Dataset Information

Identification of host RNAs that interact with EBV noncoding RNA EBER2.

ABSTRACT: Epstein-Barr virus (EBV) expresses an abundant nuclear noncoding RNA called EBER2, which interacts with and acts as a guide RNA for the host transcription factor PAX5. This ribonucleoprotein complex localizes to the terminal repeat (TR) regions of the EBV genome via RNA-RNA interactions between EBER2 and nascent transcripts originating from these target sites. Given the fact that EBER2 base pairs with a viral RNA, we developed a protocol to identify EBER2-interacting RNAs in a transcriptome-wide manner. Our approach entails psoralen-mediated crosslinking, selection with antisense oligonucleotides targeting EBER2, and RNase V1 digestion coupled to next-generation sequencing. The use of RNase V1 circumvents the need of extensive computational analysis post data acquisition to search for predicted RNA hybrids, as the RNase V1 cleavage site marks the region of RNA duplex formation. As proof of principle, we show that our approach correctly identifies the known EBER2 interaction with TR RNAs. Moreover, we identify the host functional noncoding RNAs MRP, H1, and 7SL RNAs as well as three putative enhancer RNAs as candidate EBER2-interacting RNAs. As all of these gene loci exhibit PAX5 occupancy, we propose that EBER2 is recruited to these sites through its binding partner PAX5 and forms RNA-RNA interactions with nascent transcripts on chromatin. Thus, our novel approach facilitates the identification of targeted RNA-RNA-interactions and minimizes the need of downstream computational analyses to predict RNA duplexes.

SUBMITTER: Nanni AV

PROVIDER: S-EPMC6284586 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Identification of host RNAs that interact with EBV noncoding RNA EBER2.

Nanni Adalena V AV Lee Nara N

RNA biology 20180918 9

Epstein-Barr virus (EBV) expresses an abundant nuclear noncoding RNA called EBER2, which interacts with and acts as a guide RNA for the host transcription factor PAX5. This ribonucleoprotein complex localizes to the terminal repeat (TR) regions of the EBV genome via RNA-RNA interactions between EBER2 and nascent transcripts originating from these target sites. Given the fact that EBER2 base pairs with a viral RNA, we developed a protocol to identify EBER2-interacting RNAs in a transcriptome-wide ...[more]

PMID: 30176159

Similar Datasets

Project description:In Epstein-Barr virus (EBV) latent infection, the EBV-encoded RNAs EBER1 and EBER2 accumulate in the host cell nucleus to ~10(6) copies. While the expression of EBERs in cell lines is associated with transformation, a mechanistic explanation of their roles in EBV latency remains elusive. To identify EBER-specific gene expression features, we compared the proteome and mRNA transcriptome from BJAB cells (an EBV-negative B lymphoma cell line) stably transfected with an empty plasmid or with one carrying both EBER genes. We identified ~1800 proteins with at least 2 SILAC pair measurements, of which only 8 and 12 were up- and downregulated ≥ 2-fold, respectively. One upregulated protein was PIK3AP1, a B-cell specific protein adapter known to activate the PI3K-AKT signaling pathway, which regulates alternative splicing and translation in addition to its pro-survival effects. In the mRNA-seq data, the mRNA levels for some of the proteins changing in the SILAC data did not change. We instead observed isoform switch events. We validated the most relevant findings with biochemical assays. These corroborated the upregulation of PIK3AP1 and AKT activation in BJAB cells expressing high levels of both EBERs and EBNA1 (a surrogate of Burkitt's lymphoma EBV latency I) relative to those expressing only EBNA1. The mRNA-seq data in these cells showed multiple upregulated oncogenes whose mRNAs are enriched for 3´-UTR AU-rich elements (AREs), such as ccl3, ccr7, il10, vegfa and zeb1. The CCL3, CCR7, IL10 and VEGFA proteins promote cell proliferation and are associated with EBV-mediated lymphomas. In EBV latency, ZEB1 represses the transcription of ZEBRA, an EBV lytic phase activation factor. We previously found that EBER1 interacts with AUF1 in vivo and proposed stabilization of ARE-containing mRNAs. Thus, the ~10(6) copies of EBER1 may promote not only cell proliferation due to an increase in the levels of ARE-containing genes like ccl3, ccr7, il10, and vegfa, but also the maintenance of latency, through higher levels of zeb1.

Project description:UNLABELLED:Neisseria meningitidis is a leading cause of bacterial meningitis and septicemia, affecting infants and adults worldwide. N. meningitidis is also a common inhabitant of the human nasopharynx and, as such, is highly adapted to its niche. During bacteremia, N. meningitidis gains access to the blood compartment, where it adheres to endothelial cells of blood vessels and causes dramatic vascular damage. Colonization of the nasopharyngeal niche and communication with the different human cell types is a major issue of the N. meningitidis life cycle that is poorly understood. Here, highly saturated random transposon insertion libraries of N. meningitidis were engineered, and the fitness of mutations during routine growth and that of colonization of endothelial and epithelial cells in a flow device were assessed in a transposon insertion site sequencing (Tn-seq) analysis. This allowed the identification of genes essential for bacterial growth and genes specifically required for host cell colonization. In addition, after having identified the small noncoding RNAs (sRNAs) located in intergenic regions, the phenotypes associated with mutations in those sRNAs were defined. A total of 383 genes and 8 intergenic regions containing sRNA candidates were identified to be essential for growth, while 288 genes and 33 intergenic regions containing sRNA candidates were found to be specifically required for host cell colonization. IMPORTANCE:Meningococcal meningitis is a common cause of meningitis in infants and adults. Neisseria meningitidis (meningococcus) is also a commensal bacterium of the nasopharynx and is carried by 3 to 30% of healthy humans. Under some unknown circumstances, N. meningitidis is able to invade the bloodstream and cause either meningitis or a fatal septicemia known as purpura fulminans. The onset of symptoms is sudden, and death can follow within hours. Although many meningococcal virulence factors have been identified, the mechanisms that allow the bacterium to switch from the commensal to pathogen state remain unknown. Therefore, we used a Tn-seq strategy coupled to high-throughput DNA sequencing technologies to find genes for proteins used by N. meningitidis to specifically colonize epithelial cells and primary brain endothelial cells. We identified 383 genes and 8 intergenic regions containing sRNAs essential for growth and 288 genes and 33 intergenic regions containing sRNAs required specifically for host cell colonization.

Project description:Liver fibrosis occurs as a result of chronic liver lesions, which may subsequently develop into liver cirrhosis and hepatocellular carcinoma. The involvement of long noncoding RNAs (lncRNAs) in liver fibrosis is being increasingly recognized. However, the exact mechanisms and functions of the majority of lncRNAs are poorly characterized. In the present study, the hepatotoxic substance carbon tetrachloride (CCl4) was employed to induce liver fibrosis in an animal model and agenome‑wide identification of lncRNAs in fibrotic liver tissues compared with CCl4 untreated liver tissues was performed using RNA sequencing. Sprague‑Dawley rats were treated with CCl4 for 8 weeks. Histopathogical alterations were observed in liver tissues, and serum levels of alanine aminotransferase, aspartate aminotransferase, transforming growth factor‑β1 and tumor necrosis factor‑α were significantly higher, in the CCl4‑treated group compared with the CCl4 untreated group. RNA sequencing of liver tissues demonstrated that 231 lncRNAs and 1,036 mRNAs were differentially expressed between the two groups. Furthermore, bioinformatics analysis demonstrated that the differentially expressed mRNAs were predominantly enriched in 'ECM‑receptor interaction', 'PI3K‑Akt signaling pathway' and 'focal adhesion' pathways, all of which are essential for liver fibrosis development. Validation of 12 significantly aberrant lncRNAs by reverse transcription‑quantitative polymerase chain reaction indicated that the expression patterns of 11 lncRNAs were consistent with the sequencing data. Furthermore, overexpression of lncRNA NR_002155.1, which was markedly downregulated in CCl4‑treated liver tissues, was demonstrated to inhibit HSC‑T6 cell proliferation in vitro. In conclusion, the present study determined the expression patterns of mRNAs and lncRNAs in fibrotic liver tissue induced by CCl4. The identified differentially expressed lncRNAs may serve as novel diagnostic biomarkers and therapeutic targets for liver fibrosis.

Project description:BackgroundMajor depressive disorder (MDD) is a common and serious mental illness. Many novel genes in MDD have been characterized by high-throughput methods such as microarrays or sequencing. Recently, noncoding RNAs (ncRNAs) were suggested to be involved in the complicated environmental-genetic regulatory network of MDD occurrence; however, the interplay among RNA species, including protein-coding RNAs and ncRNAs, in MDD remains unclear.AimTo investigate the RNA expression datasets downloaded from a public database and construct a network based on differentially expressed long noncoding RNA (lncRNAs), microRNAs (miRNAs), and mRNAs between MDD and controls.MethodsGene expression data were searched in NCBI Gene Expression Omnibus using the search term "major depressive disorder." Six array datasets from humans were related to the search term: GSE19738, GSE32280, GSE38206, GSE52790, GSE76826, and GSE81152. These datasets were processed for initial assessment and subjected to quality control and differential expression analysis. Differentially expressed lncRNAs, miRNAs, and mRNAs were determined, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses were performed, and protein-protein interaction network was generated. The results were analyzed for their association with MDD.ResultsAfter analysis, 3 miRNAs, 12 lncRNAs, and 33 mRNAs were identified in the competing endogenous RNA network. Two of these miRNAs were earlier shown to be involved in psychiatric disorders, and differentially expressed mRNAs were found to be highly enriched in pathways related to neurogenesis and neuroplasticity as per Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses. The expression of hub gene fatty acid 2-hydroxylase was enriched, and the encoded protein was found to be involved in myelin formation, indicating that neurological development and signal transduction are involved in MDD pathogenesis.ConclusionThe present study presents candidate ncRNAs involved in the neurogenesis and neuroplasticity pathways related to MDD.

Dataset Information

Identification of host RNAs that interact with EBV noncoding RNA EBER2.

Publications

Identification of host RNAs that interact with EBV noncoding RNA EBER2.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets