Generation and gene expression profiles of Grevy's zebra iPSCs

Induced pluripotent stem cells (iPSCs) can provide biological resource for functional and conservation research for various species. However, the understanding of species variations of mammalian iPSCs is still limited. Here, we report the first generation of iPSCs from the endangered species Grevy's zebra (Equus grevyi; gz-iPSCs). We reprogram primary fibroblasts with human reprogramming transcription factors, OCT3/4, SOX2, KLF4, and c-MYC, with the retroviral method and confirmed the pluripotency and differentiation potential. In light of RNA sequencing analysis, generated gz-iPSCs robustly express genes associated with pluripotency and reprogramming processes, including epithelial-to-mesenchymal and mesenchymal-to-epithelial transitions (EMT-MET). Comparative transcriptomics with other species reveals patterns of gene expressions among mammalian PSCs and detects evolutionary conservation of pluripotency-associated genes and plausible importance of translation process. This work will aid in providing biological resource for this endangered species and enables new insight into the evolution of the mammalian PSCs. Overall design: We analyzed gene expression to characterize Grevy's zebra iPSCs reprogrammed from primary fibroblasts. Total RNA from Grevy's zebra iPSCs and fibroblasts, each at three different generations, and RNA-seq were performed using Oxford Nanopore Technologies long-read cDNA sequencing. The sequence reads were mapped to the horse reference genome, EquCab3.0, because reference genome is lacking in the Grevy's zebra at the time of analysis. The differentially expressed genes were estimated between iPSCs and fibroblasts, and Grevy's zebra transcriptome was further compared with other mammalian PSCs.