Many biological processes are regulated by RNA-RNA interactions 1, nonetheless it remains formidable to analyze the entire RNA interactome. We developed a method, MARIO (MApping Rna-rna Interactions in vivO), to map protein-assisted RNA-RNA interactions in vivo. By circumventing the selection for a specific RNA-binding protein 2-5, our approach vastly expands the identifiable portion of the RNA interactome. Using this technology, we mapped the RNA interactome in mouse embryonic stem cells, which was composed of 46,780 RNA-RNA interactions. The RNA interactome was a scale-free network, with several lincRNAs and mRNAs emerging as hubs. We validated an interaction between two hubs, Malat1 and Slc2a3 using single molecule RNA fluorescence in situ hybridization. Base pairing was observed at the interaction sites of long RNAs, and was particularly strong in transposonRNA-mRNA and lincRNA-mRNA interactions. This reveals a new type of regulatory sequences acting in trans. Consistent with their hypothesized roles, the RNA interaction sites were more evolutionarily conserved than other regions of the transcripts. MARIO also provided new information on RNA structures, by simultaneously revealing the footprint of single stranded regions and the spatially proximal sites of each RNA. The unbiased mapping of the protein-assisted RNA interactome with minimum perturbation of cell physiology will greatly expand our capacity to investigate RNA functions. Overall design: Three (3) ESC samples were treated with one (1) type of antisense oligonucleotides as is described in Kretz M. et al. (Nature. 2013 Jan 10;493(7431):231-5) to show the RNA interaction among specific RNAs and verify the results from MARIO
Mapping RNA-RNA interactome and RNA structure in vivo by MARIO.
No sample metadata fields
View SamplesThe subunits of voltage-gated calcium channels regulate surface expression and gating of CaV1 and CaV2 1 subunits, and thus contribute to neuronal excitability, neurotransmitter release and calcium-induced gene regulation. In addition certain subunits are targeted into the nucleus, where they directly interact with the epigenetic machinery. Whereas their involvement in this multitude of functions is reflected by a great molecular heterogeneity of isoforms derived from four genes and abundant alternative splicing, little is known about the roles of individual variants in specific neuronal functions. In the present study, an alternatively spliced 4 subunit lacking the variable N-terminus (4e) is identified. It is highly expressed in mouse cerebellum and cultured cerebellar granule cells (CGC) and modulates P/Q-type calcium currents in tsA cells and CaV2.1 surface expression in neurons. Compared to the other two known full-length 4 variants (4a, 4b) 4e is most abundantly expressed in the distal axon, but lacks nuclear targeting properties. To examine the importance of nuclear targeting of 4 subunits for transcriptional regulation, we performed whole genome expression profiling of CGCs from lethargic mice individually reconstituted with 4a, 4b, and 4e. Notably, the number of genes regulated by each 4 splice variant correlated with the rank order of their nuclear targeting properties (4b> 4a> 4e). Together these findings support isoform-specific functions of 4 splice variant in neurons, with 4b playing a dual role in channel modulation and gene regulation, while the newly detected 4e variant serves exclusively in calcium channel-dependent functions.
Differential neuronal targeting of a new and two known calcium channel β4 subunit splice variants correlates with their regulation of gene expression.
Specimen part
View SamplesEye development is a multistep process that requires specific inductive signals and precise morphogenetic movements, starting early during development in the eye-field, a well-definite region of the anterior neural plate. It has been demonstrated that a gene network of eye field transcription factors (EFTFs) contributes to specify the neural and retinal fate of the eye field. Among these EFTFs, Xrx1 is involved in proliferation and neurogenesis in the eye field and is necessary for the correct development of the retina.
Brief report: Rx1 defines retinal precursor identity by repressing alternative fates through the activation of TLE2 and Hes4.
Specimen part
View SamplesHematopoietic Stem/Progenitor cells (HSPCs) are endowed with the role of maintaining a diverse pool of blood cells throughout the human life. Despite recent efforts, the nature of the early cell fate decisions remains contentious. Using single-cell RNA-Seq, we show that existing approaches to stratify bone marrow CD34+ cells reveal a hierarchically-structured transcriptional landscape of hematopoietic differentiation. Still, this landscape misses important early fate decisions. We here provide a broader transcriptional profiling of bone marrow lineage negative hematopoietic progenitors that recovers a key missing branchpoint into basophils and expands our understanding of the underlying structure of early adult human haematopoiesis. We also show that this map has strong similarities in topology and gene expression to that found in mouse. Finally, we identify the sialomucin CD164, as a reliable marker for the earliest branches of HSPCs specification and we showed how its use can foster the design of alternative transplantation cell products. Overall design: Single-cell mRNA sequencing of freshly isolated hematopoietic progenitors from human bone marrow. Sample HSC (Donor A) represents 1282 single cells. Sample MPP (Donor A) represents 215 single cells. Sample MLP (Donor A) represents 123 single cells. Sample PreB/NK (Donor A) represents 592 single cells. Sample MEP (Donor A) represents 1211 single cells. Sample CMP (Donor A) represents 1576 single cells. Sample GMP (Donor A) represents 1012 single cells. Sample Lin-CD34+CD164+ (Donor B) represents 6343 single cells. Sample Lin-CD34-CD164high (Donor B) represents 4434 single cells. Sample Lin-CD34lowCD164high (Donor B) represents 4266 single cells. Sample Lin-CD34-CD164low (Donor B) represents 358 single cells.
A comprehensive single cell transcriptional landscape of human hematopoietic progenitors.
Specimen part, Subject
View SamplesAt present, medical treatments of synchronous and metachronous liver metastases from colorectal cancer are not differentiated. The aim of the study was to analyze the gene expression profiling of synchronous and metachronous lesions in order to identify molecular signatures as possible basis for choice of systemic therapies. Fresh tissues specimens from metastases of 18 patients undergone liver surgery were collected (10 synchronous and 8 metachronous lesions). Gene expression profiling was studied using Affymetrix platform. Two different profiles were identified. Pathway related to the Epidermal Growth Factor receptor (EGFr) was upregulated in metachronous lesions whereas pathways mainly related to inflammation in synchronous lesions. Real Time-PCR, Western Blotting and ELISA confirmed that the metachronous lesions had the overexpression of EGFr, but the synchronous ones had the overexpression of Cyclo-oxygenase 2 (COX-2). These results suggest that synchronous or metachronous liver metastases from colorectal cancer could be differently treated on the basis of different molecular pathways.
Gene expression profiling of liver metastases from colorectal cancer as potential basis for treatment choice.
Specimen part
View SamplesThis SuperSeries is composed of the SubSeries listed below.
A molecular portrait of gastrointestinal stromal tumors: an integrative analysis of gene expression profiling and high-resolution genomic copy number.
Sex, Age, Specimen part
View SamplesIn addition to KIT and PDGFRA mutations, sequential accumulation of other genetic events is involved in the development and progression of gastrointestinal stromal tumors (GISTs). Until recently, the significance of these other alterations has not been thoroughly investigated. The combination of gene expression profiling and high-resolution genomic copy number analysis offers a detailed molecular portrait of GISTs, providing an essential comprehensive knowledge necessary to guide the discovery of novel target genes involved in tumor development and progression.
A molecular portrait of gastrointestinal stromal tumors: an integrative analysis of gene expression profiling and high-resolution genomic copy number.
Sex, Age, Specimen part
View SamplesCategorisation of LGGs related to their lesion site (infratentorial vs. supratentorial)
Molecular fingerprinting reflects different histotypes and brain region in low grade gliomas.
No sample metadata fields
View SamplesRNAseq analysis of YAP and Myc induced in quiescent and confluent 3T9 fibroblasts Overall design: RNAseq analysis of YAP and Myc induced in quiescent and confluent 3T9 fibroblasts
Transcriptional integration of mitogenic and mechanical signals by Myc and YAP.
Specimen part, Cell line, Subject
View SamplesTumors driven by activation of the transcription factor Myc generally show oncogene addiction. However, the gene-expression programs that depend upon sustained Myc activity in those tumors remain unknown. We have addressed this issue in a model of liver carcinoma driven by a reversible tet-Myc transgene, combining gene expression profiling with the mapping of Myc and RNA Polymerase II on chromatin. Switching off the oncogene in advanced carcinomas revealed that Myc is required for the continuous activation and repression of distinct sets of genes, constituting no more than half of those deregulated during tumor progression, and an even smaller subset of all Myc-bound genes. We further showed that a Myc mutant unable to associate with the co-repressor protein Miz1 is defective in the initiation of liver tumorigenesis. Altogether, our data provide the first detailed analysis of a Myc-dependent transcriptional program in a fully developed carcinoma, revealing that the critical effectors of Myc in tumor maintenance must be included within defined subsets (ca. 1,300 each) of activated and repressed genes. Overall design: RNAseq samples of control liver (n=11), tet-Myc tumors (n=16), tet-Myc tumors with short-term Myc inactivation (n=8), tet-MycVD tumors (n=11)
Identification of MYC-Dependent Transcriptional Programs in Oncogene-Addicted Liver Tumors.
Specimen part, Cell line, Subject
View Samples