Research

Understanding Naïve and Primed Pluripotent States

Being able to generate all cell types, mouse embryonic stem cells are a most valuable tool for research. They can be found in the developing mouse embryo in two distinct states: naïve – in the blastocyst, and primed – in the post-implantation epiblast. These two states are distinct in various aspects, most notable, only naïve cells can contribute efficiently to chimera. Naïve and primed cells can be sustained in-vitro, and are dependent on distinct signaling. In human, naïve stem cells were out of reach for a long time. We investigate the regulation of naïve and primed pluripotent stem cell in mouse and human. Specifically, we were able to maintain human stem cells in a “naive” state, with distinct molecular and functional properties, including enhanced ability to contribute to cross-species mouse chimeric embryos (Gafni et al, 2013). In addition, we found that mRNA methylation has a critical role in facilitating degradation of pluripotent genes, an essential step during the switch from naïve to primed states, both in-vitro and in-vivo (Geula et al, 2014). Our current studies involve elucidating molecular regulation of these states across different species, and define how their molecular architecture dictates their functional competence.

 

Naive human stem cells share defining molecular features with mouse ES cells

a, WIBR3 human ES cells were stably transfected with the different engineered BAC reporter constructs indicated. GFP levels were evaluated in genetically matched cells by flow cytometry. MFI, mean fluorescence intensity. Average MFI values of 3 biological replicates per sample are indicated. b, Representative confocal images obtained after double immunostaining for OCT4 and H3K27me3 on naive, primed and differentiated samples (obtained from naive cells). Average percentages of H3K27me3 foci-positive nuclei out of 150–200 cells per sample (arrowheads) are indicated. EBs, embryonic body differentiated cells. c, Unbiased global transcriptional cross-species hierarchical clustering of naive and primed pluripotent cells from mice and humans. Correlation matrix of gene expression was clustered using Spearman correlation and average linkage. Colour bar indicates correlation strength. Each row/column represents an independent cell line or clone. mEpiSCs, murine EpiSCs; mESC, mouse ES cell; miPSC, mouse iPS cell. NOD indicates non-obese diabetic ICR mice. d, Naive and primed human ES cells were double immunostained for TFE3 and OCT4. Representative confocal images are shown for WIBR3 human ES cells. Insets are enlargements of the dashed boxes. Scale bars, 50µm.