Similar Datasets
Project description:Differences in codon frequency between genomes, genes, or positions along a gene, modulate transcription and translation efficiency, leading to phenotypic and functional differences. Integrative studies quantifying the phenotypic consequences of codon usage bias at different molecular and cellular levels in human cells are lacking. Here, we present a multiscale analysis of the effects of synonymous codon recoding during heterologous gene expression in human cells. Six synonymous versions of an antibiotic resistance gene were generated, fused to a fluorescent reporter, and independently expressed in HEK293 cells. Multiscale phenotype was analysed by means of: quantitative transcriptome and proteome assessment, as proxies for gene expression; cellular fluorescence, as a proxy for single-cell level expression; and real-time cell proliferation in absence or presence of antibiotic, as a proxy for the cell fitness. We show that differences in codon usage bias strongly impact the molecular and cellular phenotype: (i) they result in large differences in mRNA and in protein levels, as well in mRNA-to-protein ratio; (ii) they introduce unpredicted splicing events; (iii) they lead to reproducible phenotypic heterogeneity; and (iv) they lead to a trade-off between the benefit of antibiotic resistance and the burden of heterologous expression. In human cells in culture, codon usage bias modulates gene expression by modifying mRNA availability and suitability for translation, leading to differences in protein levels and eventually eliciting functional phenotypic changes.
2024-05-24 | PXD038324 | Pride
Project description:Investigating PCR bias in TnSeq
| PRJNA684144 | ENA
Project description:To characterize the rules governing exon recognition during splicing, we analyzed RNA-seq datasets and identified ~4,000 GC-rich and ~5,000 AT-rich exons, labelled GC-exons and AT-exons, respectively whose inclusion depends on different sets of splicing factors. We show that a high GC-load is associated with predicted RNA secondary structures at 5'ss and that GC-exons are dependent on U1 snRNP-associated proteins. Meanwhile, a high AT-load is associated with a large number of decoy splicing-related signals upstream exons such as the number of branchpoints and SF1- or U2AF65-binding sites and AT-exons are dependent on U2 snRNP-associated proteins. Nucleotide composition bias also influences local chromatin organization. Since the GC content of exons correlates with that of their hosting-genes, -isochores and – topologically-associated domains, we propose that regional nucleotide composition bias leaves a footprint locally, at the exon level, inducing, during splicing, constraints that are alleviated by the local chromatin organization and specific splicing factors.
2019-10-25 | GSE138397 | GEO
Project description:Study of the consequences of expression of Sf3b1K700E in haematopoietic cells on splicing of messenger RNA and on transcript expression levels
2015-07-21 | E-ERAD-379 | biostudies-arrayexpress
Project description:The clustering of mass spectra is a critical component of many proteomics applications. The clustering validation science is just as important, having evolved side by side with the clustering algorithms themselves. In this work, we build on Rieder et al. 's cluster validation framework, and we discuss the problem of selection bias in cluster validation measures; we introduce an assessment measure that is biased toward the number of peptide ion species; we introduce a cluster assessment framework for proteomics and demonstrate its importance by evaluating the performance of 8 clustering algorithms in 7 proteomics datasets, and we discuss the tradeoff between assessment measures. Finally, the validation methods presented here can be of broad applicability be-yond the clustering of mass spectra. This PRIDE entry describes in detail sample preparation, LC-MS/MS analysis, and protein identification of one of the proteomics datasets used in this work (> 10 kDa Bothrops jararaca snake venom proteome).
2022-03-16 | PXD022124 | Pride
Project description:Here we show that the pioneer transcription factors Foxa1 and Foxa2 are required to regulate RNA splicing during positive selection at the transition from CD4+CD8+ double positive (DP) to single positive (SP thymocyte). Conditional deletion of both Foxa1 and Foxa2 from DP thymocytes led to a partial arrest at the transition from DP to SP, with a reduction in the number of cells undergoing positive selection, and reduced the number and maturation of both CD4SP and CD8SP populations, with a reduction in peripheral naïve CD4+ T-cells. Foxa1 and Foxa2 were required for physiological levels of expression of several genes which encoding splicing factors (Mbnl1, Sf3b1, Hnrnpa1) in cells undergoing positive selection. Within the positively selecting CD69+DP cells, alternative RNA splicing was dysregulated leading to more than 859 significantly differentially expressed exons compared to control, with many genes important for T cell development and for RNA splicing showing differentially used exons.
2021-07-13 | GSE169602 | GEO
Project description:Recurrent mutations in the spliceosome are observed in several human cancers but their functional and therapeutic significance remain elusive. SF3B1, the most frequently mutated component of the spliceosome in cancer, is involved in the recognition of the branch point sequence (BPS) during selection of the 3’ splice site (ss) in RNA splicing. Here, we report that common and tumor-specific splicing aberrations are induced by SF3B1 mutations and establish aberrant 3’ ss selection as the most frequent splicing defect. Strikingly, mutant SF3B1 utilizes a BPS that differs from that used by wild-type SF3B1 and requires the canonical 3’ ss to enable aberrant splicing during the second step. Approximately 50% of the aberrantly spliced mRNAs are subjected to nonsense-mediated decay resulting in downregulation of gene and protein expression. These findings ascribe functional significance to the consequences of SF3B1 mutations in cancer. 72 samples, including two sets of patient data and cell lines with two additional technical replicates each
2015-09-08 | E-GEOD-72790 | biostudies-arrayexpress
Project description:Recurrent mutations in the spliceosome are observed in several human cancers but their functional and therapeutic significance remain elusive. SF3B1, the most frequently mutated component of the spliceosome in cancer, is involved in the recognition of the branch point sequence (BPS) during selection of the 3’ splice site (ss) in RNA splicing. Here, we report that common and tumor-specific splicing aberrations are induced by SF3B1 mutations and establish aberrant 3’ ss selection as the most frequent splicing defect. Strikingly, mutant SF3B1 utilizes a BPS that differs from that used by wild-type SF3B1 and requires the canonical 3’ ss to enable aberrant splicing during the second step. Approximately 50% of the aberrantly spliced mRNAs are subjected to nonsense-mediated decay resulting in downregulation of gene and protein expression. These findings ascribe functional significance to the consequences of SF3B1 mutations in cancer.
2015-09-08 | GSE72790 | GEO
Project description:Neuronal alternative splicing is dynamically regulated in a spatiotemporal fashion. We previously found that STAR family proteins (SAM68, SLM1, SLM2) regulate spatiotemporal alternative splicing in the nervous system. However, the whole aspect of alternative splicing programs governed by STARs remains unclear. We deciphered the alternative splicing programs of SAM68 and SLM1 proteins using transcriptomics. We reveal that SAM68 and SLM1 encode distinct alternative splicing programs; SAM68 preferentially controls alternative last exon (ALE) splicing. Interleukin 1-receptor accessory protein (Il1rap) is a novel target for SAM68. The usage of Il1rap ALEs results in mainly two variants encoding two functionally different isoforms, a membrane-bound (mIL1RAcP) and a soluble (sIL1RAcP) type. The brain exclusively expresses mIL1RAcP. SAM68 knockout results in remarkable conversion into sIL1RAcP in the brain, which significantly disturbs IL1RAcP neuronal function. Thus, we uncover the critical role of proper neuronal isoform selection through ALE choice by the SAM68-specific splicing program.
2019-12-29 | GSE110258 | GEO
Project description:Small molecule splicing modifiers have been extensively described which target the generic splicing machinery and thus have low target specificity. We have identified potent splicing modifiers with unprecedented high selectively, correcting the splicing deficit of the SMN2 (survival motor neuron 2) gene in Spinal Muscular Atrophy (SMA). Here we show that they directly bind to two sites of the SMN2 pre-mRNA, thereby stabilizing a novel ribonucleoprotein (RNP) complex in the SMN2 gene that is critical for the high target specificity of these small molecules over other genes. In addition to the therapeutic potential of these molecules for treatment of SMA, our work may have wide-ranging consequences for further research to identify small molecules that target splicing correction of specific genes by interacting with tertiary RNA structures.
2017-08-11 | GSE86001 | GEO