Transcription factor heterogeneity and epiblast pluripotency

Rodrigo Osorno 0 Ian Chambers () 0 0 Institute for Stem Cell Research, MRC Centre for Regenerative Medicine, School of Biological Sciences, University of Edinburgh , King's Buildings, West Mains Road, Edinburgh EH9 3JQ , UK Articles on similar topics can be found in the following collections Receive free email alerts when new articles cite this article - sign up in the box at the top right-hand corner of the article or click here References Subject collections Email alerting service Transcription factor heterogeneity and epiblast pluripotency Rodrigo Osorno and Ian Chambers* Stem cells are defined by the simultaneous possession of the seemingly incongruent properties of selfrenewal and multi-lineage differentiation potential. To maintain a stem cell population, these opposing forces must be balanced. Transcription factors that function to direct pluripotent cell identity are not all equally distributed throughout the pluripotent cell population. While Oct4 levels are relatively homogeneous, other transcription factors, such as Nanog, are more heterogeneously expressed. Moreover, Oct4 positive cells fluctuate between states of high Nanog expression associated with a high probability of self-renewal and low Nanog expression associated with an increased propensity to differentiate. As embryonic stem (ES) cells transit to the more developmentally advanced epiblast stem cell (EpiSC) state, the levels of pluripotency transcription factors are modulated. Such modulations are blunted in cells that overexpress Nanog and this may underlie the resistance of Nanog-overexpressing cells to transit to an EpiSC state. Interestingly, increasing the levels of Nanog in EpiSC can facilitate reversion to the ES cell state. Together these observations suggest that Nanog lies close to the top of the hierarchy of pluripotent transcription factor regulation. 1. INTRODUCTION Stem cells are defined by the possession of the dual properties of self-renewal and differentiation potential. The cells with the greatest developmental potential are cells that have the ability to contribute to each of the three embryonic germ layers and are known as pluripotent stem cells. Pluripotent stem cells exist in the early embryo and in teratocarcinomas, tumours that spontaneously arise in the testes of specific mouse strains (reviewed by [1,2]). The first pluripotent cells to be isolated and propagated in vitro, while retaining demonstrable pluripotency, were derived indirectly from teratocarcinomas following their in vivo passage as ascites and are referred to as embryonal carcinoma (EC) cells [3]. EC cells are the undifferentiated cells of teratocarcinomas, and conditions tuned to the in vitro propagation of these cells [4] allowed the subsequent isolation of pluripotent embryonic stem (ES) cells [5,6]. Mouse ES cells are pluripotent stem cells derived directly from the inner cell mass (ICM) of the blastocyst between embryonic day (E)3.5 and E4.5. At E4.5, the blastocyst contains three cell types; the trophectoderm, the hypoblast and the epiblast. While the trophectoderm and the hypoblast contribute to extraembryonic tissues, the epiblast gives rise to all cell types of the developing embryo proper. This pre-implantation epiblast cell population contains cells that have the ability to differentiate into derivatives One contribution of 15 to a Discussion Meeting Issue What next for stem cell biology? The evolving biology of cell reprogramming. of all three somatic lineages [7 9] and the germline [10]. ES cells retain these pluripotent characteristics provided they are maintained in appropriate culture conditions, e.g. in leukaemia inhibitory factor (LIF)/ bone morphogenetic protein (BMP). At around E5.5, the embryo implants into the uterus and the epiblast undergoes molecular and cellular changes. However, post-implantation epiblast cells remain pluripotent and can give rise to cell lines in vitro, called epiblast stem cells (EpiSCs) [11,12]. EpiSCs differ from ES cells in that they have a flattened colony morphology, inefficient clonal propagation and rather than requiring LIF/BMP to maintain their pluripotency, require activin/fibroblast growth factor (FGF). EpiSCs are demonstrably pluripotent as they can generate teratocarcinomas containing differentiated cells of each of the somatic germ layers. However, EpiSCs lack the capacity to integrate into a pre-implantation embryo and thus contribute to the soma or germline of chimeric animals. This may suggest that EpiSCs have reduced potency relative to ES cells, or alternatively, that they have features incompatible with incorporation into the blastocyst. The observation that EpiSCs in culture can give rise to primordial germ cells (PGC) indicates that at least some cells in an EpiSC culture have retained germline potential [13]. Here, we discuss how transcription factors regulate the distinct identities and behaviours of these pluripotent cells. 2. GENE REGULATORY NETWORKS GOVERNING PLURIPOTENT CELL STATES Pluripotent stem cell self-renewal efficiency is governed by a gene regulatory network centred around 2230 Heterogeneity of the epiblast R. Osorno & I. Chambers 2231 the triumvirate of transcription factors Oct4, Sox2 and Nanog [14]. Oct4 belongs to the class of transcription factors that possess a bipartite POU domain composed of two DNA-binding domains, a homeodomain (POUHD) and a POU-specific domain (POUS). Based on studies conducted on Oct1, these domains are considered to be able to bind DNA separately. POUHD has a much higher affinity for DNA than POUS, but together both domains bind DNA cooperatively [15]. Oct4 is expressed in pluripotent cells and is specifically required for cells that become allocated to the interior of the blastocyst to acquire a pluripotent identity [16]. In ES cells, several Oct4 targets have been functionally identified including Oct4 [17], Opn [18], FGF4 [19], Utf1 [20], Fbx15 [21], Nanog [22], Zfp42 (Rex1) [23], Esrrb [24], Lefty1 [25], Cdx2 [26] and Sox2 [27]. In addition, global chromatin localization studies have identified thousands of sites throughout the ES cell epigenome to which Oct4 binds and which are therefore hypothesized to be involved in the control of expression of nearby genes [28 30]. The expression level of Oct4 is a critical determinant of the phenotype of ES cells and derivatives. In line with the in vivo phenotype, deletion of Oct4 results in differentiation of cells to a trophectodermal type [31]. More surprisingly, the elevation of Oct4 levels causes differentiation of ES cells into a mixed cell population that expresses markers of endoderm and mesoderm [31]. Therefore, Oct4 expression needs to be constrained within tight boundaries in order to sustain ES cell self-renewal [31]. Sox2 is a member of the Sry-related HMG box family of transcription factors that interact with DNA through binding to the minor groove. Sox2 shares many of the same DNA targets as Oct4 [32,33], with many of the char (...truncated)