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| Status |
Public on Feb 16, 2019 |
| Title |
Allele-specific expression profiling of imprinted genes in mouse isogenic pluripotent tissues and cell lines |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
Genomic imprinting, resulting in parent-of-origin specific gene expression, plays a critical role in mammalian development. Here, we perform allele-specific RNA-Seq on isogenic B6D2F1 mice to assay imprinted genes in tissues from early embryonic stages and in pluripotent cell lines. For the cell lines, we include embryonic stem cells (ESCs) and epiblast stem cells (EpiSCs) derived from fertilized embryos or from embryos obtained after nuclear transfer (NT), as well as B6D2F1 ESCs and EpiSCs derived after parthenogenetic activation (PGA). Notably, the PGA-derived cell lines contain a mosaic genotype due to genomic recombination occurring in the parental B6D2F1 oocyte during meiosis. As the homozygous genomic regions of the PGA-derived cells are not compatible with allele-specific RNA-Seq, we developed an RNA-Seq based genotyping strategy that allows identification of the informative heterozygous regions. Global imprinting analysis shows that ESC lines largely lose imprints as compared to their corresponding embryonic tissues. Fertilized EpiSC and EpiSC-NT lines generally maintain imprinted gene expression. However, two EpiSC-NT lines show aberrant silencing of Rian and Meg3, two critically imprinted genes in mouse iPSCs. EpiSC-PGA lines display loss of imprinting, with known paternally-expressed genes being silenced and known maternally-expressed genes consistently showing doubled expression. Interestingly, most female EpiSC lines show monoallelic expression of Xist and full skewing of X inactivation, suggesting a (near) clonal origin. Together, our analysis provides a comprehensive overview of imprinted gene expression in pluripotency and provides a benchmark to allow identification of cell lines that faithfully maintain imprinted gene expression and therefore retain full developmental potential.
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| Overall design |
We include RNA-Seq on conventional B6D2F1 ESCs derived from fertilized embryos, three ESC lines derived from nuclear transfer embryos generated using B6D2F1 mouse embryonic fibroblast (MEF) donor nuclei (ESC-NT1) or B6D2F1 cumulus donor nuclei (ESC-NT2 and ESC-NT3), as well as three ESC lines derived from PGA embryos generated using B6D2F1 oocytes (ESC-PGA). Also included is RNA-Seq of three fertilized B6D2F1 EpiSC lines, three EpiSC lines derived from nuclear transfer embryos generated using a B6D2F1 cumulus donor nuclei (EpiSC-NT) and two EpiSC-PGA lines. For fertilized ESCs, ESC-NTs and ESC-PGAs we include 1 replica each of H3K4me3 and H3K27me3 ChIP-Seq epigenetic profiling.
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| Contributor(s) |
Dirks RA, van Mierlo G, Kerstens HH, Maruotti J, Bernardo AS, Kobolák J, Bock I, Pedersen RA, Dinnyés A, Huynen MA, Stunnenberg HG, Jouneau A, Marks H |
| Citation(s) |
30767785 |
| Submission date |
Jul 12, 2017 |
| Last update date |
Jul 25, 2021 |
| Contact name |
Hendrik Marks |
| E-mail(s) |
h.marks@ncmls.ru.nl
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| Organization name |
Radboud University Nijmegen, RIMLS
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| Department |
Molecular Biology
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| Street address |
Geert Grooteplein 26/28
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| City |
Nijmegen |
| ZIP/Postal code |
6525GA |
| Country |
Netherlands |
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| Platforms (3) |
| GPL11002 |
Illumina Genome Analyzer IIx (Mus musculus) |
| GPL13112 |
Illumina HiSeq 2000 (Mus musculus) |
| GPL19057 |
Illumina NextSeq 500 (Mus musculus) |
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| Samples (22)
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| Relations |
| BioProject |
PRJNA393966 |
| SRA |
SRP111681 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE101292_RAW.tar |
128.8 Mb |
(http)(custom) |
TAR (of WIG) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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