RNA sequencing of blastocyst-stage mouse embryos in the presence or absence of RNA decay activator UPF2
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ABSTRACT:
PROVIDER: PRJNA789489 | ENA |
REPOSITORIES: ENA
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Project description:Purpose: The goal of this study is to compare transcriptome profiles of whole blastocysts in the presence or absence of UPF2, via an approach optimized for low-input samples. Methods: Blastocyst mRNA profiles of wild-type (WT) and Upf2 knockout (Upf2−/−) mice were generated by deep sequencing, using Illumina 2500. The sequence reads that passed quality filters were analyzed at the transcript isoform level with DESeq2 and with IsoformSwitchAnalyzeR. Results: We identified transcripts targeted for decay by UPF2 and the NMD pathway in the blastocyst via multiple methods. We found that NMD regulates a cohort of RNAs involved in cell survival.
2022-03-16 | GSE191039 | GEO
Project description:We have HA-tagged two nonsense-mediated decay proteins, UPF1 and UPF2, in the malaria parasite Plasmodium falciparum. We then performed co-immunoprecipitation experiments to determine the protein-protein interactions of these nonsense-mediated decay components.
2021-04-16 | PXD023910 | Pride
Project description:Nonsense-mediated decay (NMD) is an evolutionarily conserved surveillance mechanism that targets mRNAs undergoing premature translation termination for rapid degradation as well as normal physiological transcripts. From yeast to humans, NMD requires the function of three conserved Up-frameshift (Upf) factors (Upf1, Upf2, Upf3). Interestingly, in humans, mutations in UPF2 and UPF3B are associated with intellectual disability and autism spectrum disorder (ASD). However, the neurobiological mechanism by which deficient NMD leads to neurodevelopmental disorders remains unknown. Consequently, no treatment is currently available. Here we report that mice lacking Upf2 in the murine forebrain (Upf2 fb-KO mice) show endophenotypes associated with ASD and deficits in learning and memory. In addition, Upf2-mediated deficient NMD leads to abnormal long-term potentiation (LTP) in the hippocampus. Like patients with mutations in UPF2, Upf2 fb-KO mice showed neuroanatomical abnormalities including a smaller corpus callosum. Surprisingly, transcriptomic analysis revealed elevated mRNA expression of immune-related genes in the hippocampus of Upf2-fb-KO mice, which was accompanied by increased inflammation and progressive infiltration of peripheral immune cells. More importantly, treatment with an FDA-approved immunosuppressant, cyclophosphamide (CyP), significantly reduces brain inflammation, improves the neuroanatomical defects and the deficits in LTP and memory deficits, and reverses some of the ASD-like behaviors, notably the social deficits. Collectively, our results reveal the biological basis underlying neurodevelopmental disorders associated with dysfunctional NMD. Moreover, these findings suggest that immunosuppressant drugs, like CyP, may provide a new therapeutic approach for the treatment of patients with mutations in core NMD components.
2020-06-29 | GSE81591 | GEO
Project description:Over one million cases of gastric cancer are diagnosed each year and metastatic disease continues to have a poor prognosis. A significant proportion of gastric tumors have defects in the DNA damage response pathway creating therapeutic opportunities through synthetic lethal approaches. In particular, several small molecule inhibitors of Ataxia-Telangiectasia Mutated and Rad3-related protein kinase (ATR), a key regulator of the DNA Damage response, are now in clinical development as targeted agents for gastric cancer. Here, we sought to discover genetic determinants of response and resistance to ATRi in gastric cancer cells. We performed a large-scale CRISPR interference screen to discover determinants of resistance to ATRi in gastric cancer. Top hits were validated and further evaluated to define mechanisms of resistance. We identified components of the nonsense-mediated decay pathway and the UPF2 gene, in particular, as mediators of sensitivity to ATRi. We show loss of UPF2 causes resistance to ATRi through the abrogation of ATRi mediated cell cycle arrest and find evidence that UPF2 regulates R-loops. Our results uncover a novel role for NMD factors in R-loop formation and resistance to ATRi in gastric cancer cells, potentially through R-loop formation.
2022-11-06 | PXD034887 | Pride
Project description:The Upf1, Upf2 and Upf3 proteins cooperate to implement nonsense-mediated decay. We used DNA microarrays for profiling RNA expression levels of upf2 and upf3 mutants
2013-06-03 | E-MTAB-1325 | biostudies-arrayexpress
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).
2012-05-01 | E-GEOD-26561 | biostudies-arrayexpress
Project description:All eukaryotes studied to date have the capacity to detect and degrade mRNAs harboring premature translation termination codons (PTCs) in a process called nonsense-mediated mRNA decay (NMD) (reviewed in Wagner E & Lykke-Andersen J, 2002). This surveillance system allows the cell to prevent the expression of potentially harmful truncated proteins. trans-acting factors required for NMD in Drosophila are the proteins UPF1, UPF2, UPF3, SMG-1, SMG-5 and SMG-6. To identify mRNAs naturally regulated by NMD, we analyzed expression profiles in Drosophila cells RNAi-depleted of known NMD factors using high-density oligonucleotide arrays.
2005-11-01 | E-MEXP-202 | biostudies-arrayexpress
Project description:mRNA level is controlled by factors that mediate both mRNA synthesis and decay, including the 5’ to 3’ exonuclease Xrn1 - a major mRNA synthesis and decay factor. Here we show that nucleocytoplasmic shuttling of several mRNA decay factors plays a key role in determining both mRNA synthesis and decay. Shuttling is regulated by RNA-controlled binding of the karyopherin Kap120 to two nuclear localization sequences (NLSs) in Xrn1, location of one of which is conserved from yeast to human. The decaying RNA binds and masks NLS1, establishing a link between mRNA decay and Xrn1 shuttling. Preventing Xrn1 import, either by deleting KAP120 or mutating the two Xrn1 NLSs, compromise transcription and, unexpectedly, also the cytoplasmic decay, uncovering a cytoplasmic decay pathway that initiates in the nucleus. Most mRNAs are degraded by both the “classical” and the novel pathways, the ratio between them represents a full spectrum. Importantly, Xrn1 shuttling is required for proper adaptation to environmental changes, in particular to ever changing environmental fluctuations.
2022-09-19 | PXD036414 | Pride
Project description:Comparative trascriptomic analysis between porcine early-blastocyst and Hatched blastocyst collected around day 5-6 direct comparison with dye-swap; two different arrays with 6 Samples each
2015-06-21 | E-GEOD-38882 | biostudies-arrayexpress
Project description:Comparative trascriptomic analysis between porcine early-blastocyst and Hatched blastocyst collected around day 5-6
2015-06-21 | GSE38882 | GEO