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| Status |
Public on Dec 06, 2017 |
| Title |
Transcriptomal changes during differentiation of human iPS cells to PGC-like cells |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Pluripotent stem cell-derived human primordial germ cell (PGC)-like cells (hPGCLCs) may provide important opportunities to study human PGCs. We produced CD38+ hPGCLCs with a high efficiency [~43% of FACS-sorted embryoid body (EB) cells] from primed-pluripotency induced pluripotent stem cells (iPSCs) via 72-hour reprogramming towards ERK-independent naïve pluripotency. RNA-seq confirmed transcriptomal consistency of our hPGCLCs with hPGCLCs previously produced using various other methods. Immunohistochemical studies identified hPGCLCs exclusively at the outermost surface layer of EBs mostly as scattered cells, and live cell imaging revealed actively migrating hPGCLCs forming cellular protrusions. Both CD38+ hPGCLCs and CD38- EB cells significantly expressed PRDM1 and TFAP2C, but PRDM1 mRNA of CD38- cells lacked the 3’-untranslated region with let-7 and other miRNA binding sites known to regulate PRDM1 mRNA stability. Genes expressed specifically in hPGCLCs included early-PGC markers KIT, NANOS3, and SOX17, and cell migration genes, and their promoter sequences were enriched with TFAP2C binding motif. Whereas all hPGCLCs strongly expressed the CXCR4 chemotaxis receptor, its ligand CXCL12/SDF1 was not significantly expressed in the whole EBs. Exposure of hPGCLCs to CXCL12/SDF1 induced cell migration and anti-apoptosis genes; in contrast, genes involved in nuclear division were suppressed. Our study demonstrates a transcriptomal consistency of hPGCLCs produced using varying methods as well as distinct roles and/or regulation of PRDM1 and TFAP2C expression in development of human and mouse PGCs. Relevance of hPGCLCs to early-stage human embryonic PGCs randomly migrating in the midline region of human embryos before initiating directional chemotaxis under CXCR4-CXCL12/SDF1 signaling has also been suggested.
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| Overall design |
Human iPS cells with primed pluripotency were converted to naïve pluripotency for 72 hours and then subjected to embryoid body formation, from which CD38-positive PGC-like cells and CD38-negative cells were isolated by FACS. Total RNA was extracted from FACS-enriched cells and subjected to RNA-seq.
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| Contributor(s) |
Mitsunaga S, Shioda K, Shioda T |
| Citation(s) |
29087313 |
| |
| Submission date |
Aug 22, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Toshi Shioda |
| E-mail(s) |
tshioda@mgh.harvard.edu
|
| Phone |
(617) 726-3425
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| Organization name |
MGH Cancer Center
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| Department |
Center for Cancer Research
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| Lab |
Molecular Profiling Lab
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| Street address |
Building 149 - 7th Floor, 13th Street
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| City |
Charlestown |
| State/province |
MA |
| ZIP/Postal code |
02129 |
| Country |
USA |
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| Platforms (1) |
| GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
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| Samples (56)
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| Relations |
| BioProject |
PRJNA399518 |
| SRA |
SRP115953 |
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