Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of mouse T-cell thymocytes and bone marrow-derived macrophages dervied from Upf2fl/fl;LysMCre and Upf2fl/fl;LckCre mice to test the importance of nonsense-mediated mRNA decay on global transcription

ABSTRACT: Knockdown experiments in transformed human cells, suggested that nonsense-mediated mRNA decay (NMD) components had extensive effects on the regulation of a significant fraction of the transcriptome In order to test the importance of NMD on global transcription we subjected BMDMs and T-cell thymocytes derived from Upf2fl/fl;LysMCre and Upf2fl/fl;LckCre mice, respectively, to microarray-based gene expression profiling. Both these cell types display strong, if not complete, reduction of NMD as assayed by transfection with reporter constructs or sequencing of Tcrb transcripts. Upf2¯/¯ BMDMs displayed significant changes in the expression of 182 genes (234 probe sets) of which 179 were upregulated.

ORGANISM(S): Mus musculus

SUBMITTER: Charlotte Schjerling

PROVIDER: E-MEXP-1371 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

NMD is essential for hematopoietic stem and progenitor cells and for eliminating by-products of programmed DNA rearrangements.

Weischenfeldt Joachim J Damgaard Inge I Bryder David D Theilgaard-Mönch Kim K Thoren Lina A LA Nielsen Finn Cilius FC Jacobsen Sten Eirik W SE Nerlov Claus C Porse Bo Torben BT

Genes & development 20080501 10

Nonsense-mediated mRNA decay (NMD) is a post-transcriptional surveillance process that eliminates mRNAs containing premature termination codons (PTCs). NMD has been hypothesized to impact on several aspects of cellular function; however, its importance in the context of a mammalian organism has not been addressed in detail. Here we use mouse genetics to demonstrate that hematopoietic-specific deletion of Upf2, a core NMD factor, led to the rapid, complete, and lasting cell-autonomous extinction ...[more]

PMID: 18483223

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Project description:A major task in analyzing alternative splicing (AS) by RNA-Seq is to explore and quantify the differential representation of various splice classes and the protein-coding potential of the mapped reads. To this end, we have generated a streamlined bioinformatic pipeline available to the scientific community, which maps RNA-Seq reads to a complex combinatorial database of exon-exon junctions for the unique junction discovery, PTC detection and identification of splice isoforms. Hence, we have incorporated a splice isoform inference and PTC detection algorithm, which facilitates a highly accurate mapping and prediction of splice isoform junctions and nonsense-mediated mRNA decay (NMD)-susceptibility. We used this pipeline to investigate the complexity of the transcriptome and global role of AS-NMD in vivo in mammalian cells, by taking advantage of our Upf2 conditional knock-out mouse. Using tissue-specific Cre deleter strains, we have previously demonstrated in vivo that NMD acts to degrade many transcripts that results from AS. Also, ablating UPF2 (a core NMD factor) in the mouse leads to rapid mortality and collapse of most organs tested (Thoren et al., 2010; Weischenfeldt et al., 2008), suggesting an essential function for NMD in AS. To explore the effect of NMD on global splicing and to generate and validate an attractive bioinformatic pipeline for the scientific community to study AS and NMD, we chose to analyze two different mammalian organ systems with distinct phenotypes upon UPF2 ablation to test the robustness and dynamic range of the analysis. In one end of the spectrum, we chose to analyze the liver, wherein removal of UPF2 results in failure in liver metabolism and a high mortality rate (Thoren et al., 2010). To profile a less affected tissue, we chose to analyze bone marrow-derived macrophages (BMMs). These are macrophages that differentiate in vitro and Upf2 deleted BMMs are completely devoid of NMD activity but nevertheless show no morphological or functional phenotype compared to wild-type controls (Weischenfeldt et al., 2008). Thus, we were able to make a direct comparison between these two tissues to demonstrate the potency of our Isoform and PTC detection pipeline described below, which will allow the scientific community to analyze transcriptome data for global AS and NMD-susceptibility. Examination of UPF2 WT vs. KO in two different murine tissues (liver, bone marrow-derived macrophages).

Dataset Information

Transcription profiling of mouse T-cell thymocytes and bone marrow-derived macrophages dervied from Upf2fl/fl;LysMCre and Upf2fl/fl;LckCre mice to test the importance of nonsense-mediated mRNA decay on global transcription

Publications

NMD is essential for hematopoietic stem and progenitor cells and for eliminating by-products of programmed DNA rearrangements.

Similar Datasets

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