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1.
BMC Genomics ; 22(1): 642, 2021 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-34482814

RESUMO

BACKGROUND: RNA polymerase II plays critical roles in transcription in eukaryotic organisms. C-terminal Domain Phosphatase-like 1 (CPL1) regulates the phosphorylation state of the C-terminal domain of RNA polymerase II subunit B1, which is critical in determining RNA polymerase II activity. CPL1 plays an important role in miRNA biogenesis, plant growth and stress responses. Although cpl1 mutant showes delayed-flowering phenotype, the molecular mechanism behind CPL1's role in floral transition is still unknown. RESULTS: To study the role of CPL1 during the floral transition, we first tested phenotypes of cpl1-3 mutant, which harbors a point-mutation. The cpl1-3 mutant contains a G-to-A transition in the second exon, which results in an amino acid substitution from Glu to Lys (E116K). Further analyses found that the mutated amino acid (Glu) was conserved in these species. As a result, we found that the cpl1-3 mutant experienced delayed flowering under both long- and short-day conditions, and CPL1 is involved in the vernalization pathway. Transcriptome analysis identified 109 genes differentially expressed in the cpl1 mutant, with 2 being involved in floral transition. Differential expression of the two flowering-related DEGs was further validated by qRT-PCR. CONCLUSIONS: Flowering genetic pathways analysis coupled with transciptomic analysis provides potential genes related to floral transition in the cpl1-3 mutant, and a framework for future studies of the molecular mechanisms behind CPL1's role in floral transition.


Assuntos
Proteínas de Arabidopsis , Arabidopsis , Flores/fisiologia , Arabidopsis/genética , Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Flores/genética , Regulação da Expressão Gênica de Plantas , Mutação , Fosfoproteínas Fosfatases/genética , Proteínas de Ligação a RNA/genética , Fatores de Transcrição/metabolismo
2.
Science ; 373(6557): 882-889, 2021 08 20.
Artigo em Inglês | MEDLINE | ID: mdl-34413232

RESUMO

Eukaryotic genomes contain domesticated genes from integrating viruses and mobile genetic elements. Among these are homologs of the capsid protein (known as Gag) of long terminal repeat (LTR) retrotransposons and retroviruses. We identified several mammalian Gag homologs that form virus-like particles and one LTR retrotransposon homolog, PEG10, that preferentially binds and facilitates vesicular secretion of its own messenger RNA (mRNA). We showed that the mRNA cargo of PEG10 can be reprogrammed by flanking genes of interest with Peg10's untranslated regions. Taking advantage of this reprogrammability, we developed selective endogenous encapsidation for cellular delivery (SEND) by engineering both mouse and human PEG10 to package, secrete, and deliver specific RNAs. Together, these results demonstrate that SEND is a modular platform suited for development as an efficient therapeutic delivery modality.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Ligação a DNA/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/metabolismo , Animais , Proteínas Reguladoras de Apoptose/química , Proteínas Reguladoras de Apoptose/genética , Capsídeo/metabolismo , Linhagem Celular , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Vesículas Extracelulares/metabolismo , Edição de Genes , Vetores Genéticos , Humanos , Camundongos , Neurônios/metabolismo , Domínios Proteicos , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/genética , Retroelementos , Transfecção , Regiões não Traduzidas , Regulação para Cima
3.
Nat Commun ; 12(1): 4980, 2021 08 17.
Artigo em Inglês | MEDLINE | ID: mdl-34404792

RESUMO

Proximity labeling (PL) with genetically-targeted promiscuous enzymes has emerged as a powerful tool for unbiased proteome discovery. By combining the spatiotemporal specificity of PL with methods for functional protein enrichment, we show that it is possible to map specific protein subclasses within distinct compartments of living cells. In particular, we develop a method to enrich subcompartment-specific RNA binding proteins (RBPs) by combining peroxidase-catalyzed PL with organic-aqueous phase separation of crosslinked protein-RNA complexes ("APEX-PS"). We use APEX-PS to generate datasets of nuclear, nucleolar, and outer mitochondrial membrane (OMM) RBPs, which can be mined for novel functions. For example, we find that the OMM RBP SYNJ2BP retains specific nuclear-encoded mitochondrial mRNAs at the OMM during translation stress, facilitating their local translation and import of protein products into the mitochondrion during stress recovery. Functional PL in general, and APEX-PS in particular, represent versatile approaches for the discovery of proteins with novel function in specific subcellular compartments.


Assuntos
RNA Mensageiro/metabolismo , RNA Mitocondrial/metabolismo , Proteínas de Ligação a RNA/metabolismo , Técnicas de Inativação de Genes , Células HEK293 , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Mitocôndrias/metabolismo , Membranas Mitocondriais/metabolismo , Fosforilação , Proteoma/metabolismo , Proteômica , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética
4.
Mol Cell ; 81(15): 3041-3042, 2021 08 05.
Artigo em Inglês | MEDLINE | ID: mdl-34358456

RESUMO

Einstein et al. (2021) uncover a novel role for the RNA-binding protein YTHDF2, one of the m6A reader proteins, in TNBC proliferation and survival. This study demonstrates the clinical potential of targeting a specific reader protein in the treatment of breast cancer.


Assuntos
Proteínas de Ligação a RNA , Proteínas de Ligação a RNA/genética
5.
Nat Commun ; 12(1): 4696, 2021 08 04.
Artigo em Inglês | MEDLINE | ID: mdl-34349113

RESUMO

Productive ribosomal RNA (rRNA) compaction during ribosome assembly necessitates establishing correct tertiary contacts between distant secondary structure elements. Here, we quantify the response of the yeast proteome to low temperature (LT), a condition where aberrant mis-paired RNA folding intermediates accumulate. We show that, at LT, yeast cells globally boost production of their ribosome assembly machinery. We find that the LT-induced assembly factor, Puf6, binds to the nascent catalytic RNA-rich subunit interface within the 60S pre-ribosome, at a site that eventually loads the nuclear export apparatus. Ensemble Förster resonance energy transfer studies show that Puf6 mimics the role of Mg2+ to usher a unique long-range tertiary contact to compact rRNA. At LT, puf6 mutants accumulate 60S pre-ribosomes in the nucleus, thus unveiling Puf6-mediated rRNA compaction as a critical temperature-regulated rescue mechanism that counters rRNA misfolding to prime export competence.


Assuntos
Núcleo Celular/metabolismo , Proteínas de Ligação a RNA/metabolismo , Subunidades Ribossômicas Maiores de Eucariotos/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Transporte Ativo do Núcleo Celular , Temperatura Baixa , GTP Fosfo-Hidrolases/metabolismo , Mutação , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Proteoma/metabolismo , Dobramento de RNA , Precursores de RNA/química , Precursores de RNA/metabolismo , RNA Ribossômico/química , RNA Ribossômico/metabolismo , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/genética , Subunidades Ribossômicas Maiores de Eucariotos/química , Ribossomos/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/fisiologia , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética
6.
Nucleic Acids Res ; 49(15): 8535-8555, 2021 09 07.
Artigo em Inglês | MEDLINE | ID: mdl-34358317

RESUMO

Gene deletion and gene expression alteration can lead to growth defects that are amplified or reduced when a second mutation is present in the same cells. We performed 154 genetic interaction mapping (GIM) screens with query mutants related with RNA metabolism and estimated the growth rates of about 700 000 double mutant Saccharomyces cerevisiae strains. The tested targets included the gene deletion collection and 900 strains in which essential genes were affected by mRNA destabilization (DAmP). To analyze the results, we developed RECAP, a strategy that validates genetic interaction profiles by comparison with gene co-citation frequency, and identified links between 1471 genes and 117 biological processes. In addition to these large-scale results, we validated both enhancement and suppression of slow growth measured for specific RNA-related pathways. Thus, negative genetic interactions identified a role for the OCA inositol polyphosphate hydrolase complex in mRNA translation initiation. By analysis of suppressors, we found that Puf4, a Pumilio family RNA binding protein, inhibits ribosomal protein Rpl9 function, by acting on a conserved UGUAcauUA motif located downstream the stop codon of the RPL9B mRNA. Altogether, the results and their analysis should represent a useful resource for discovery of gene function in yeast.


Assuntos
Genes Fúngicos , RNA Fúngico/metabolismo , Saccharomyces cerevisiae/genética , Alelos , Deleção de Genes , Pleiotropia Genética , Fosfatos de Inositol/metabolismo , Iniciação Traducional da Cadeia Peptídica , Estabilidade de RNA , RNA Mensageiro/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/fisiologia , Proteínas Ribossômicas/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/fisiologia
7.
BMC Res Notes ; 14(1): 311, 2021 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-34391474

RESUMO

OBJECTIVE: RNA-binding proteins (RBPs) are important regulators of gene expression that influence mRNA splicing, stability, localization, transport, and translational control. In particular, RBPs play an important role in neurons, which have a complex morphology. Previously, we showed that there are many RBPs that play a conserved role in dendrite development in Drosophila dendritic arborization neurons and Caenorhabditis elegans (C. elegans) PVD neurons including the cytoplasmic polyadenylation element binding proteins (CPEBs), Orb in Drosophila and CPB-3 in C. elegans, and the DEAD box RNA helicases, Me31B in Drosophila and CGH-1 in C. elegans. During these studies, we observed that fluorescently-labeled CPB-3 and CGH-1 localize to cytoplasmic particles that are motile, and our research aims to further characterize these RBP-containing particles in live neurons. RESULTS: Here we extend on previous work to show that CPB-3 and CGH-1 localize to motile particles within dendrites that move at a speed consistent with microtubule-based transport. This is consistent with a model in which CPB-3 and CGH-1 influence dendrite development through the transport and localization of their mRNA targets. Moreover, CPB-3 and CGH-1 rarely localize to the same particles suggesting that these RBPs function in discrete ribonucleoprotein particles (RNPs) that may regulate distinct mRNAs.


Assuntos
Proteínas de Caenorhabditis elegans , Caenorhabditis elegans , Animais , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Dendritos , RNA Nucleotidiltransferases , Proteínas de Ligação a RNA/genética , Células Receptoras Sensoriais/metabolismo
8.
Nat Commun ; 12(1): 4872, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34381052

RESUMO

The Netrin-1 receptor UNC5B is an axon guidance regulator that is also expressed in endothelial cells (ECs), where it finely controls developmental and tumor angiogenesis. In the absence of Netrin-1, UNC5B induces apoptosis that is blocked upon Netrin-1 binding. Here, we identify an UNC5B splicing isoform (called UNC5B-Δ8) expressed exclusively by ECs and generated through exon skipping by NOVA2, an alternative splicing factor regulating vascular development. We show that UNC5B-Δ8 is a constitutively pro-apoptotic splicing isoform insensitive to Netrin-1 and required for specific blood vessel development in an apoptosis-dependent manner. Like NOVA2, UNC5B-Δ8 is aberrantly expressed in colon cancer vasculature where its expression correlates with tumor angiogenesis and poor patient outcome. Collectively, our data identify a mechanism controlling UNC5B's necessary apoptotic function in ECs and suggest that the NOVA2/UNC5B circuit represents a post-transcriptional pathway regulating angiogenesis.


Assuntos
Apoptose , Vasos Sanguíneos/crescimento & desenvolvimento , Receptores de Netrina/metabolismo , Isoformas de RNA/metabolismo , Processamento Alternativo , Animais , Neoplasias do Colo/irrigação sanguínea , Neoplasias do Colo/metabolismo , Células Endoteliais , Humanos , Morfogênese , Neovascularização Patológica/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Receptores de Netrina/genética , Netrina-1/metabolismo , Isoformas de RNA/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Análise de Sobrevida , Peixe-Zebra
9.
Nat Commun ; 12(1): 4909, 2021 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-34389707

RESUMO

In bacteria, trans-translation is the main rescue system, freeing ribosomes stalled on defective messenger RNAs. This mechanism is driven by small protein B (SmpB) and transfer-messenger RNA (tmRNA), a hybrid RNA known to have both a tRNA-like and an mRNA-like domain. Here we present four cryo-EM structures of the ribosome during trans-translation at resolutions from 3.0 to 3.4 Å. These include the high-resolution structure of the whole pre-accommodated state, as well as structures of the accommodated state, the translocated state, and a translocation intermediate. Together, they shed light on the movements of the tmRNA-SmpB complex in the ribosome, from its delivery by the elongation factor EF-Tu to its passage through the ribosomal A and P sites after the opening of the B1 bridges. Additionally, we describe the interactions between the tmRNA-SmpB complex and the ribosome. These explain why the process does not interfere with canonical translation.


Assuntos
Proteínas de Escherichia coli/genética , Escherichia coli/genética , Biossíntese de Proteínas/genética , RNA Bacteriano/genética , Proteínas de Ligação a RNA/genética , Ribossomos/genética , Sítios de Ligação/genética , Microscopia Crioeletrônica , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Modelos Moleculares , Conformação de Ácido Nucleico , Ligação Proteica , Domínios Proteicos , RNA Bacteriano/química , RNA Bacteriano/metabolismo , RNA Mensageiro/química , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA de Transferência/química , RNA de Transferência/genética , RNA de Transferência/metabolismo , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/metabolismo , Ribossomos/metabolismo , Ribossomos/ultraestrutura
10.
Nat Commun ; 12(1): 4908, 2021 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-34389711

RESUMO

C9ORF72 hexanucleotide GGGGCC repeat expansion is the most common genetic cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Repeat-containing RNA mediates toxicity through nuclear granules and dipeptide repeat (DPR) proteins produced by repeat-associated non-AUG translation. However, it remains unclear how the intron-localized repeats are exported and translated in the cytoplasm. We use single molecule imaging approach to examine the molecular identity and spatiotemporal dynamics of the repeat RNA. We demonstrate that the spliced intron with G-rich repeats is stabilized in a circular form due to defective lariat debranching. The spliced circular intron, instead of pre-mRNA, serves as the translation template. The NXF1-NXT1 pathway plays an important role in the nuclear export of the circular intron and modulates toxic DPR production. This study reveals an uncharacterized disease-causing RNA species mediated by repeat expansion and demonstrates the importance of RNA spatial localization to understand disease etiology.


Assuntos
Proteína C9orf72/genética , Núcleo Celular/metabolismo , Íntrons/genética , Biossíntese de Proteínas/genética , RNA/genética , Transporte Ativo do Núcleo Celular/genética , Esclerose Amiotrófica Lateral/genética , Esclerose Amiotrófica Lateral/metabolismo , Proteína C9orf72/metabolismo , Linhagem Celular Tumoral , Núcleo Celular/genética , Expansão das Repetições de DNA/genética , Dipeptídeos/genética , Dipeptídeos/metabolismo , Demência Frontotemporal/genética , Demência Frontotemporal/metabolismo , Predisposição Genética para Doença/genética , Células HEK293 , Humanos , Microscopia de Fluorescência , Proteínas de Transporte Nucleocitoplasmático/genética , Proteínas de Transporte Nucleocitoplasmático/metabolismo , RNA/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/genética
11.
Cancer Sci ; 112(9): 3835-3845, 2021 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-34219323

RESUMO

Tumor-induced angiogenesis is important for further progression of solid tumors. The initiation of tumor angiogenesis is dictated by a shift in the balance between proangiogenic and antiangiogenic gene expression programs. However, the potential mechanism controlling the expression of angiogenesis-related genes in the tumor cells, especially the process mediated by RNA-binding protein (RBP) remains unclear. SAMD4A is a conserved RBP across fly to mammals, and is believed to play an important role in controlling gene translation and stability. In this study, we identified the potential role of SAMD4A in modulating angiogenesis-related gene expression and tumor progression in breast cancer. SAMD4A expression was repressed in breast cancer tissues and cells and low SAMD4A expression in human breast tumor samples was strongly associated with poor survival of patients. Overexpression of SAMD4A inhibited breast tumor angiogenesis and caner progression, whereas knockdown of SAMD4A demonstrated a reversed effect. Mechanistically, SAMD4A was found to specifically destabilize the proangiogenic gene transcripts, including C-X-C motif chemokine ligand 5 (CXCL5), endoglin (ENG), interleukin 1ß (IL1ß), and angiopoietin 1 (ANGPT1), by directly interacting with the stem-loop structure in the 3' untranslated region (3'UTR) of these mRNAs through its sterile alpha motif (SAM) domain, resulting in the imbalance of angiogenic genes expression. Collectively, our results suggest that SAMD4A is a novel breast tumor suppressor that inhibits tumor angiogenesis by specifically downregulating the expression of proangiogenic genes, which might be a potential antiangiogenic target for breast cancer therapy.


Assuntos
Neoplasias da Mama/irrigação sanguínea , Regulação Neoplásica da Expressão Gênica , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas Repressoras/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Animais , Progressão da Doença , Feminino , Células HEK293 , Humanos , Células MCF-7 , Glândulas Mamárias Humanas/citologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas de Ligação a RNA/genética , Proteínas Repressoras/genética , Transfecção , Carga Tumoral/genética , Proteínas Supressoras de Tumor/genética , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Mol Cell ; 81(15): 3082-3095.e6, 2021 08 05.
Artigo em Inglês | MEDLINE | ID: mdl-34197738

RESUMO

To understand how chromatin domains coordinate gene expression, we dissected select genetic elements organizing topology and transcription around the Prdm14 super enhancer in mouse embryonic stem cells. Taking advantage of allelic polymorphisms, we developed methods to sensitively analyze changes in chromatin topology, gene expression, and protein recruitment. We show that enhancer insulation does not rely strictly on loop formation between its flanking boundaries, that the enhancer activates the Slco5a1 gene beyond its prominent domain boundary, and that it recruits cohesin for loop extrusion. Upon boundary inversion, we find that oppositely oriented CTCF terminates extrusion trajectories but does not stall cohesin, while deleted or mutated CTCF sites allow cohesin to extend its trajectory. Enhancer-mediated gene activation occurs independent of paused loop extrusion near the gene promoter. We expand upon the loop extrusion model to propose that cohesin loading and extrusion trajectories originating at an enhancer contribute to gene activation.


Assuntos
Fator de Ligação a CCCTC/metabolismo , Cromatina/genética , Elementos Facilitadores Genéticos , Animais , Fator de Ligação a CCCTC/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Cromatina/metabolismo , Proteínas Cromossômicas não Histona/genética , Proteínas Cromossômicas não Histona/metabolismo , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Expressão Gênica , Camundongos , Células-Tronco Embrionárias Murinas , Coativador 2 de Receptor Nuclear/genética , Regiões Promotoras Genéticas , Proteínas de Ligação a RNA/genética , Fatores de Transcrição/genética
13.
Mol Cell ; 81(15): 3048-3064.e9, 2021 08 05.
Artigo em Inglês | MEDLINE | ID: mdl-34216543

RESUMO

RNA-binding proteins (RBPs) are critical regulators of post-transcriptional gene expression, and aberrant RBP-RNA interactions can promote cancer progression. Here, we interrogate the function of RBPs in cancer using pooled CRISPR-Cas9 screening and identify 57 RBP candidates with distinct roles in supporting MYC-driven oncogenic pathways. We find that disrupting YTHDF2-dependent mRNA degradation triggers apoptosis in triple-negative breast cancer (TNBC) cells and tumors. eCLIP and m6A sequencing reveal that YTHDF2 interacts with mRNAs encoding proteins in the MAPK pathway that, when stabilized, induce epithelial-to-mesenchymal transition and increase global translation rates. scRibo-STAMP profiling of translating mRNAs reveals unique alterations in the translatome of single cells within YTHDF2-depleted solid tumors, which selectively contribute to endoplasmic reticulum stress-induced apoptosis in TNBC cells. Thus, our work highlights the therapeutic potential of RBPs by uncovering a critical role for YTHDF2 in counteracting the global increase of mRNA synthesis in MYC-driven breast cancers.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proteínas de Ligação a RNA/genética , Adenosina/análogos & derivados , Adenosina/metabolismo , Animais , Morte Celular/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Genes myc , Humanos , Camundongos Nus , Camundongos Transgênicos , Biossíntese de Proteínas , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
14.
Int J Mol Sci ; 22(14)2021 Jul 12.
Artigo em Inglês | MEDLINE | ID: mdl-34299096

RESUMO

Circular RNAs (circRNAs) are increasingly recognized as having a role in cancer development. Their expression is modified in numerous cancers, including hepatocellular carcinoma (HCC); however, little is known about the mechanisms of their regulation. The aim of this study was to identify regulators of circRNAome expression in HCC. Using publicly available datasets, we identified RNA binding proteins (RBPs) with enriched motifs around the splice sites of differentially expressed circRNAs in HCC. We confirmed the binding of some of the candidate RBPs using ChIP-seq and eCLIP datasets in the ENCODE database. Several of the identified RBPs were found to be differentially expressed in HCC and/or correlated with the overall survival of HCC patients. According to our bioinformatics analyses and published evidence, we propose that NONO, PCPB2, PCPB1, ESRP2, and HNRNPK are candidate regulators of circRNA expression in HCC. We confirmed that the knocking down the epithelial splicing regulatory protein 2 (ESRP2), known to be involved in the maintenance of the adult liver phenotype, significantly changed the expression of candidate circRNAs in a model HCC cell line. By understanding the systemic changes in transcriptome splicing, we can identify new proteins involved in the molecular pathways leading to HCC development and progression.


Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/patologia , Regulação Neoplásica da Expressão Gênica , Redes Reguladoras de Genes , Neoplasias Hepáticas/patologia , RNA Circular/genética , Proteínas de Ligação a RNA/metabolismo , Apoptose , Biomarcadores Tumorais/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Proliferação de Células , Biologia Computacional , Perfilação da Expressão Gênica , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , MicroRNAs/genética , Prognóstico , Proteínas de Ligação a RNA/genética , Taxa de Sobrevida , Células Tumorais Cultivadas
15.
Nat Commun ; 12(1): 4288, 2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34257283

RESUMO

The commonly mutated human KRAS oncogene encodes two distinct KRAS4A and KRAS4B proteins generated by differential splicing. We demonstrate here that coordinated regulation of both isoforms through control of splicing is essential for development of Kras mutant tumors. The minor KRAS4A isoform is enriched in cancer stem-like cells, where it responds to hypoxia, while the major KRAS4B is induced by ER stress. KRAS4A splicing is controlled by the DCAF15/RBM39 pathway, and deletion of KRAS4A or pharmacological inhibition of RBM39 using Indisulam leads to inhibition of cancer stem cells. Our data identify existing clinical drugs that target KRAS4A splicing, and suggest that levels of the minor KRAS4A isoform in human tumors can be a biomarker of sensitivity to some existing cancer therapeutics.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Células-Tronco Neoplásicas/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Proteínas de Ligação a RNA/metabolismo , Células A549 , Animais , Western Blotting , Proliferação de Células , Citometria de Fluxo , Xenoenxertos , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Camundongos , Camundongos Knockout , Reação em Cadeia da Polimerase , Proteínas Proto-Oncogênicas p21(ras)/genética , Proteínas de Ligação a RNA/genética
16.
Int J Mol Sci ; 22(11)2021 Jun 02.
Artigo em Inglês | MEDLINE | ID: mdl-34199367

RESUMO

TDP-43 is an RNA-binding protein that has been robustly linked to the pathogenesis of a number of neurodegenerative disorders, including amyotrophic lateral sclerosis and frontotemporal dementia. While mutations in the TARDBP gene that codes for the protein have been identified as causing disease in a small subset of patients, TDP-43 proteinopathy is present in the majority of cases regardless of mutation status. This raises key questions regarding the mechanisms by which TDP-43 proteinopathy arises and spreads throughout the central nervous system. Numerous studies have explored the role of a variety of cellular functions on the disease process, and nucleocytoplasmic transport, protein homeostasis, RNA interactions and cellular stress have all risen to the forefront as possible contributors to the initiation of TDP-43 pathogenesis. There is also a small but growing body of evidence suggesting that aggregation-prone TDP-43 can recruit physiological TDP-43, and be transmitted intercellularly, providing a mechanism whereby small-scale proteinopathy spreads from cell to cell, reflecting the spread of clinical symptoms observed in patients. This review will discuss the potential role of the aforementioned cellular functions in TDP-43 pathogenesis, and explore how aberrant pathology may spread, and result in a feed-forward cascade effect, leading to robust TDP-43 proteinopathy and disease.


Assuntos
Proteínas de Ligação a DNA/genética , Agregação Patológica de Proteínas/genética , Proteínas de Ligação a RNA/genética , Proteinopatias TDP-43/genética , Esclerose Amiotrófica Lateral/genética , Esclerose Amiotrófica Lateral/patologia , Demência Frontotemporal/genética , Demência Frontotemporal/patologia , Humanos , Mutação/genética , Proteinopatias TDP-43/patologia
17.
Int J Mol Sci ; 22(13)2021 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-34203267

RESUMO

Insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1) is an RNA-binding protein and serves as a post-transcriptional fine-tuner regulating the expression of mRNA targets. However, the clinicopathological roles of IGF2BP1 in colorectal cancer (CRC) remains limited. Thus, we aimed to elucidate the clinical significance and biomarker potentials of IGF2BP1 in CRC. A total of 266 specimens from two sets of CRC patients were collected. IGF2BP1 expression was studied by immunohistochemical (IHC) staining. The Kaplan-Meier survival plot and a log-rank test were used for survival analysis. The Cox proportional hazards model was applied to determine the survival impact of IGF2BP1. Public datasets sets from The Cancer Genome Atlas (TCGA) and Human Cancer Metastasis Database (HCMDB), receiver operating characteristic (ROC) plotter, and two CRC cell lines, HCT-116 and DLD-1, were used for validating our findings. We showed that IGF2BP1 was overexpressed in tumor specimens compared to 13 paired normal parts by examining the immunoreactivity of IGF2BP1 (p = 0.045). The increased expression of IGF2BP1 in primary tumor parts was observed regardless of metastatic status (p < 0.001) in HCMDB analysis. IGF2BP1 expression was significantly associated with young age (59.6% vs. 46.7%, p-value = 0.043) and advanced stage (61.3% vs. 40.0%, p-value = 0.001). After controlling for confounding factors, IGF2BP1 remained an independent prognostic factor (HR = 1.705, p-value = 0.005). TCGA datasets analysis indicated that high IGF2BP1 expression showed a lower 5-year survival rate (58% vs. 65%) in CRC patients. The increased expression of IGF2BP1 in chemotherapy non-responder rectal cancer patients was observed using a ROC plotter. Overexpression of IGF2BP1 promoted the colony-forming capacity and 5-fluorouracil and etoposide resistance in CRC cells. Here, IGF2BP1 was an independent poor prognostic marker in CRC patients and contributed to aggressive phenotypes in CRC cell lines.


Assuntos
Biomarcadores/metabolismo , Neoplasias Colorretais/metabolismo , Proteínas de Ligação a RNA/metabolismo , Biomarcadores/química , Neoplasias Colorretais/genética , Células HCT116 , Humanos , Estimativa de Kaplan-Meier , Prognóstico , Modelos de Riscos Proporcionais , Proteínas de Ligação a RNA/genética , Curva ROC
18.
Biomed Pharmacother ; 139: 111719, 2021 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-34233389

RESUMO

Poly(rC)-binding protein 2 (PCBP2) is an RNA-binding protein that is characterized by its ability to interact with poly(C) with high affinity in a sequence-specific manner. PCBP2 contains three K homology domains, which are consensus RNA-binding domains that play a role in recognizing and combining with RNA and DNA. The specific structure and localization of PCBP2 lay the foundation for its multiple roles in transcriptional, posttranscriptional, and translational processes, even in iron metabolism. Numerous studies have indicated that PCBP2 expression is increased in many cancer types. PCBP2 is considered as an oncogene that promotes tumorigenesis, development of cancer cells, and metastasis. Here, we summarized the current evidence regarding PCBP2 in the proliferation, migration, invasion of cancer cells, and drug resistance, aiming to clarify the molecular mechanisms of PCBP2 in cancer. Results from this review suggest that an in-depth study of PCBP2 in cancer may provide novel biomarkers for prognostic or therapeutic purposes.


Assuntos
Neoplasias/metabolismo , Proteínas de Ligação a RNA , Animais , Expressão Gênica , Humanos , Neoplasias/genética , Neoplasias/mortalidade , Prognóstico , Proteínas de Ligação a RNA/química , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo
19.
J Coll Physicians Surg Pak ; 31(8): 937-940, 2021 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-34320711

RESUMO

OBJECTIVE: To investigate the expression of miR-22-3p in breast cancer and the mechanism of targeting PLAGL2 to inhibit the invasion and migration in human breast cancer. STUDY DESIGN: An experimental study. PLACE AND DURATION OF STUDY: Department of Oncology and Department of General Surgery, The People's Hospital of China Three Gorges University, China, from March 2019 to December 2020. METHODOLOGY: The miR-22-3p expression level in 41 paired human primary breast invasive ductal carcinoma tissues and para-cancer tissues was obtained by real-time fluorescence quantitative reverse transcriptase PCR (qRT-PCR). The effect of miR-22-3p on the proliferation of breast cancer cells was detected by growth curve method. Online software TargetScan was used to predict the target genes of miR-22-3p. The prediction results were verified by luciferase reporter gene assay and qRT⁃PCR. RESULTS: MiR-22-3p expression was significantly decreased in the breast cancer tissues than in para⁃carcinoma normal breast tissues (p<0.05). Over-expression of miR-22-3p can inhibit the proliferation of MCF-7 cells significantly. Pleomorphic adenoma gene-like protein 2(PLAGL2) is the predicted target gene of miR-22-3p. MiR-22-3p binds to its predicted target gene PLAGL2-3'UTR. The expression of miR-22-3p was negatively correlated with PLAGL2 in MCF-7 cells. CONCLUSION: MiR-22-3p could suppress the proliferation of breast cancer by targeting PLAGL2. This suggests that miR-22-3p may be a strategy of choice for targeted therapy of breast cancer. Key Words: Breast cancer, MiR-22-3p, PLAGL2, Cell proliferation.


Assuntos
Neoplasias da Mama , MicroRNAs , Neoplasias da Mama/genética , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , China , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , MicroRNAs/genética , Proteínas de Ligação a RNA/genética , Fatores de Transcrição/genética
20.
Viruses ; 13(7)2021 06 25.
Artigo em Inglês | MEDLINE | ID: mdl-34202160

RESUMO

The product of the interferon-stimulated gene C19orf66, Shiftless (SHFL), restricts human immunodeficiency virus replication through downregulation of the efficiency of the viral gag/pol frameshifting signal. In this study, we demonstrate that bacterially expressed, purified SHFL can decrease the efficiency of programmed ribosomal frameshifting in vitro at a variety of sites, including the RNA pseudoknot-dependent signals of the coronaviruses IBV, SARS-CoV and SARS-CoV-2, and the protein-dependent stimulators of the cardioviruses EMCV and TMEV. SHFL also reduced the efficiency of stop-codon readthrough at the murine leukemia virus gag/pol signal. Using size-exclusion chromatography, we confirm the binding of the purified protein to mammalian ribosomes in vitro. Finally, through electrophoretic mobility shift assays and mutational analysis, we show that expressed SHFL has strong RNA binding activity that is necessary for full activity in the inhibition of frameshifting, but shows no clear specificity for stimulatory RNA structures.


Assuntos
Códon de Terminação/genética , Coronavirus/genética , Mudança da Fase de Leitura do Gene Ribossômico/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas Virais/metabolismo , Sequência de Bases , Escherichia coli/genética , Regulação Viral da Expressão Gênica , Humanos , Vírus da Leucemia Murina/genética , Proteínas com Motivo de Reconhecimento de RNA , RNA Viral/genética , Replicação Viral
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