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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Aug 01, 2018 |
| Title |
Deconstructing lincRNA regulation during ESC to NPC differentiation |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
Other
Non-coding RNA profiling by high throughput sequencing
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| Summary |
Using single-cell and bulk RNA-sequencing, we profiled mouse embryonic stem cells and neural precursor cells to identify lincRNAs with cell type- and subpopulation-specific expression. In contrast to mRNAs, we found that lincRNAs are prone to prematurely terminate transcription, are less abundant across single cells, and are degraded by the nuclear exosome. These properties indicate that lincRNA genes widely function without requiring their RNA product. To test this prediction experimentally, we developed a method that separates the role of a lincRNA gene from the role of its RNA product. This revealed RNA-independent lincRNA gene activities, underscoring the importance of distinguishing the roles of transcription, the genomic locus, and the RNA product when studying non-coding loci, for which we are providing suitable tools.
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| Overall design |
Two mESC lines (BL6xCAST and CASTxBL6) were differentiated into neural precursor cells, as well as being sampled in 2i+LIF and in serum+LIF medium. Bulk RNA-seq was performed for all timepoints (2i+LIF, serum+LIF, day3, day6 and day8 of differentiation). Most timepoints were also sampled by single-cell RNA-seq (96 cells per sample, 7 samples in total). Cell lines with lincRNA genomic deletions, or with the hammerhead ribozyme genomically integrated into lincRNA loci, were also analysed by bulk RNA-seq (140 cell lines in total). We also performed CRosslinking and Analysis of CDNAs (CRAC) in wild-type cells or cells where endogenous Mtr4 is tagged with a FLAG-Bio tag, to examine transcriptome-wide targets of the nuclear RNA surveillance factor Mtr4. For normalisation, we performed NET-seq. To measure global RNA half-lives, we treated wild-type mESCs with actinomycin D, and performed bulk RNA-seq at several timepoints, in triplicate (3 wells of cells were analysed in parallel per timepoint).
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| Contributor(s) |
Tuck A, Natarajan K, Mohn F |
| Citation(s) |
30456373 |
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Alex C Tuck, Kedar Nath Natarajan, Greggory M Rice, Jason Borawski, Fabio Mohn, Aneliya Rankova, Matyas Flemr, Alice Wenger, Razvan Nutiu, Sarah Teichmann, and Marc Bühler. Distinctive features of lincRNA gene expression suggest widespread RNA-independent functions. Life Science Alliance 2018 Jul;1(4):e201800124. DOI:10.26508/lsa.201800124
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| Submission date |
Nov 29, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Alex Charles Tuck |
| Organization name |
Friedrich Miescher Institute for Biomedical Research
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| Street address |
Maulbeerstrasse 66
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| City |
Basel |
| ZIP/Postal code |
4058 |
| Country |
Switzerland |
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| Platforms (2) |
| GPL13112 |
Illumina HiSeq 2000 (Mus musculus) |
| GPL17021 |
Illumina HiSeq 2500 (Mus musculus) |
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| Samples (840)
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| Relations |
| BioProject |
PRJNA420368 |
| SRA |
SRP125831 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE107493_RAW.tar |
26.6 Gb |
(http)(custom) |
TAR (of BIGWIG) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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