Generation of Systemic Chimeras via Rabbit Induced Pluripotent Stem Cells Reprogrammed with KLF2, ERAS, and PRMT6

Little is known about the molecular underpinnings of pluripotent stem cells’ (PSCs) ability to colonize the epiblast of preimplantation embryos and generate chimeras. In our study, using rabbit PSCs as a model system, we conducted unbiased screening of a cDNA library that encodes a panel of 36 pluripotency factors. From this screening, we identified KLF2, ERAS and PRMT6, whose overexpression confers the ability for self-renewal in a KOSR/FGF2-free culture medium supplemented with LIF, activin A, PKC and WNT inhibitors. The reprogrammed cells acquired transcriptomic and epigenetic features of naive pluripotency, including the reactivation of the 2 ^nd X-chromosome. Leveraging these PSC lines, we determined the transcriptomic signature of embryonic colonization-competence, demonstrating transcriptional repression of genes involved in MAPK, WNT, HIPPO, and EPH signaling pathways, alongside the activation of genes involved in amino-acid metabolism, NF-kB signaling, and p53 pathway. Remarkably, a subset of reprogrammed cells, expressing CD75 at a high level, gained the ability to produce chimeric fetuses with a high contribution from PSCs in all lineages.