Role of Zscan4 in secondary murine iPSC derivation mediated by protein extracts of ESC or iPSC.

Abstract

Previously, we found that the delivery of mouse ES (mES) cell-derived proteins to adult fibroblasts enables the full reprogramming of these cells, converting them to mouse pluripotent stem cells (protein-iPS cells) without transduction of defined factors. During reprogramming, global gene expression and epigenetic status such as DNA methylation and histone modifications convert from somatic to ES-equivalent status. mES cell extract-derived iPS cells are biologically and functionally indistinguishable from mES cells in its potential in differentiation both in vitro and in vivo. Furthermore, these cells show complete developmental potency. However, the efficiency of generating iPS by treatment with extract from mES cells is still low. In this report, we demonstrated that protein extracts of mouse iPS cells that were previously generated by mES cell extract treatment were able to reprogram somatic cells to become ES-like cells (secondary protein-iPS cells). We confirmed that fetal animals (E12.5) could be derived from these cells. Surprisingly, the efficiency of forming Oct4-positive colonies was remarkably improved by treatment of somatic cells with mouse iPS cell extract in comparison to treatment with mES cell extract. By screening the genes differentially expressed between mouse iPS and mES cells, Zscan4, which is known to enhance telomere elongation and stabilize genomic DNA, was identified as a strong candidate to promote efficiency of reprogramming. Interestingly, treatment with protein extracted from mES cells overexpressing Zscan4 enhanced formation of Oct4-positive colonies. Our results provide an efficient and safe strategy for reprogramming somatic cells by using mouse iPS cell extract. Zscan4 might be a key molecule involved in the demonstrated improvement of reprogramming efficiency.