Project description:This SuperSeries is composed of the following subset Series: GSE31745: Primary effusion lymphoma cell lines BC-1 and BC-3 GSE31746: BJAB Cell Lines Transduced with lentiviral vector pNL-SIN-CMV-AcGFP expressing KSHV miRNAs miR-K1, miR-K12-11, or miR-K4-3p GSE32109: microRNA Targetome Analysis of Latently KSHV-infected Primary Effusion Lymphoma Cell lines Using PAR-CLIP [Illumina] Refer to individual Series
Project description:BJAB cells were infected with pNL-SIN-CMV-AcGFP (see accession number GSE8867 for data) or pNL-SIN-CMV-AcGFP expressing KSHV miR-K1, miR-K12-11 or miR-K4-3p and sorted 48 hours after infection. 12 or 16 days after transduction, cytoplasmic RNA was harvested and gene expression analysis of independent BJAB cell pools was performed using Human Operon v3.0.2 arrays. Each sample was run against Universal Human Reference RNA, Stratagene. The samples listed here were processed in parallel to those with accession number GSE8867, which includes all matched control cell lines. 6 independent B cell pools each expressing KSHV miR-K1, miRK12-11, or miR-K4-3p are included. Matched controls include unmodified BJAB (3 arrays) and cells transducted with parental vector pNL-SIN-CMV-AcGFP (6 replicates).
Project description:BJAB cells were infected with pNL-SIN-CMV-AcGFP (see accession number GSE8867 for data) or pNL-SIN-CMV-AcGFP expressing KSHV miR-K1, miR-K12-11 or miR-K4-3p and sorted 48 hours after infection. 12 or 16 days after transduction, cytoplasmic RNA was harvested and gene expression analysis of independent BJAB cell pools was performed using Human Operon v3.0.2 arrays. Each sample was run against Universal Human Reference RNA, Stratagene. The samples listed here were processed in parallel to those with accession number GSE8867, which includes all matched control cell lines.
Project description:Transcriptional profiling of BJAB cells expressing miR-K12-9 and BCBL cells treated with miR-K12-9 inhibitor. To identify host RNA targets of KSHV miRNAs, we took advantage of the observation that RNAs targeted by miRNAs often display small reductions in their steady-state levels, perhaps as a result of their impaired translation. Accordingly, we examined cellular transcript accumulation by array-based expression profiling under four sets of conditions in which KSHV miRNAs were expressed or inhibited. Cells transfected with negative control miRNA compared to miR-K12-9 or negative miRNA inhibitor compared to miR-K12-9 inhibitor.
Project description:Transcriptional profiling of BJAB cells expressing miR-K12-9 and BCBL cells treated with miR-K12-9 inhibitor. To identify host RNA targets of KSHV miRNAs, we took advantage of the observation that RNAs targeted by miRNAs often display small reductions in their steady-state levels, perhaps as a result of their impaired translation. Accordingly, we examined cellular transcript accumulation by array-based expression profiling under four sets of conditions in which KSHV miRNAs were expressed or inhibited.
Project description:Transcriptional profiling of BJAB cells expressing miR-K12-6-5p. To identify host RNA targets of KSHV miRNAs, we took advantage of the observation that RNAs targeted by miRNAs often display small reductions in their steady-state levels, perhaps as a result of their impaired translation. Accordingly, we examined cellular transcript accumulation by array-based expression profiling under four sets of conditions in which KSHV miRNAs were expressed. Cells transfected with negative control miRNA compared to miR-K12-6-5p.
Project description:All metazoan eukaryotes express microRNAs (miRNAs), ~22 nt long regulatory RNAs that can repress the expression of mRNAs bearing complementary sequences. Several DNA viruses also express miRNAs in infected cells, suggesting a role in viral replication and pathogenesis. While specific viral miRNAs have been shown to autoregulate viral mRNAs or downregulate cellular mRNAs via novel target sites, the function of the majority of viral miRNAs remains unknown. Here, we report that the miR-K12-11 miRNA encoded by Kaposi’s Sarcoma Associated Herpesvirus (KSHV) shows significant homology to cellular miR-155, including the entire miRNA “seed” region. Using a range of assays, we demonstrate that expression of physiological levels of miRK12- 11 or miR-155 results in the downregulation of an extensive set of common mRNA targets, including genes with known roles in cell growth regulation. Our findings indicate that viral miR-K12-11 functions as an ortholog of cellular miR-155 and has likely evolved to exploit a pre-existing gene regulatory pathway in B-cells. Moreover, the known etiological role of miR-155 in B-cell transformation suggests the possibility that miR-K12-11 may contribute to the induction of KSHV positive B-cell tumors in infected patients. BJAB cells were infected and sorted 48 hours after infection. 12 to 16 days after transduction, gene expression analysis of 10 independent BJAB cell pools, expressing AcGFP only or AcGPF and miR-K12-11, was performed using Human Operon v3.0.2 arrays. Each ample was run against Universal Human Reference RNA, Stratagene. # of Arrays: BJAB = 3 AcGFP only = 10 AcGFP miR-K12-11 = 11
Project description:Transcriptional profiling of BJAB cells expressing miR-K12-6-5p. To identify host RNA targets of KSHV miRNAs, we took advantage of the observation that RNAs targeted by miRNAs often display small reductions in their steady-state levels, perhaps as a result of their impaired translation. Accordingly, we examined cellular transcript accumulation by array-based expression profiling under four sets of conditions in which KSHV miRNAs were expressed.
Project description:Herpesviruses are known to encode micro (mi)RNAs and to use them to regulate the expression of both viral and cellular genes. The genome of Kaposi’s sarcoma herpesvirus (KSHV) encodes a cluster of twelve miRNAs, which are abundantly expressed during both latency and lytic infection. Relatively few cellular targets of KSHV miRNAs are known. Here, we used a microarray expression profiling approach to analyze the transcriptome of both B lymphocytes and endothelial cells stably expressing KSHV miRNAs and monitor the changes induced by the presence of these miRNAs. We generated a list of potential cellular targets by looking for miRNA seed-match-containing transcripts that were significantly down regulated upon KSHV miRNAs expression. Interestingly, the overlap of putative targets identified in B lymphocytes and endothelial cells was minimal, suggesting a tissue-specific target-regulation by viral miRNAs. Among the putative targets, we identified caspase 3, a critical factor for the control of apoptosis, which we validated using luciferase reporter assays and western blotting. In functional assays we obtained further evidence that KSHV miRNAs indeed protect cells from apoptosis. Single-channel hybridization to Affymetrix oligonucleotides microarrays. DG75 and EA.hy926 cells were transducted in triplicates with the K10/12 and K12/12 lentiviral constructs expressing the KSHV miRNAs, or with a GFP-expressing construct as control. The K10/12 construct expressed only the ten intronic miRNAs miR-K12-1 to 9 and miR-K12-11 while the K12/12 construct also expressed miR-K12-10 and miR-K12-12 in addition.