Generation of somatic chimeras using genetically-reprogrammed rabbits induced pluripotent stem cells [KEPi]

Little is known about the molecular underpinnings of pluripotent stem cells (PSCs)' ability to colonize preimplantation embryo epiblast to generate chimeras. Using rabbit PSCs as a model system, we conducted unbiased screening of a cDNA library encoding a panel of 36 pluripotency factors. From this, we identified KLF2, ERAS and PRMT6 whose overexpression endows the ability to self-renew 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 naïve pluripotency, including the reactivation of the 2nd X-chromosome. Leveraging on 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, and activation of genes involved in amino-acid metabolism, NF-kB signaling, and p53 pathway. Remarkably, these reprogrammed cells acquired the ability to produce chimeric embryos and fetuses with high contribution from PSCs in all major organs. Overall design: Transcriptomic profiling of chimeric competent KEP rabbit iPS cells.