Identification of novel, highly expressed retroviral microRNAs in cells infected by bovine foamy virus
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ABSTRACT:
ORGANISM(S): Bos taurus
PROVIDER: GSE67620 | GEO | 2015/04/07
SECONDARY ACCESSION(S): PRJNA280497
REPOSITORIES: GEO
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Project description:While numerous viral microRNAs (miRNAs) expressed by DNA viruses, especially herpesvirus family members, have been reported, there have been very few reports of miRNAs derived from RNA viruses. Here we describe three miRNAs expressed by bovine foamy virus (BFV), a member of the spumavirus subfamily of retroviruses, in both BFV-infected cultured cells and BFV-infected cattle. All three viral miRNAs are initially expressed in the form of an ∼ 122-nucleotide (nt) pri-miRNA, encoded within the BFV long terminal repeat U3 region, that is subsequently cleaved to generate two pre-miRNAs that are then processed to yield three distinct, biologically active miRNAs. The BFV pri-miRNA is transcribed by RNA polymerase III, and the three resultant mature miRNAs were found to contribute a remarkable ∼ 70% of all miRNAs expressed in BFV-infected cells. These data document the second example of a retrovirus that is able to express viral miRNAs by using embedded proviral RNA polymerase III promoters. Small RNA profiling of Bovine Foamy Virus (BFV)-infected cells through total small RNA deep sequencing
2015-04-07 | E-GEOD-67620 | biostudies-arrayexpress
Project description:The cellular abundance of mature microRNAs (miRNAs) is dictated by the efficiency of nuclear processing of primary miRNA transcripts (pri-miRNAs) into pre-miRNA intermediates. The Microprocessor complex of Drosha and DGCR8 carries this out, but it has been unclear what controls Microprocessor's differential processing of various pri-miRNAs. Here, we show that Drosophila DGCR8 (Pasha) directly associates with the C terminal domain of the RNA polymerase II elongation complex when it is phosphorylated by the Cdk9 kinase (pTEFb). When association is blocked by loss of Cdk9 activity, a global change in pri-miRNA processing is detected. Processing of pri-miRNAs with a UGU sequence motif in their apical junction domain increases, while processing of pri-miRNAs lacking this motif decreases. Therefore, phosphorylation of RNA polymerase II recruits Microprocessor for co-transcriptional processing of non-UGU pri-miRNAs that would otherwise be poorly processed. In contrast, UGU-positive pri-miRNAs are robustly processed by Microprocessor independent of RNA polymerase association.
2017-09-15 | GSE103234 | GEO
Project description:Micro (mi)RNAs are small non-coding RNAs with key regulatory functions. Recent advances in the field allowed researchers to identify their targets. However, much less is known regarding the regulation of miRNA themselves. The accumulation of these tiny regulators can be modulated at various levels during their biogenesis from the transcription of the primary transcript (pri-miRNA) to the stability of the mature miRNA. Here, we studied the importance of the pri-miRNA secondary structure for the regulation of mature miRNAs accumulation. To this end, we used the KaposiM-bM-^@M-^Ys sarcoma herpesvirus, which encodes a cluster of twelve pre-miRNAs. Using small RNA profiling and quantitative northern blot analysis, we measured the absolute amount of each mature miRNAs in different cellular context. We found that the difference in expression between the least and most expressed viral miRNA could be as high as 60-fold. Using high-throughput selective 2M-bM-^@M-^Y-hydroxyl acylation analyzed by primer extension (hSHAPE), we then determined the secondary structure of the long primary transcript. We found that highly expressed miRNAs derived from optimally structured regions within the pri-miRNA. Finally, we confirmed the importance of the local structure by swapping stem-loops for highly and lowly expressed miRNAs, which resulted in a perturbed accumulation of the mature miRNA. Examination of sRNA profiles in 3 independent B cell lines expressing KSHV miRNAs or infected with KSHV, without replicate
2014-06-02 | E-GEOD-53296 | biostudies-arrayexpress
Project description:Maturation of canonical microRNA (miRNA) is initiated by DROSHA that cleaves the primary transcript (pri-miRNA). Over 1,800 miRNA loci are annotated in humans, but it remains largely unknown if and at which sites the pri-miRNAs are cleaved by DROSHA. Here we performed in vitro processing on a full set of human pri-miRNAs (miRBase v21) followed by sequencing. This comprehensive profiling enabled us to classify miRNAs based on DROSHA-dependence and map their cleavage sites with respective processing efficiency measures. Only 758 pri-miRNAs are confidently processed by DROSHA, while the majority may be non-canonical or false entries. Analyses of the DROSHA-dependent pri-miRNAs show key cis-elements for processing. We observe widespread alternative processing as well as unproductive cleavage events such as “nick” or “inverse” processing. SRSF3 is a broad-acting auxiliary factor modulating alternative processing and suppressing unproductive processing. The profiling data and methods developed in this study will allow systematic analyses of miRNA regulation.
2021-07-28 | PXD025767 | Pride
Project description:MicroRNAs (miRNAs) are small regulatory RNAs processed from stem-loop regions of primary transcripts (pri-miRNAs), with the choice of stem-loops for initial processing largely determining what becomes a miRNA. To identify sequence and structural features influencing this choice, we determined cleavage efficiencies of >50,000 variants of three human pri-miRNAs, focusing on the regions intractable to previous high-throughput analyses. Our analyses revealed a mismatched motif in the basal stem region, a preference for maintaining or improving base-pairing throughout the remainder of the stem, and a narrow stem-length preference of 35±1 base pairs. Incorporating these features with previously identified features, including three primary-sequence motifs, yielded a unifying model defining mammalian pri-miRNAs, in which motifs help orient processing and increase efficiency, with the presence of more motifs compensating for structural defects. This model enables generation of artificial pri-miRNAs, designed de novo, without reference to any natural sequence, yet processed more efficiently than natural pri-miRNAs. Three major experiments are included in the submitted data. 1) Pools of synthetic DNA containing barcodes and pri-miRNA variants were sequenced to infer the barcode-pri-miRNA linkages, and the pri-miRNA variants were assayed for their cleavage efficiency ("selection"), which can be quantified by comparing their representations in the input and after cleavage. 2) Artificial miRNAs produced in HEK293T cells were sequenced. 3) Transcriptomes of artificial miRNA-expressed cells were sequenced to quantify mRNA changes.
2015-09-24 | E-GEOD-67937 | biostudies-arrayexpress
Project description:Background. The pathogenesis of influenza A virus subtype H5N1 (hearafter, "H5N1") infection in humans is not completely understood, although hypercytokinemia is thought to play a role. We previously reported that most H5N1 viruses induce high cytokine responses in human macrophages, whereas some H5N1 viruses induce only a low level of cytokine production similar to that induced by seasonal viruses. Methods. To identify the viral molecular determinants for cytokine induction of H5N1 viruses in human macrophages, we generated a series of reassortant viruses between the high cytokine inducer A/Vietnam/UT3028II/03 clone 2 (VN3028IIcl2) and the low inducer A/Indonesia/UT3006/05 (IDN3006), and evaluated cytokine expression in human macrophages. Results. Viruses possessing the acidic polymerase (PA) gene of VN3028IIcl2 exhibited high levels of hypercytokinemia-related cytokine expression in human macrophages, compared with IDN3006, but showed no substantial differences in viral growth in these cells. Further, the PA gene of VN3028IIcl2 conferred enhanced virulence in mice. Conclusions. These results demonstrate that the PA gene of VN3028IIcl2 affects cytokine production in human macrophages and virulence in mice. These findings provide new insights into the cytokine-mediated pathogenesis of H5N1 infection in humans. Human monocyte-derived macrophages were mock-infected, or infected with IDN3006, VN3028IIcl2, or IDN3006/cl2PA at a multiplicity of infection (MOI) of 2. At 6 hours post-infection (hpi), the cells were harvested and subjected to microarray analysis (three technical replicates per each group).
2012-09-30 | E-GEOD-40711 | biostudies-arrayexpress
Project description:Porcine cytomegalovirus (PCMV; genus Cytomegalovirus, subfamily Betaherpesvirinae, family Herpesviridae) is an immunosuppressive virus that mainly inhibits the immune function of T lymphocytes and macrophages, which has caused great distress to the farming industry. In this study, we obtained the miRNA expression profiles of PCMV-infected and control porcine macrophages, PCMV-infected and control porcine tissues via high-throughput sequencing. The comprehensive analysis of miRNA profiles showed that 306 miRNA database annotated and 295 novel pig-encoded miRNAs were detected. Gene Ontology (GO) analysis of the target genes of miRNAs in PCMV infected porcine macrophages showed that the differentially expressed miRNAs are mainly involved in immune and metabolic process. This is the first report of the miRNA transcriptome in PCMV infected porcine macrophages and PCMV infected tissues and the analysis of the miRNA regulatory mechanism during PCMV infection. Further research into the regulatory mechanisms of miRNAs during immunosuppressive viral infections will contribute to the treatment and prevention of immunosuppressive viruses. miRNA expression profiling of PCMV-infected and control porcine macrophages; PCMV-infected and control porcine tissues via high-throughput sequencing.
2015-12-19 | E-GEOD-76156 | biostudies-arrayexpress
Project description:Porcine cytomegalovirus (PCMV; genus Cytomegalovirus, subfamily Betaherpesvirinae, family Herpesviridae) is an immunosuppressive virus that mainly inhibits the immune function of T lymphocytes and macrophages, which has caused great distress to the farming industry. In this study, we obtained the miRNA expression profiles of PCMV-infected and control porcine macrophages, PCMV-infected and control porcine tissues via high-throughput sequencing. The comprehensive analysis of miRNA profiles showed that 306 miRNA database annotated and 295 novel pig-encoded miRNAs were detected. Gene Ontology (GO) analysis of the target genes of miRNAs in PCMV infected porcine macrophages showed that the differentially expressed miRNAs are mainly involved in immune and metabolic process. This is the first report of the miRNA transcriptome in PCMV infected porcine macrophages and PCMV infected tissues and the analysis of the miRNA regulatory mechanism during PCMV infection. Further research into the regulatory mechanisms of miRNAs during immunosuppressive viral infections will contribute to the treatment and prevention of immunosuppressive viruses.
2015-12-19 | GSE76156 | GEO
Project description:SE-Associated Protein 1 (SEAP1), is a conserved protein in eukaryotes. Its orthologs, SART3 and PRP24 from metazoans and yeast play essential roles in splicing. Here, we report that SE-Associated Protein 1 (SEAP1), is required for miRNA biogenesis in Aradopsis. Lack of SEAP1 causes embryo lethality and knockdown of SEAP1 by an artificial miRNA (amiRSEAP1) causes pleiotropic growth defects and reduction in miRNA accumulation, accompanied with increased levels of target transcripts. SEAP1 associates with the RNase III enzyme Dicer-like 1(DCL1) complex, which processes primary miRNA transcripts (pri-miRNAs) into miRNAs, and seems to promote the interaction of the DCL1 complexes with pri-miRNAs. SEAP1 is also required for pri-miRNA accumulation. However, it does not affect the transcription of pri-miRNAs, suggesting it may indirectly or directly stabilize pri-miRNAs. SEAP1 also affects the spicing of pri-mRNAs. These results suggest that SEPA1 promotes miRNA biogenesis by positively modulating pri-miRNA splicing, processing and stability. We also find that intron retention in amiRSEAP1 is altered at global levels, linking SEAP1 to spicing. Collectively, our results uncover the function of SEAP1 in plants.
2021-04-12 | GSE171865 | GEO
Project description:MiRNAs are key regulators of gene expression. They are processed from primary miRNA transcripts (pri-miRNAs), most of which are transcribed by DNA-dependent polymerase II (Pol II). MiRNA levels need to be precisely controlled to maintain various biological processes. In this manuscript, we report a discovery that STV1, a conserved ribosomal protein, acts in miRNA biogenesis in Arabidopsis. A portion of STV1 localizes at the nucleus and binds pri-miRNAs. Using pri-miR172b as a reporter, we show that STV1 binds the stem-loop with a short 5’ flanking sequencing within pri-miRNAs. Lack of STV1 in stv1-1 reduced the association of pri-miRNAs with their processing complex. These data suggest that STV1 promotes miRNA biogenesis through facilitating the recruitment of pri-miRNAs to their processing complex. Furthermore, we show that STV1 indirectly influence the occupancy of Pol II at the promoters of miRNA coding genes (MIR) and MIR promoter activities. Based on these results, we propose that STV1 refines the accumulation of miRNAs through its combined effects on pri-miRNA processing and transcription. This study uncovers an extra-ribsomal function of STV1.
2017-03-06 | GSE91013 | GEO