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1.
Life Sci ; 240: 117019, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31678554

RESUMO

AIMS: Long noncoding RNA melanotransferrin antisense RNA (MFI2-AS1) plays a vital role in the development of multiple diseases. This study aimed to investigate the effect of this lncRNA on osteoarthritis progression and explore the interaction among MFI2-AS1, microRNA (miR)-130a-3p and transcription factor 4 (TCF4). METHODS: Forty-six knee osteoarthritis tissues and 28 normal samples were collected. Human chondrocytes C28/I2 cells treated by lipopolysaccharide (LPS) were used as the model of osteoarthritis. The expression levels of MFI2-AS1, miR-130a-3p and TCF4 were detected by quantitative real-time polymerase chain reaction or western blot. LPS-induced chondrocytes injury was investigated by cell viability, apoptosis, inflammatory response and extracellular matrix degradation using MTT, flow cytometry, enzyme-linked immunosorbent assay and western blot. The target association between miR-130a-3p and MFI2-AS1 or TCF4 was confirmed by luciferase reporter assay and RNA immunoprecipitation. RESULTS: MFI2-AS1 expression was increased in osteoarthritis tissues and LPS-treated C28/I2 cells. Silence of MFI2-AS1 attenuated LPS-induced viability suppression, apoptosis production, inflammatory response and extracellular matrix degradation. MFI2-AS1 was validated as a decoy of miR-130a-3p and TCF4 was confirmed as a target of miR-130a-3p. miR-130a-3p overexpression inhibited LPS-induced cell injury in C28/I2 cells by decreasing TCF4 expression. Moreover, knockdown of MFI2-AS1 alleviated LPS-induced cell injury in C28/I2 cells by mediating miR-130a-3p and TCF4. CONCLUSION: Knockdown of MFI2-AS1 increased cell viability but suppressed apoptosis, inflammatory response and extracellular matrix degradation in LPS-treated chondrocytes by increasing miR-130a-3p and decreasing TCF4, indicating a novel target for the treatment of osteoarthritis.


Assuntos
Técnicas de Silenciamento de Genes , Lipopolissacarídeos , MicroRNAs/genética , Osteoartrite/genética , Osteoartrite/prevenção & controle , RNA Antissenso/genética , RNA Longo não Codificante/genética , Fator de Transcrição 4/genética , Animais , Linhagem Celular , Proliferação de Células , Sobrevivência Celular , Condrócitos , Humanos
2.
DNA Cell Biol ; 38(9): 1013-1021, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31386568

RESUMO

Increasing evidence has indicated that long noncoding RNAs (lncRNAs) could participate in diverse cancers. Among these, lymphoid enhancer-binding factor 1 antisense RNA 1 (LEF1-AS1) was recently identified as an oncogenic lncRNA, but little is known about its function in non-small-cell lung cancer (NSCLC). In the present study, we found that LEF1-AS1 was markedly upregulated in lung cancer tissues and could promote NSCLC cell proliferation and migration in vivo and in vitro. LEF1-AS1 could bind with miR-489 and further negatively regulate miR-489 to promote SRY-related HMG box transcription factor 4 (SOX4) expression. In conclusion, these data suggested that LEF1-AS1 promoted NSCLC tumorigenesis dependent on the miR-489-SOX4 axis and implicated the potential application of LEF1-AS1 for the prognosis and treatment of NSCLC.


Assuntos
Carcinoma Pulmonar de Células não Pequenas/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Pulmonares/genética , Fator 1 de Ligação ao Facilitador Linfoide/genética , RNA Antissenso/genética , RNA Longo não Codificante/genética , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Feminino , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Fator 1 de Ligação ao Facilitador Linfoide/metabolismo , Masculino , MicroRNAs/genética , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Fatores de Transcrição SOXC/genética , Fatores de Transcrição SOXC/metabolismo , Regulação para Cima
3.
BMC Cancer ; 19(1): 771, 2019 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-31382922

RESUMO

BACKGROUND: Long non-coding RNAs (lncRNAs) represent a substantial portion of the human transcriptome. LncRNAs present a very stringent cell-type/tissue specificity being potential candidates for therapeutical applications during aging and disease. As example, targeting of MALAT1, a highly conserved lncRNA originally identified in metastatic non-small cell lung cancer, has shown promising results in cancer regression. Nevertheless, the regulation and specificity of MALAT1 have not been directly addressed. Interestingly, MALAT1 locus is spanned by an antisense transcript named TALAM1. METHODS: Here using a collection of breast cancer cells and in vitro and in vivo migration assays we characterized the dynamics of expression and demonstrated that TALAM1 regulates and synergizes with MALAT1 during tumorigenesis. RESULTS: Down-regulation of TALAM1 was shown to greatly impact on the capacity of breast cancer cells to migrate in vitro or to populate the lungs of immunocompromised mice. Additionally, we demonstrated that TALAM1 cooperates with MALAT1 in the regulation of the properties guiding breast cancer aggressiveness and malignancy. CONCLUSIONS: By characterizing this sense/anti-sense pair we uncovered the complexity of MALAT1 locus regulation, describing new potential candidates for cancer targeting.


Assuntos
Neoplasias da Mama/genética , Neoplasias da Mama/patologia , RNA Antissenso/genética , RNA Longo não Codificante/genética , Transcrição Genética/genética , Animais , Carcinogênese/genética , Movimento Celular , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Células HeLa , Humanos , Pulmão/patologia , Células MCF-7 , Camundongos , Camundongos SCID , Metástase Neoplásica , Transfecção , Transplante Heterólogo , Regulação para Cima/genética
4.
Malar J ; 18(1): 294, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31462239

RESUMO

BACKGROUND: Insecticides are still at the core of insect pest and vector control programmes. Several lines of evidence indicate that ABC transporters are involved in detoxification processes against insecticides, including permethrin and other pyrethroids. In particular, the ABCG4 gene, a member of the G subfamily, has consistently been shown to be up-regulated in response to insecticide treatments in the mosquito malaria vector Anopheles stephensi (both adults and larvae). METHODS: To verify the actual involvement of this transmembrane protein in the detoxification process of permethrin, bioassays on larvae of An. stephensi, combining the insecticide with a siRNA, specifically designed for the inhibition of ABCG4 gene expression were performed. Administration to larvae of the same siRNA, labeled with a fluorescent molecule, was effected to investigate the systemic distribution of the inhibitory RNA into the larval bodies. Based on siRNA results, similar experiments using antisense Vivo-Morpholinos (Vivo-MOs) were effected. These molecules, compared to siRNA, are expected to guarantee a higher stability in environmental conditions and in the insect gut, and present thus a higher potential for future in-field applications. RESULTS: Bioassays using two different concentrations of siRNA, associated with permethrin, led to an increase of larval mortality, compared with results with permethrin alone. These outcomes confirm that ABCG4 transporter plays a role in the detoxification process against the selected insecticide. Moreover, after fluorescent labelling, it was shown the systemic dissemination of siRNA in different body districts of An. stephensi larvae, which suggest a potential systemic effect of the molecule. At the same time, results of Vivo-MO experiments were congruent with those obtained using siRNA, thus confirming the potential of ABCG4 inhibition as a strategy to increase permethrin susceptibility in mosquitoes. For the first time, Vivo-MOs were administered in water to larvae, with evidence for a biological effect. CONCLUSIONS: Targeting ABCG4 gene for silencing through both techniques resulted in an increased pyrethroid efficacy. These results open the way toward the possibility to exploit ABCG4 inhibition in the context of integrated programmes for the control An. stephensi mosquitoes and malaria transmission.


Assuntos
Anopheles/genética , Resistência a Inseticidas/genética , Inseticidas , Morfolinos/administração & dosagem , Piretrinas , RNA Antissenso/genética , Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Animais , Bioensaio , Larva/genética , Malária/prevenção & controle , Morfolinos/genética , Controle de Mosquitos , Mosquitos Vetores , Interferência de RNA , RNA Interferente Pequeno
5.
Artif Cells Nanomed Biotechnol ; 47(1): 3172-3179, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31352804

RESUMO

We aimed to determine the roles and possible mechanism of long non-coding RNA SDPR-AS in the processes of non-small cell lung cancer (NSCLC). The expressions of SDPR-AS and SDPR in different subtypes of lung cancer (AC, SCC, LCC and SCLC) tissues and cells were determined. Three NSCLC cells were infused with pcDNA-SDPR-AS, pcDNA3.1, sh-SDPR-AS, sh-NC, si-SDPR and si-NC. The effects of SDPR-AS dysregulation on cell behaviors (including cell viability, colony forming ability, migration, invasion and apoptosis) were assessed. Moreover, the combined effects of SDPR-AS overexpression and SDPR-AS knockdown on H522 cell behaviors and the levels of p-p38 and p-ERK were investigated. SDPR-AS was lowly expressed in NSCLC tissues and cells, but had no changes in SCLC tissues and cells. Down-regulation of SDPR-AS enhanced the proliferation, migration and invasion and inhibited apoptosis of NSCLC cells (H522, H661 and H520). Overexpression of SDPR-AS exhibited opposite effects. Moreover, SDPR was positively regulated by SDPR-AS and effects of SDPR-AS on the cell biological processes of NSCLC cells were through regulation of SDPR. Besides, the levels of p-p38 and p-ERK were significantly decreased after SDPR-AS overexpression, which were dramatically changeover by SDPR knockdownsimultaneously. Our findings indicate that SDPR-AS was lowly expressed in NSCLC cells and down-regulation of SDPR-AS may promote the malignant behaviors of NSCLC cells possible through regulating SDPR expression and involving in p38 MAPK/ERK signaling pathway. SDPR-AS may serve as a prospective target for NSCLC diagnosis and therapy.


Assuntos
Carcinogênese/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/patologia , Sistema de Sinalização das MAP Quinases/genética , Proteínas de Ligação a Fosfato/genética , RNA Antissenso/genética , RNA Longo não Codificante/genética , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose/genética , Carcinoma Pulmonar de Células não Pequenas/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Neoplasias Pulmonares/genética , Masculino , Pessoa de Meia-Idade , Invasividade Neoplásica/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo
6.
BMC Genomics ; 20(1): 601, 2019 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-31331261

RESUMO

BACKGROUND: Long intergenic non-coding RNAs (lincRNAs) can act as regulators of expression of protein-coding genes. Trans-natural antisense transcripts (trans-NATs) are a type of lincRNAs that contain sequence complementary to mRNA from other loci. The regulatory potential of trans-NATs has been poorly studied in eukaryotes and no example of trans-NATs regulating gene expression in plants are reported. The goal of this study was to identify lincRNAs, and particularly trans-NATs, in Arabidopsis thaliana that have a potential to regulate expression of target genes in trans at the transcriptional or translational level. RESULTS: We identified 1001 lincRNAs using an RNAseq dataset from total polyA+ and polysome-associated RNA of seedlings grown under high and low phosphate, or shoots and roots treated with different phytohormones, of which 550 were differentially regulated. Approximately 30% of lincRNAs showed conservation amongst Brassicaceae and 25% harbored transposon element (TE) sequences. Gene co-expression network analysis highlighted a group of lincRNAs associated with the response of roots to low phosphate. A total of 129 trans-NATs were predicted, of which 88 were significantly differentially expressed under at least one pairwise comparison. Five trans-NATs showed a positive correlation between their expression and target mRNA steady-state levels, and three showed a negative correlation. Expression of four trans-NATs positively correlated with a change in target mRNA polysome association. The regulatory potential of these trans-NATs did not implicate miRNA mimics nor siRNAs. We also looked for lincRNAs that could regulate gene expression in trans by Watson-Crick DNA:RNA base pairing with target protein-encoding loci. We identified 100 and 81 with a positive or negative correlation, respectively, with steady-state level of their predicted target. The regulatory potential of one such candidate lincRNA harboring a SINE TE sequence was validated in a protoplast assay on three distinct genes containing homologous TE sequence in their promoters. Construction of networks highlighted other putative lincRNAs with multiple predicted target loci for which expression was positively correlated with target gene expression. CONCLUSIONS: This study identified lincRNAs in Arabidopsis with potential in regulating target gene expression in trans by both RNA:RNA and RNA:DNA base pairing and highlights lincRNAs harboring TE sequences in such activity.


Assuntos
Pareamento de Bases , RNA Antissenso/genética , RNA Longo não Codificante/genética , Cromatina/genética , Elementos de DNA Transponíveis/genética , Redes Reguladoras de Genes , Loci Gênicos/genética , Regiões Promotoras Genéticas/genética
7.
Asia Pac J Clin Oncol ; 15(5): e191-e196, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31309731

RESUMO

AIM: Nicotinamide Nucleotide Transhydrogenase (NNT) gene encodes a protein, which is an important antioxidative enzyme that converts NADH to NADPH. This enzyme provides a significant proportion of the entire NADPH resource in the mitochondria. Previous reports have shown possible contribution of this gene in the carcinogenesis process. METHODS: In the current research, we evaluated expression levels of NNT gene and a naturally occurring antisense RNA (NNT-AS1) in gastric cancer specimens compared to their corresponding adjacent noncancerous tissues (ANCTs). RESULTS: Both NNT1 and NNT-AS1 genes were significantly downregulated in tumor tissues compared to ANCTs (expression ratio = 0.369, p = .045 and expression ratio = 0.368, p = .043, respectively). Transcript levels of NNT1 and NNT-AS1 were associated with the location of the primary tumor (p = .003 and .002, respectively). Moreover, expressions of both genes were significantly elevated in tumors with lymphatic/vascular invasion compared to tumors without lymphatic/vascular invasion (p = .001 and p = .005). No other remarkable associations were noticed between transcript levels of genes in tumor tissues and patients' information. Based on the area under curve (AUC) values in the receiver operating characteristic (ROC) curves, the diagnostic power of NNT1 and NNT-AS1 were estimated to be 0.62 and 0.63, respectively. CONCLUSIONS: Although we demonstrated dysregulation of NNT1 and NNT-AS1 in gastric tumor specimens in association with clinical data of patients, these two genes are not supposed to be appropriate biomarkers for gastric cancer.


Assuntos
Biomarcadores Tumorais/metabolismo , NADP Trans-Hidrogenase Específica para A ou B/metabolismo , RNA Antissenso/metabolismo , Neoplasias Gástricas/patologia , Adolescente , Adulto , Biomarcadores Tumorais/genética , Estudos de Casos e Controles , Regulação para Baixo , Feminino , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas Mitocondriais/genética , Proteínas Mitocondriais/metabolismo , NADP Trans-Hidrogenase Específica para A ou B/genética , Prognóstico , RNA Antissenso/genética , Neoplasias Gástricas/enzimologia , Neoplasias Gástricas/genética , Adulto Jovem
8.
In Vitro Cell Dev Biol Anim ; 55(7): 522-532, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31264061

RESUMO

Emerging evidences exposed that long noncoding RNAs (lncRNAs) play important roles in various tumor progression including breast cancer (BC). However, the role of lncRNA ADP-dependent glucokinase antisense RNA 1 (ADPGK-AS1) in BC progression remains undiscovered. Hence, this study aimed to investigate the role of ADPGK-AS1 in BC. qRT-PCR was performed to investigate ADPGK-AS1 expression level in BC tissues and cell lines. The effect of ADPGK-AS1 knockdown on BC cellular process was assessed by loss-of-function assay. Luciferase reporter and RIP assay were performed to investigate the combination between ADPGK-AS1 and miR-3196. The combination between miR-3196 and orthodenticle homeobox 1 (OTX1) was verified by luciferase reporter assay. Finally, rescue assays were performed to confirm the effects of ADPGK-AS1/miR-3196/OTX1 axis on BC development. ADPGK-AS1 expression level was upregulated in BC tissues and cell lines. High expression of ADPGK-AS1 predicted poor prognosis for BC patients. Functionally, ADPGK-AS1 promoted cell proliferation, migration, induced epithelial-mesenchymal transition (EMT) process, and suppressed cell apoptosis. Mechanistically, ADPGK-AS1 acted as a miR-3196 sponge to release OTX1 in BC cells. Currently, ADPGK-AS1 acted as a competing endogenous RNA (ceRNA) via modulating miR-3196/OTX1 axis in BC.


Assuntos
Neoplasias da Mama/genética , Regulação Neoplásica da Expressão Gênica/genética , Glucoquinase/genética , MicroRNAs/genética , Fatores de Transcrição Otx/genética , RNA Longo não Codificante/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Transição Epitelial-Mesenquimal/genética , Feminino , Técnicas de Silenciamento de Genes , Humanos , Células MCF-7 , Pessoa de Meia-Idade , RNA Antissenso/genética
9.
Medicine (Baltimore) ; 98(24): e15982, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31192939

RESUMO

BACKGROUND: FEZ family zinc finger 1 antisense RNA 1 (FEZF1-AS1), as a novel lncRNA, was reported to be up-regulated in various cancers and involved in tumor progression. This study systematically assessed the prognostic value of FEZF1-AS1 in solid tumors. METHODS: Web of Science, PubMed, EMBASE, Chinese National Knowledge Infrastructure, and Wanfang databases were searched for eligible studies that evaluated the prognostic role of FEZF1-AS1 expression in cancer patients. Pooled hazard ratios (HRs) and combined odds ratios (ORs) with their 95% confidence intervals (CIs) were calculated. The meta-analysis was conducted using Stata/SE 14.1. RESULTS: Fifteen original studies involving 1378 patients were enrolled. Pooled results showed that increased expression of FEZF1-AS1 significantly correlated with shorter overall survival (OS) in cancer patients (HR 2.04, 95% CI 1.60-2.47), and also shorter disease-free survival (DFS) (HR 2.08, 95% CI 1.27-2.89). Additionally, the combined ORs indicated that increased FEZF1-AS1 expression was significantly associated with lymph node metastasis (OR 3.35, 95% CI 1.98-5.67), distant metastasis (OR 3.10, 95% CI 1.86-5.15), poor tumor differentiation (OR 2.90, 95% CI 1.45-5.80), high depth of tumor invasion (OR 2.72, 95% CI 1.36-5.43), and advanced clinical stage (OR 2.76, 95% CI 1.75-4.35). Expression analysis using the Gene Expression Profiling Interactive Analysis database indicated that the expression of FEZF1-AS1 was higher in tumor tissues than that in the corresponding normal tissues. The results of survival analysis revealed that increased FEZF1-AS1 expression was correlated with poor OS and DFS in cancer patients. CONCLUSIONS: LncRNA FEZF1-AS1 may serve as a valuable prognostic biomarker for clinical outcomes in various solid tumors.


Assuntos
Neoplasias/genética , RNA Antissenso/genética , RNA Longo não Codificante/genética , Fatores de Transcrição/genética , Biomarcadores Tumorais/biossíntese , Biomarcadores Tumorais/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Metástase Linfática , Neoplasias/metabolismo , Razão de Chances , Prognóstico , RNA Antissenso/biossíntese , Proteínas Repressoras , Análise de Sobrevida , Regulação para Cima
10.
Nucleic Acids Res ; 47(15): e88, 2019 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-31147705

RESUMO

Small non-coding RNAs (sRNAs) regulate numerous cellular processes in all domains of life. Several approaches have been developed to identify them from RNA-seq data, which are efficient for eukaryotic sRNAs but remain inaccurate for the longer and highly structured bacterial sRNAs. We present APERO, a new algorithm to detect small transcripts from paired-end bacterial RNA-seq data. In contrast to previous approaches that start from the read coverage distribution, APERO analyzes boundaries of individual sequenced fragments to infer the 5' and 3' ends of all transcripts. Since sRNAs are about the same size as individual fragments (50-350 nucleotides), this algorithm provides a significantly higher accuracy and robustness, e.g., with respect to spontaneous internal breaking sites. To demonstrate this improvement, we develop a comparative assessment on datasets from Escherichia coli and Salmonella enterica, based on experimentally validated sRNAs. We also identify the small transcript repertoire of Dickeya dadantii including putative intergenic RNAs, 5' UTR or 3' UTR-derived RNA products and antisense RNAs. Comparisons to annotations as well as RACE-PCR experimental data confirm the precision of the detected transcripts. Altogether, APERO outperforms all existing methods in terms of sRNA detection and boundary precision, which is crucial for comprehensive genome annotations. It is freely available as an open source R package on https://github.com/Simon-Leonard/APERO.


Assuntos
Algoritmos , Escherichia coli/genética , Genoma Bacteriano , RNA Bacteriano/genética , RNA Mensageiro/genética , Pequeno RNA não Traduzido/genética , Salmonella enterica/genética , Conjuntos de Dados como Assunto , Enterobacteriaceae/genética , Enterobacteriaceae/metabolismo , Escherichia coli/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Internet , RNA Antissenso/classificação , RNA Antissenso/genética , RNA Antissenso/metabolismo , RNA Bacteriano/classificação , RNA Bacteriano/metabolismo , RNA Mensageiro/classificação , RNA Mensageiro/metabolismo , Pequeno RNA não Traduzido/classificação , Pequeno RNA não Traduzido/metabolismo , Salmonella enterica/metabolismo , Análise de Sequência de RNA , Software
11.
BMC Genomics ; 20(1): 477, 2019 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-31185909

RESUMO

BACKGROUND: Global RNA sequencing technologies have revealed widespread RNA polymerase II (Pol II) transcription outside of gene promoters. Small 5'-capped RNA sequencing (Start-seq) originally developed for the detection of promoter-proximal Pol II pausing has helped improve annotation of Transcription Start Sites (TSSs) of genes as well as identification of non-genic regulatory elements. However, apart from the most well studied genomes of human and mouse, mammalian transcription has not been profiled with sufficiently high precision. RESULTS: We prepared and sequenced Start-seq libraries from rat (Rattus norgevicus) primary neural progenitor cells. Over 48 million uniquely mappable reads from two independent biological replicates allowed us to define the TSSs of 7365 known genes in the rn6 genome, reannotating 2503 TSSs by more than 5 base pairs, characterize promoter-associated antisense transcription, and profile Pol II pausing. By combining TSS data with polyA-selected RNA sequencing, we also identified thousands of potential new genes producing stable RNA as well as non-genic transcripts representing possible regulatory elements. CONCLUSIONS: Our study has produced the first Start-seq dataset for the rat. Apart from profiling transcription initiation, our data reaffirm the prevalence of Pol II pausing across the rat genome and indicate conservation of pausing mechanisms across metazoan genomes. We suggest that pausing location, at least in mammals, is constrained by a distance from initiation of transcription, whether it occurs at or outside of a gene promoter. Abundant antisense transcription initiation around protein coding genes indicates that Pol II recruited to the vicinity of a promoter is distributed to available start sites of transcription at either DNA strand. Transcriptome profiling of neural progenitors presented here will facilitate further studies of other rat cell types as well as other organisms.


Assuntos
Genômica , Células-Tronco Neurais/metabolismo , RNA Polimerase II/metabolismo , Iniciação da Transcrição Genética , Animais , Feminino , Gravidez , RNA Antissenso/genética , Ratos , Ratos Sprague-Dawley , Análise de Sequência de RNA , Sítio de Iniciação de Transcrição
12.
Nat Immunol ; 20(7): 824-834, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31209403

RESUMO

Multiple genome-wide studies have identified associations between outcome of human immunodeficiency virus (HIV) infection and polymorphisms in and around the gene encoding the HIV co-receptor CCR5, but the functional basis for the strongest of these associations, rs1015164A/G, is unknown. We found that rs1015164 marks variation in an activating transcription factor 1 binding site that controls expression of the antisense long noncoding RNA (lncRNA) CCR5AS. Knockdown or enhancement of CCR5AS expression resulted in a corresponding change in CCR5 expression on CD4+ T cells. CCR5AS interfered with interactions between the RNA-binding protein Raly and the CCR5 3' untranslated region, protecting CCR5 messenger RNA from Raly-mediated degradation. Reduction in CCR5 expression through inhibition of CCR5AS diminished infection of CD4+ T cells with CCR5-tropic HIV in vitro. These data represent a rare determination of the functional importance of a genome-wide disease association where expression of a lncRNA affects HIV infection and disease progression.


Assuntos
Regulação da Expressão Gênica , Variação Genética , Infecções por HIV/genética , Infecções por HIV/virologia , HIV-1 , RNA Antissenso/genética , RNA Longo não Codificante/genética , Receptores CCR5/genética , Regiões 3' não Traduzidas , Alelos , Biomarcadores , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/virologia , Membrana Celular/metabolismo , Genes Reporter , Genótipo , Infecções por HIV/metabolismo , Humanos , Desequilíbrio de Ligação , Polimorfismo de Nucleotídeo Único , Grupos Populacionais/genética , Prognóstico , Estabilidade de RNA , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores CCR5/metabolismo , Carga Viral
13.
Chin Med J (Engl) ; 132(13): 1591-1598, 2019 Jul 05.
Artigo em Inglês | MEDLINE | ID: mdl-31205077

RESUMO

BACKGROUND: Natural anti-sense transcripts (NATs), which are transcribed from the complementary DNA strand of annotated genes, exert regulatory function of gene expression. Increasing studies recognized anti-sense transcription widespread throughout human cytomegalovirus (HCMV) genome, whereas the anti-sense transcription of RNA1.2 gene locus has never been investigated. In this study, the transcription of the RNA1.2 anti-sense strand was investigated in clinically isolated HCMV strain. METHODS: Strand-specific high-through RNA-sequencing (RNA-seq) was performed to find possible anti-sense transcripts (ASTs). For analyzing and visualization of RNA-seq data sets, Integrative Genomics Viewer software was applied. To confirm these possibilities, Northern blotting and rapid amplification of cDNA ends (RACE) were used. RESULTS: Transcription of the opposite strand of RNA1.2 gene locus was detected by RNA-sequencing using RNAs extracted from human embryonic lung fibroblasts infected with HCMV clinical isolate HAN. At least three HCMV NATs, named RNA1.2 AST 1, RNA1.2 AST2, and RNA1.2 AST3, were characterized by Northern blotting and RACE analyses. These RNA1.2 ASTs orientated from the complementary strand of RNA1.2 locus during the late phase of HCMV infection. The 5'- and 3'-termini of these transcripts were located within the opposite sequence of the predicted RNA1.2 gene. CONCLUSION: A cluster of novel NATs was transcribed from the opposite sequence of the HCMV RNA1.2 gene region.


Assuntos
Citomegalovirus/genética , RNA Antissenso/genética , Northern Blotting , Células Cultivadas , Genoma Viral/genética , Humanos , Software
14.
Surg Infect (Larchmt) ; 20(6): 472-479, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31038392

RESUMO

Background: Methicillin-resistant Staphylococcus aureus (MRSA) is an urgent medical problem in osteomyelitis. The YycFG two-component regulatory system (TCS) allows bacteria to adapt rapidly to physical, chemical, and biological stresses. The recombinant plasmid shuttle vector was used to overexpress an antisense RNA (asRNA) to inhibit target gene expression by sequence-specific double-stranded RNA complex degradation. In the current study, antisense yycG RNA (ASyycG)-overexpression MRSA clinical isolates were constructed. Methods: Bacterial growth was monitored, and biofilm biomass was determined by crystal violet microtiter assay. Quantitative reverse transcription polymerase chain reaction analysis was used to identify expression of yycF/G/H and icaA/D in MRSA and ASyycG strains. The expression of YycG protein was quantified by Western blot assays. The antibiotic resistance of ASyycG strains was compared with that of the MRSA strains. Results: The ASyycG strains showed a decrease in growth rate compared with the MRSA strains. Of note, overexpression of ASyycG led to a reduction in biofilm formation and adhesion force. ASyycG strains had decreased expressions of the yycF/G/H and icaA/D. Furthermore, Western blot data showed that expression of the YycG protein decreased by 40% in ASyycG strains compared with MRSA strains. In addition, the effect of yycG asRNA improved the susceptibility of ASyycG strains to cefoxitin. Conclusions: The ASyycG strains inhibited biofilm organization and increased antibiotic sensitivity, which may be attributed to altered intracellular polysaccharide construction.


Assuntos
Biofilmes/crescimento & desenvolvimento , Histidina Quinase/antagonistas & inibidores , Staphylococcus aureus Resistente à Meticilina/enzimologia , Staphylococcus aureus Resistente à Meticilina/crescimento & desenvolvimento , RNA Antissenso/metabolismo , Antibacterianos/farmacologia , Cefoxitina/farmacologia , Perfilação da Expressão Gênica , Histidina Quinase/genética , Humanos , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/genética , Testes de Sensibilidade Microbiana , RNA Antissenso/genética
15.
PLoS Genet ; 15(5): e1008144, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-31086376

RESUMO

Long noncoding RNAs (lncRNAs) participate in various biological processes such as apoptosis. The function of lncRNAs is closely correlated with their localization within the cell. While regulatory potential of many lncRNAs has been revealed at specific subcellular location, the biological significance of discrete distribution of an lncRNA in different cellular compartments remains largely unexplored. Here, we identified an lncRNA antisense to the pro-apoptotic gene PYCARD, named PYCARD-AS1, which exhibits a dual nuclear and cytoplasmic distribution and is required for the PYCARD silencing in breast cancer cells. The PYCARD-regulated apoptosis is controlled by PYCARD-AS1; moreover, PYCARD-AS1 regulates apoptosis in a PYCARD-dependent manner, indicating that PYCARD is a critical downstream target of PYCARD-AS1. Mechanistically, PYCARD-AS1 can localize to the PYCARD promoter, where it facilitates DNA methylation and H3K9me2 modification by recruiting the chromatin-suppressor proteins DNMT1 and G9a. Moreover, PYCARD-AS1 and PYCARD mRNA can interact with each other via their 5' overlapping region, leading to inhibition of ribosome assembly in the cytoplasm for PYCARD translation. This study reveals a mechanism whereby an lncRNA works at different cellular compartments to regulate the pro-apoptotic gene PYCARD at both the epigenetic and translational levels, contributing to the PYCARD-regulated apoptosis, and also sheds new light on the role of discretely distributed lncRNAs in diverse biological processes.


Assuntos
Apoptose/genética , Proteínas Adaptadoras de Sinalização CARD/genética , Regulação da Expressão Gênica/genética , Proteínas Reguladoras de Apoptose/genética , Proteínas Adaptadoras de Sinalização CARD/metabolismo , Linhagem Celular , Núcleo Celular/metabolismo , Proliferação de Células , Citoplasma/metabolismo , Metilação de DNA/genética , Epigênese Genética , Epigenômica , Células HEK293 , Humanos , Células MCF-7 , Regiões Promotoras Genéticas/genética , RNA Antissenso/genética , RNA Longo não Codificante/genética , Transdução de Sinais/genética
16.
Methods Mol Biol ; 1986: 17-33, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31115883

RESUMO

The DNA microarray is a powerful, flexible, nonbiased discovery technology. Microarrays can be used to assess processes from gene expression to long noncoding RNAs to specific pathologies, as well as many others. This chapter describes the protocol for DNA microarray analysis of differential gene expression using DNA sequences spotted on microscope slides.


Assuntos
Vidro/química , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Animais , Células Cultivadas , Humanos , RNA/genética , RNA/isolamento & purificação , RNA Antissenso/genética
17.
Plant Cell Physiol ; 60(8): 1646-1655, 2019 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-31093664

RESUMO

Upon nitrogen deficiency, some filamentous cyanobacteria differentiate specialized cells, called heterocysts, devoted to N2 fixation. Heterocysts appear regularly spaced along the filaments and exhibit structural and metabolic adaptations, such as loss of photosynthetic CO2 fixation or increased respiration, to provide a proper microaerobic environment for its specialized function. Heterocyst development is under transcriptional control of the global nitrogen regulator NtcA and the specific regulator HetR. Transcription of a large number of genes is induced or repressed upon nitrogen deficiency specifically in cells undergoing differentiation. In recent years, the HetR regulon has been described to include heterocyst-specific trans-acting small RNAs and antisense RNAs (asRNAs), suggesting that there is an additional layer of post-transcriptional regulation involved in heterocyst development. Here, we characterize in the cyanobacterium Nostoc (Anabaena) sp. PCC 7120 an asRNA, that we call as_glpX, transcribed within the glpX gene encoding the Calvin cycle bifunctional enzyme sedoheptulose-1,7-bisphosphatase/fructose-1,6-bisphosphatase (SBPase). Transcription of as_glpX is restricted to heterocysts and is induced very early during the process of differentiation. Expression of as_glpX RNA promotes the cleavage of the glpX mRNA by RNase III, resulting in a reduced amount of SBPase. Therefore, the early expression of this asRNA could contribute to the quick shut-down of CO2 fixation in those cells in the filament that are undergoing differentiation into heterocysts. In summary, as_glpX is the first naturally occurring asRNA shown to rapidly and dynamically regulate metabolic transformation in Nostoc heterocysts. The use of antisense transcripts to manipulate gene expression specifically in heterocysts could became a useful tool for metabolic engineering in cyanobacteria.


Assuntos
Nostoc/metabolismo , RNA Antissenso/metabolismo , Anabaena/genética , Anabaena/metabolismo , Dióxido de Carbono/metabolismo , Frutose-Bifosfatase/genética , Frutose-Bifosfatase/metabolismo , Regulação Bacteriana da Expressão Gênica/genética , Engenharia Metabólica , Nostoc/genética , Monoéster Fosfórico Hidrolases/genética , Monoéster Fosfórico Hidrolases/metabolismo , RNA Antissenso/genética , Ribonuclease III/genética , Ribonuclease III/metabolismo
18.
Genes (Basel) ; 10(4)2019 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-30959844

RESUMO

Antisense RNAs (asRNAs) are present in diverse organisms and play important roles in gene regulation. In this work, we mapped the primary antisense transcriptome in the halophilic archaeon Halobacterium salinarum NRC-1. By reanalyzing publicly available data, we mapped antisense transcription start sites (aTSSs) and inferred the probable 3' ends of these transcripts. We analyzed the resulting asRNAs according to the size, location, function of genes on the opposite strand, expression levels and conservation. We show that at least 21% of the genes contain asRNAs in H. salinarum. Most of these asRNAs are expressed at low levels. They are located antisense to genes related to distinctive characteristics of H. salinarum, such as bacteriorhodopsin, gas vesicles, transposases and other important biological processes such as translation. We provide evidence to support asRNAs in type II toxin⁻antitoxin systems in archaea. We also analyzed public Ribosome profiling (Ribo-seq) data and found that ~10% of the asRNAs are ribosome-associated non-coding RNAs (rancRNAs), with asRNAs from transposases overrepresented. Using a comparative transcriptomics approach, we found that ~19% of the asRNAs annotated in H. salinarum belong to genes with an ortholog in Haloferax volcanii, in which an aTSS could be identified with positional equivalence. This shows that most asRNAs are not conserved between these halophilic archaea.


Assuntos
Perfilação da Expressão Gênica , Halobacterium salinarum/genética , RNA Antissenso/genética , Transcriptoma/genética , Regulação da Expressão Gênica em Archaea/genética , Genoma Arqueal/genética , RNA não Traduzido/genética , Ribossomos/genética , Sítio de Iniciação de Transcrição
19.
Biochimie ; 164: 3-16, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-30995539

RESUMO

Prokaryotes encounter constant and often brutal modifications to their environment. In order to survive, they need to maintain fitness, which includes adapting their protein expression patterns. Many factors control gene expression but this review focuses on just one, namely antisense RNAs (asRNAs), a class of non-coding RNAs (ncRNAs) characterized by their location in cis and their perfect complementarity with their targets. asRNAs were considered for a long time to be trivial and only to be found on mobile genetic elements. However, recent advances in methodology have revealed that their abundance and potential activities have been underestimated. This review aims to illustrate the role of asRNA in various physiologically crucial functions in both archaea and bacteria, which can be regrouped in three categories: cell maintenance, horizontal gene transfer and virulence. A literature survey of asRNAs demonstrates the difficulties to characterize and assign a role to asRNAs. With the aim of facilitating this task, we describe recent technological advances that could be of interest to identify new asRNAs and to discover their function.


Assuntos
Archaea , Bactérias , Fenômenos Fisiológicos Bacterianos/genética , Transferência Genética Horizontal/genética , RNA Antissenso , Virulência/genética , Archaea/genética , Archaea/patogenicidade , Archaea/fisiologia , Bactérias/genética , Bactérias/patogenicidade , Regulação da Expressão Gênica em Archaea , Regulação Bacteriana da Expressão Gênica , RNA Antissenso/genética , RNA Antissenso/fisiologia , RNA Arqueal/genética , RNA Arqueal/fisiologia , RNA Bacteriano/genética , RNA Bacteriano/fisiologia
20.
Cell Prolif ; 52(3): e12564, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30938030

RESUMO

OBJECTIVES: Thyroid carcinoma (TC) represents a malignant neoplasm affecting the thyroid. Current treatment strategies include the removal of part of the thyroid; however, this approach is associated with a significant risk of developing hypothyroidism. In order to adequately understand the expression profiles of TNRC6C-AS1 and STK4 and their potential functions in TC, an investigation into their involvement with Hippo signalling pathway and the mechanism by which they influence TC apoptosis and autophagy were conducted. METHODS: A microarray analysis was performed to screen differentially expressed lncRNAs associated with TC. TC cells were employed to evaluate the role of TNRC6C-AS1 by over-expression or silencing means. The interaction of TNRC6C-AS1 with methylation of STK4 promoter was evaluated to elucidate its ability to elicit autophagy, proliferation and apoptosis. RESULTS: TNRC6C-AS1 was up-regulated while STK4 was down-regulated, where methylation level was elevated. STK4 was verified as a target gene of TNRC6C-AS1, which was enriched by methyltransferase. Methyltransferase's binding to STK4 increased expression of its promoter. Over-expressed TNRC6C-AS1 inhibited STK4 by promoting STK4 methylation and reducing the total protein levels of MST1 and LATS1/2. The phosphorylation of YAP1 phosphorylation was decreased, which resulted in the promotion of SW579 cell proliferation and tumorigenicity. CONCLUSION: Based on our observations, we subsequently confirmed the anti-proliferative, pro-apoptotic and pro-autophagy capabilities of TNRC6C-AS1 through STK4 methylation via the Hippo signalling pathway in TC.


Assuntos
Proteínas Serina-Treonina Quinases/genética , RNA Longo não Codificante/antagonistas & inibidores , RNA Longo não Codificante/genética , Proteínas de Ligação a RNA/antagonistas & inibidores , Proteínas de Ligação a RNA/genética , Neoplasias da Glândula Tireoide/genética , Neoplasias da Glândula Tireoide/metabolismo , Animais , Apoptose/genética , Autofagia/genética , Sequência de Bases , Linhagem Celular Tumoral , Proliferação de Células/genética , Desmetilação do DNA , Regulação para Baixo , Xenoenxertos , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Modelos Biológicos , RNA Antissenso/genética , Transdução de Sinais , Neoplasias da Glândula Tireoide/patologia , Regulação para Cima
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