathways also emerged as a contributor to cancer. By modulating cell survival, differentiation, migration and drug resistance, RNA modifications add another regulatory layer of complexity to most aspects of tumourigenesis.


Assuntos
Diferenciação Celular/genética , Neoplasias/genética , Processamento Pós-Transcricional do RNA/genética , RNA/genética , Linhagem da Célula/genética , Movimento Celular/genética , Proliferação de Células/genética , Sobrevivência Celular/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias/patologia , RNA/metabolismo
13.
Int J Mol Sci ; 20(9)2019 May 06.
Artigo em Inglês | MEDLINE | ID: mdl-31064115

RESUMO

Although the large majority of mitochondrial proteins are nuclear encoded, for their correct functioning mitochondria require the expression of 13 proteins, two rRNA, and 22 tRNA codified by mitochondrial DNA (mtDNA). Once transcribed, mitochondrial RNA (mtRNA) is processed, mito-ribosomes are assembled, and mtDNA-encoded proteins belonging to the respiratory chain are synthesized. These processes require the coordinated spatio-temporal action of several enzymes, and many different factors are involved in the regulation and control of protein synthesis and in the stability and turnover of mitochondrial RNA. In this review, we describe the essential steps of mitochondrial RNA synthesis, maturation, and degradation, the factors controlling these processes, and how the alteration of these processes is associated with human pathologies.


Assuntos
Doenças Mitocondriais/genética , Processamento Pós-Transcricional do RNA , Estabilidade de RNA , RNA Mitocondrial/genética , Animais , Humanos , Doenças Mitocondriais/metabolismo , RNA Mitocondrial/metabolismo
14.
Mol Cells ; 42(4): 301-312, 2019 Apr 30.
Artigo em Inglês | MEDLINE | ID: mdl-31091556

RESUMO

Post-transcriptional regulation underlies the circadian control of gene expression and animal behaviors. However, the role of mRNA surveillance via the nonsense-mediated mRNA decay (NMD) pathway in circadian rhythms remains elusive. Here, we report that Drosophila NMD pathway acts in a subset of circadian pacemaker neurons to maintain robust 24 h rhythms of free-running locomotor activity. RNA interference-mediated depletion of key NMD factors in timeless-expressing clock cells decreased the amplitude of circadian locomotor behaviors. Transgenic manipulation of the NMD pathway in clock neurons expressing a neuropeptide PIGMENT-DISPERSING FACTOR (PDF) was sufficient to dampen or lengthen free-running locomotor rhythms. Confocal imaging of a transgenic NMD reporter revealed that arrhythmic Clock mutants exhibited stronger NMD activity in PDF-expressing neurons than wild-type. We further found that hypomorphic mutations in Suppressor with morphogenetic effect on genitalia 5 (Smg5 ) or Smg6 impaired circadian behaviors. These NMD mutants normally developed PDF-expressing clock neurons and displayed daily oscillations in the transcript levels of core clock genes. By contrast, the loss of Smg5 or Smg6 function affected the relative transcript levels of cAMP response element-binding protein B (CrebB ) in an isoform-specific manner. Moreover, the overexpression of a transcriptional repressor form of CrebB rescued free-running locomotor rhythms in Smg5-depleted flies. These data demonstrate that CrebB is a rate-limiting substrate of the genetic NMD pathway important for the behavioral output of circadian clocks in Drosophila.


Assuntos
Relógios Circadianos , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila/genética , Mutação , Degradação do RNAm Mediada por Códon sem Sentido , Transativadores/metabolismo , Animais , Animais Geneticamente Modificados , Proteínas CLOCK/genética , Drosophila/metabolismo , Proteínas de Drosophila/genética , Endorribonucleases/genética , Endorribonucleases/metabolismo , Neurônios/metabolismo , Neuropeptídeos/genética , Neuropeptídeos/metabolismo , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Processamento Pós-Transcricional do RNA , Transdução de Sinais
15.
Mol Cell ; 74(3): 494-507.e8, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-30930054

RESUMO

N6-methyladenosine (m6A) is the most abundant internal modification in RNAs and plays regulatory roles in a variety of biological and physiological processes. Despite its important roles, the molecular mechanism underlying m6A-mediated gene regulation is poorly understood. Here, we show that m6A-containing RNAs are subject to endoribonucleolytic cleavage via YTHDF2 (m6A reader protein), HRSP12 (adaptor protein), and RNase P/MRP (endoribonucleases). We demonstrate that HRSP12 functions as an adaptor to bridge YTHDF2 and RNase P/MRP, eliciting rapid degradation of YTHDF2-bound RNAs. Transcriptome-wide analyses show that m6A RNAs that are preferentially targeted for endoribonucleolytic cleavage have an HRSP12-binding site and a RNase P/MRP-directed cleavage site upstream and downstream of the YTHDF2-binding site, respectively. We also find that a subset of m6A-containing circular RNAs associates with YTHDF2 in an HRSP12-dependent manner and is selectively downregulated by RNase P/MRP. Thus, our data expand the known functions of RNase P/MRP to endoribonucleolytic cleavage of m6A RNAs.


Assuntos
Adenosina/análogos & derivados , Proteínas de Choque Térmico/genética , Estabilidade de RNA/genética , Proteínas de Ligação a RNA/genética , Ribonuclease P/genética , Ribonucleases/genética , Adenosina/genética , Dioxigenase FTO Dependente de alfa-Cetoglutarato/genética , Sítios de Ligação/genética , Escherichia coli/genética , Regulação da Expressão Gênica/genética , Células HeLa , Humanos , Metiltransferases/genética , RNA/genética , Processamento Pós-Transcricional do RNA/genética , Transcriptoma/genética
16.
Mol Cell ; 74(4): 688-700.e3, 2019 05 16.
Artigo em Inglês | MEDLINE | ID: mdl-30930056

RESUMO

Mutations in RNA-processing enzymes are increasingly linked to human disease. Telomerase RNA and related noncoding RNAs require 3' end-processing steps, including oligoadenylation. Germline mutations in poly(A)ribonuclease (PARN) cause accumulation of extended human telomerase RNA (hTR) species and precipitate dyskeratosis congenita and pulmonary fibrosis. Here, we develop nascent RNAend-seq to measure processing rates of RNA precursors. We find that mature hTR derives from extended precursors but that in PARN-mutant cells hTR maturation kinetically stalls and unprocessed precursors are degraded. Loss of poly(A)polymerase PAPD5 in PARN-mutant cells accelerates hTR maturation and restores hTR processing, indicating that oligoadenylation and deadenylation set rates of hTR maturation. The H/ACA domain mediates hTR maturation by precisely defining the 3' end, recruiting poly(A)polymerase activity, and conferring sensitivity to PARN regulation. These data reveal a feedforward circuit in which post-transcriptional oligoadenylation controls RNA maturation kinetics. Similar alterations in RNA processing rates may contribute to mechanisms of RNA-based human disease.


Assuntos
Disceratose Congênita/genética , Exorribonucleases/genética , RNA Nucleotidiltransferases/genética , RNA/genética , Telomerase/genética , Disceratose Congênita/patologia , Mutação em Linhagem Germinativa/genética , Células HeLa , Humanos , Cinética , Processamento Pós-Transcricional do RNA/genética
17.
Nat Commun ; 10(1): 1757, 2019 04 15.
Artigo em Inglês | MEDLINE | ID: mdl-30988284

RESUMO

Cyclin-dependent kinase 12 (CDK12) modulates transcription elongation by phosphorylating the carboxy-terminal domain of RNA polymerase II and selectively affects the expression of genes involved in the DNA damage response (DDR) and mRNA processing. Yet, the mechanisms underlying such selectivity remain unclear. Here we show that CDK12 inhibition in cancer cells lacking CDK12 mutations results in gene length-dependent elongation defects, inducing premature cleavage and polyadenylation (PCPA) and loss of expression of long (>45 kb) genes, a substantial proportion of which participate in the DDR. This early termination phenotype correlates with an increased number of intronic polyadenylation sites, a feature especially prominent among DDR genes. Phosphoproteomic analysis indicated that CDK12 directly phosphorylates pre-mRNA processing factors, including those regulating PCPA. These results support a model in which DDR genes are uniquely susceptible to CDK12 inhibition primarily due to their relatively longer lengths and lower ratios of U1 snRNP binding to intronic polyadenylation sites.


Assuntos
Quinases Ciclina-Dependentes/genética , Dano ao DNA , Reparo do DNA/genética , Linhagem Celular Tumoral , Cromatografia Líquida de Alta Pressão , Cromatografia Líquida , Humanos , Modelos Moleculares , Fosforilação , Poliadenilação , Processamento Pós-Transcricional do RNA , Espectrometria de Massas em Tandem
18.
Virol Sin ; 34(2): 135-161, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31025296

RESUMO

Kaposi's sarcoma-associated herpesvirus (KSHV), also known as human herpesvirus-8 (HHV-8), is etiologically linked to the development of Kaposi's sarcoma, primary effusion lymphoma, and multicentric Castleman's disease. These malignancies often occur in immunosuppressed individuals, making KSHV infection-associated diseases an increasing global health concern with persistence of the AIDS epidemic. KSHV exhibits biphasic life cycles between latent and lytic infection and extensive transcriptional and posttranscriptional regulation of gene expression. As a member of the herpesvirus family, KSHV has evolved many strategies to evade the host immune response, which help the virus establish a successful lifelong infection. In this review, we summarize the current research status on the biology of latent and lytic viral infection, the regulation of viral life cycles and the related pathogenesis.


Assuntos
Herpesvirus Humano 8/fisiologia , Herpesvirus Humano 8/patogenicidade , Transcrição Genética , Replicação Viral , Animais , Hiperplasia do Linfonodo Gigante/virologia , Estudos Clínicos como Assunto , Expressão Gênica , Regulação Viral da Expressão Gênica , Interações Hospedeiro-Patógeno , Humanos , Evasão da Resposta Imune , Hospedeiro Imunocomprometido , Linfoma de Efusão Primária/virologia , Camundongos , Processamento Pós-Transcricional do RNA , Sarcoma de Kaposi/virologia , Proteínas Virais/genética , Latência Viral
19.
BMC Mol Biol ; 20(1): 11, 2019 04 08.
Artigo em Inglês | MEDLINE | ID: mdl-30961536

RESUMO

BACKGROUND: Camels possess the characteristics of salt- and drought-resistances, due to the long-time adaption to the living environment in desert. The camel resistance research on transcriptome is rare and deficient, especially reabsorption in renal cortex. Non-coding RNAs are normally considered as the RNA molecules that are not translated into proteins, their current roles remain mostly in regulation of information flux from DNA to protein, further on normal life activities and diseases. In order to reveal the mysterious veil of the post-transcriptional regulation of ncRNAs in renal cortex for the first time as far as we know, we designed and carried out the experiment of salt stress and water-deprivation stress in camel. RESULTS: By means of RNA-seq in renal cortex of Alxa Bactrian Camel (Camelus bactrianus), we identified certain significantly differential RNAs, including 4 novel lncRNAs, 11 miRNAs and 13 mRNAs under salt stress, 0 lncRNAs, 18 miRNAs and 14 mRNAs under water-deprivation stress. By data analysis, the response pathway of post-transcriptional regulation concerning salt and water-deprivation stresses was put forward, involving preventing sodium from entering the cell, purifying of water and compensating neutral amino acids by miR-193b, miR-542-5p interaction with SLC6A19 mRNA. CONCLUSION: Based on the resistance-related lncRNAs, miRNAs, and mRNAs, we proposed the post-transcriptional regulation pathway to explain how camels respond to salt and water-deprivation stresses in the ncRNAs regulation level of renal cortex for the first time, thus hoping to provide a theoretical basis for therapy of disease that is similar to high blood pressure in humans.


Assuntos
Camelus/genética , Camelus/fisiologia , Córtex Renal/fisiologia , RNA não Traduzido/genética , Estresse Salino/genética , Transcriptoma , Privação de Água , Sistemas de Transporte de Aminoácidos Neutros/genética , Animais , Regulação da Expressão Gênica , MicroRNAs/genética , Processamento Pós-Transcricional do RNA , RNA Longo não Codificante/genética , RNA Mensageiro/genética
20.
Mol Cell ; 74(3): 521-533.e6, 2019 05 02.
Artigo em Inglês | MEDLINE | ID: mdl-30952514

RESUMO

Cellular RNAs often colocalize with cytoplasmic, membrane-less ribonucleoprotein (RNP) granules enriched for RNA-processing enzymes, termed processing bodies (PBs). Here we track the dynamic localization of individual miRNAs, mRNAs, and long non-coding RNAs (lncRNAs) to PBs using intracellular single-molecule fluorescence microscopy. We find that unused miRNAs stably bind to PBs, whereas functional miRNAs, repressed mRNAs, and lncRNAs both transiently and stably localize within either the core or periphery of PBs, albeit to different extents. Consequently, translation potential and 3' versus 5' placement of miRNA target sites significantly affect the PB localization dynamics of mRNAs. Using computational modeling and supporting experimental approaches, we show that partitioning in the PB phase attenuates mRNA silencing, suggesting that physiological mRNA turnover occurs predominantly outside of PBs. Instead, our data support a PB role in sequestering unused miRNAs for surveillance and provide a framework for investigating the dynamic assembly of RNP granules by phase separation at single-molecule resolution.


Assuntos
MicroRNAs/genética , RNA Longo não Codificante/genética , RNA Mensageiro/genética , Ribonucleoproteínas/genética , Grânulos Citoplasmáticos/genética , Inativação Gênica , Células HeLa , Humanos , Processamento Pós-Transcricional do RNA/genética , RNA não Traduzido/genética , Imagem Individual de Molécula
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