Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Murine bone marrow-derived mast cells (BMMCs) transduced with pAPM lentiviruses overexpressing miR-221 vs control overexpressing miR-221m.

ABSTRACT: Bone marrow-derived mast cells were derived from three different female C57Bl/6 mice of similar age. Cells were transduced with pAPM lentiviruses either overexpressing microRNA-221 or a non functional microRNA-221 mutant, termed miR-221m, where four point-mutations were introduced in the seed sequence of miR-221. The samples overexpressing miR-221m were used as references for the corresponding miR-221 overexpressing BMMCs from the same mouse (mouse matched reference). The experiment was performed in order to gain more insights about the molecular mechanisms underlying miR-221 effects in mast cells. Two-condition experiment, BMMC overexpressing miR-221 vs BMMC overexpressing miR-221m. Biological replicates: 3 replicates overexpressing miR-221, 3 control replicates overexpressing miR-221m.

ORGANISM(S): Mus musculus

SUBMITTER: Silvia Monticelli

PROVIDER: E-GEOD-24462 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

MiR-221 influences effector functions and actin cytoskeleton in mast cells.

Mayoral Ramon J RJ Deho Lorenzo L Rusca Nicole N Bartonicek Nenad N Saini Harpreet Kaur HK Enright Anton J AJ Monticelli Silvia S

PloS one 20111012 10

Mast cells have essential effector and immunoregulatory functions in IgE-associated allergic disorders and certain innate and adaptive immune responses, but the role of miRNAs in regulating mast cell functions is almost completely unexplored. To examine the role of the activation-induced miRNA miR-221 in mouse mast cells, we developed robust lentiviral systems for miRNA overexpression and depletion. While miR-221 favored mast cell adhesion and migration towards SCF or antigen in trans-well migra ...[more]

PMID: 22022537

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Project description:Modification of microRNA sequences by the 3' addition of nucleotides to generate so-called M-bM-^@M-^\isomiRsM-bM-^@M-^] adds to the complexity of miRNA function, with recent reports showing that 3' modifications can influence miRNA stability and efficiency of target repression. Here we show that the 3' modification of miRNAs is a physiological and common post-transcriptional event that shows selectivity for specific miRNAs and is observed across species ranging from C. elegans to human. The modifications result predominantly from adenylation and uridylation, and are seen across tissue types, disease states, and developmental stages. To quantitatively profile 3' nucleotide additions, we developed and validated a novel assay based on NanoString Technologies' nCounter platform. For certain miRNAs, the frequency of modification was altered by processes such as cell differentiation, indicating that 3' modification is a biologically regulated process. To investigate the mechanism of 3' nucleotide additions, we used RNA interference to screen a panel of eight candidate miRNA nucleotidyl transferases for 3' miRNA modification activity in human cells. Multiple enzymes, including PAPD1, PAPD4, PAPD5, ZCCHC6, ZCCHC11, and TUT1, were found to govern 3' nucleotide addition to miRNAs in a miRNA-specific manner. Three of these enzymesM-bM-^@M-^SPAPD1, ZCCHC6 and TUT1M-bM-^@M-^Shave not previously been known to modify miRNAs. Collectively, our results indicate that 3' modification observed in next generation small RNA sequencing data is a biologically relevant process, and identify enzymatic mechanisms that may lead to new approaches for modulating miRNA activity in vivo. We validated the specificity of the platform by assaying three pools of synthetic RNA oligonucleotides representing canonical, variant 1, and variant 2 versions of 5 miRNAs (miR-15a, miR-15b, miR-125a-5p, miR-143 and miR-221).Each bridge pool (i.e., specific to canonical, variant 1 or variant 2) was used to assay the three mixtures of synthetic miRNA oligonucleotides. In order to assess accuracy, we also assayed mixtures of 5 synthetic miRNAs containing 60% canonical miRNA, 30% variant 1, and 10% variant 2. Triplicate technical replicates of each sample were performed.

Dataset Information

Murine bone marrow-derived mast cells (BMMCs) transduced with pAPM lentiviruses overexpressing miR-221 vs control overexpressing miR-221m.

Publications

MiR-221 influences effector functions and actin cytoskeleton in mast cells.

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