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Induced Pluripotent Stem Cells from a Marsupial, the Tasmanian Devil (Sarcophilus harrisii): Insight into the Evolution of Mammalian Pluripotency.

We demonstrate the generation of Tasmanian devil (Sarcophilus harrisii) induced pluripotent stem cells (DeviPSCs) from dermal fibroblasts by lentiviral delivery of human transcription factors. DeviPSCs display characteristic pluripotent stem cell colony morphology, with individual cells having a high nuclear-to-cytoplasmic ratio and alkaline phosphatase activity. DeviPSCs are leukemia inhibitory factor dependent and have reactivated endogenous octamer-binding transcription factor 4 [OCT4, POU domain, class 5, transcription factor 1 (POU5F1)], POU2 [POU domain, class 5, transcription factor 3 (POU5F3)], sex determining region Y-box 2 (SOX2), Nanog homeobox (NANOG) and dosage-sensitive sex reversal, adrenal hypoplasia congenita critical region on the X chromosome, gene 1 (DAX1) genes, retained a normal karyotype, and concurrently silenced exogenous human transgenes. Notably, co-expression of both OCT4 and POU2 suggests that they are representative of cells of the epiblast, the marsupial equivalent of the inner cell mass. DeviPSCs readily form embryoid bodies and in vitro teratomas containing derivatives of all three embryonic germ layers. To date, DeviPSCs have been stably maintained for more than 45 passages. Our DeviPSCs provide an invaluable resource for studies into marsupial pluripotency and development, and they may also serve as an important tool in efforts to combat the threat of devil facial tumor disease.