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| Status |
Public on Mar 02, 2019 |
| Title |
The nuclei of somatic cells fused to mouse embryonic stem cells express pluripotency markers similar to early reprogramming events |
| Organisms |
Bos taurus; Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
The mechanisms that direct nuclear reprogramming to a pluripotent state are still not fully understood. For species in which the derivation of true induced pluripotent stem (iPS) cells has not yet been completely successful, insights into the particular mechanisms that govern pluripotency can help in determining which are the appropriate conditions to induce stable reprogramming in somatic cells. Cell fusion of a somatic cell to a pluripotent cell is known to induce expression of pluripotency markers in the somatic nucleus. We hypothesized that fusion of bovine fetal fibroblasts (bFFs) to mouse embryonic stem (mES) cells would induce expression of pluripotency markers in the bFF nucleus. We have previously established a method to produce and specifically select multinucleated cells originated from both mES cell and bFF (bi-species heterokaryon), using indirect immunofluorescence on live cells and the combined use of imaging and sorting flow cytometry. With these tools in place, we analyzed 200 heterokaryons collected at 24, 48 and 72h after fusion, as well as unfused bFFs and mES cells, and analyzed using RNA-seq. We found significant changes in bovine gene expression patterns between bFFs and heterokaryons obtained 24h after fusion. Upregulation of early pluripotency markers OCT4 and KLF4, as well as hypoxia response genes, contrasted with downregulation of cell cycle inhibitors such as SST. The cytokine IL6, known to increase survival of early embryos in vitro, was also upregulated in heterokaryons, although its role and mechanism of action is still unclear. The cell fusion model presented here can be used to characterize early changes in somatic nuclear reprogramming, and to study the effect of different conditions during reprogramming.
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| Overall design |
Five samples total. Bi-species (mouse/bovine) heterokaryons were produced and sampled at 24, 48, and 72h after fusion. Samples of mouse only and bovine only cells were harvested in parallel as controls.
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| Contributor(s) |
Villafranca C, Eyestone WH, Jensen RV |
| Citation missing |
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| Submission date |
Mar 01, 2019 |
| Last update date |
Mar 03, 2019 |
| Contact name |
M. Cristina Villafranca |
| E-mail(s) |
villm07@vt.edu
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| Organization name |
Virginia Polytechnic Institute and State University
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| Street address |
205 Duckpond Drive
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| City |
Blacksburg |
| State/province |
VA |
| ZIP/Postal code |
24061 |
| Country |
USA |
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| Platforms (3) |
| GPL16417 |
Illumina MiSeq (Mus musculus) |
| GPL19646 |
Illumina MiSeq (Bos taurus) |
| GPL26244 |
Illumina MiSeq (Bos taurus; Mus musculus) |
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| Samples (5)
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| Relations |
| BioProject |
PRJNA525141 |
| SRA |
SRP187283 |
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