OCT4 is expressed in extraembryonic endoderm stem (XEN) cell progenitors during somatic cell reprogramming.

During development, progenitors of embryonic stem (ES) and extraembryonic endoderm stem (XEN) cells are concomitantly specified within the inner cell mass (ICM) of the mouse blastocyst. Similarly, XEN cells are induced (iXEN cells) alongside induced pluripotent stem (iPS) cells following overexpression of Oct4, Sox2, Klf4 and Myc (OSKM) during somatic cell reprogramming. It is unclear how or why this cocktail produces both stem cell types, but OCT4 has been associated with non-pluripotent outcomes. In this report, we show that, during OSKM reprogramming, many individual Oct4-GFP-expressing cells are fated to become iXEN cells. Interestingly, SKM alone was also sufficient to induce iXEN cell formation, likely via activation of endogenous Oct4. These observations indicate that iXEN cell formation is not strictly an artifact of Oct4 overexpression. Moreover, our results suggest that a pathway to XEN may be an integral feature of establishing pluripotency during reprogramming, as in early embryo development.

Fluorescent Reporters Distinguish Stem Cell Colony Subtypes During Somatic Cell Reprogramming.

Somatic cell reprogramming was first developed to create induced pluripotent stem (iPS) cells. Since that time, the highly dynamic and heterogeneous nature of the reprogramming process has come to be appreciated. Remarkably, a distinct type of stem cell, called induced extraembryonic endoderm (iXEN) stem cell, is also formed during reprogramming of mouse somatic cells by ectopic expression of the transcription factors, OCT4, SOX2, KLF4, and MYC (OSKM). The mechanisms leading somatic cells to adopt differing stem cell fates are challenging to resolve given that formation of either stem cell type is slow, stochastic, and rare. For these reasons, fluorescent gene expression reporters have provided an invaluable tool for revealing the path from the somatic state to pluripotency. However, no such reporters have been established for comparable studies of iXEN cell formation. In this study, we examined the expression of multiple fluorescent reporters, including Nanog, Oct4, and the endodermal genes, Gata4 and Gata6-alone and in combination, during reprogramming. We show that only simultaneous evaluation of Nanog and Gata4 reliably distinguishes iPS and iXEN cell colonies during reprogramming.

Capturing and Interconverting Embryonic Cell Fates in a Dish.

Cells of the early embryo are totipotent because they will differentiate to produce the fetus and its surrounding extraembryonic tissues. By contrast, embryonic stem (ES) cells are considered to be merely pluripotent because they lack the ability to efficiently produce extraembryonic cell types. The relatively limited developmental potential of ES cells can be explained by the observation that ES cells are derived from the embryo after its cells have already begun to specialize and lose totipotency. Meanwhile, at the time that pluripotent ES cell progenitors are specified, so are the multipotent progenitors of two extraembryonic stem cell types: trophoblast stem (TS) cells and extraembryonic endoderm stem (XEN) cells. Notably, all three embryo-derived stem cell types are capable of either self-renewing or differentiating in a lineage-appropriate manner. These three types of embryo-derived stem cell serve as paradigms for defining the genes and pathways that define and maintain unique stem cell identities. Remarkably, some of the mechanisms that maintain the specific developmental potential of each stem cell line do so by preventing conversion to another stem cell fate. This chapter highlights noteworthy studies that have identified the genes and pathways that normally limit the interconversion of stem cell identities.