Comparative Principles of DNA Methylation Reprogramming during Human and Mouse In Vitro Primordial Germ Cell Specification.

Primordial germ cell (PGC) development is characterized by global epigenetic remodeling, which resets genomic potential and establishes an epigenetic ground state. Here we recapitulate PGC specification in vitro from naive embryonic stem cells and characterize the early events of epigenetic reprogramming during the formation of the human and mouse germline. Following rapid de novo DNA methylation during priming to epiblast-like cells, methylation is globally erased in PGC-like cells.

Distinct Signaling Requirements for the Establishment of ESC Pluripotency in Late-Stage EpiSCs.

It has previously been reported that mouse epiblast stem cell (EpiSC) lines comprise heterogeneous cell populations that are functionally equivalent to cells of either early- or late-stage postimplantation development. So far, the establishment of the embryonic stem cell (ESC) pluripotency gene regulatory network through the widely known chemical inhibition of MEK and GSK3beta has been impractical in late-stage EpiSCs. Here, we show that chemical inhibition of casein kinase 1alpha (CK1alpha) induces the conversion of recalcitrant late-stage EpiSCs into ESC pluripotency.

Distinct Wnt-driven primitive streak-like populations reflect in vivo lineage precursors.

During gastrulation, epiblast cells are pluripotent and their fate is thought to be constrained principally by their position. Cell fate is progressively restricted by localised signalling cues from areas including the primitive streak. However, it is unknown whether this restriction accompanies, at the individual cell level, a reduction in potency.

The role of pluripotency gene regulatory network components in mediating transitions between pluripotent cell states.

Pluripotency is a property that early embryonic cells possess over a considerable developmental time span. Accordingly, pluripotent cell lines can be established from the pre-implantation or post-implantation mouse embryo as embryonic stem (ES) or epiblast stem (EpiSC) cell lines, respectively. Maintenance of the pluripotent phenotype depends on the function of specific transcription factors (TFs) operating within a pluripotency gene regulatory network (PGRN).

Hes1 desynchronizes differentiation of pluripotent cells by modulating STAT3 activity.

Robust development of the early embryo may benefit from mechanisms that ensure that not all pluripotent cells differentiate at exactly the same time: such mechanisms would build flexibility into the process of lineage allocation. This idea is supported by the observation that pluripotent stem cells differentiate at different rates in vitro. We use a clonal commitment assay to confirm that pluripotent cells commit to differentiate asynchronously even under uniform differentiation conditions.

Reduced Oct4 expression directs a robust pluripotent state with distinct signaling activity and increased enhancer occupancy by Oct4 and Nanog.

Embryonic stem cell (ESC) pluripotency is governed by a gene regulatory network centered on the transcription factors Oct4 and Nanog. To date, robust self-renewing ESC states have only been obtained through the chemical inhibition of signaling pathways or enforced transgene expression. Here, we show that ESCs with reduced Oct4 expression resulting from heterozygosity also exhibit a stabilized pluripotent state.

In Vivo differentiation potential of epiblast stem cells revealed by chimeric embryo formation.

Chimera formation after blastocyst injection or morula aggregation is the principal functional assay of the developmental potential of mouse embryonic stem cells (ESCs). This property, which demonstrates functional equivalence between ESCs and the preimplantation epiblast, is not shared by epiblast stem cell (EpiSC) lines. Here, we show that EpiSCs derived either from postimplantation embryos or from ESCs in vitro readily generate chimeras when grafted to postimplantation embryos in whole embryo culture.

OCT4/SOX2-independent Nanog autorepression modulates heterogeneous Nanog gene expression in mouse ES cells.

NANOG, OCT4 and SOX2 form the core network of transcription factors supporting embryonic stem (ES) cell self-renewal. While OCT4 and SOX2 expression is relatively uniform, ES cells fluctuate between states of high NANOG expression possessing high self-renewal efficiency, and low NANOG expression exhibiting increased differentiation propensity. NANOG, OCT4 and SOX2 are currently considered to activate transcription of each of the three genes, an architecture that cannot readily account for NANOG heterogeneity.

Esrrb is a direct Nanog target gene that can substitute for Nanog function in pluripotent cells.

Embryonic stem cell (ESC) self-renewal efficiency is determined by the level of Nanog expression. However, the mechanisms by which Nanog functions remain unclear, and in particular, direct Nanog target genes are uncharacterized. Here we investigate ESCs expressing different Nanog levels and Nanog(-/-) cells with distinct functionally inducible Nanog proteins to identify Nanog-responsive genes.

The developmental dismantling of pluripotency is reversed by ectopic Oct4 expression.

The transcription factors Nanog and Oct4 regulate pluripotency in the pre-implantation epiblast and in derivative embryonic stem cells. During post-implantation development, the precise timing and mechanism of the loss of pluripotency is unknown. Here, we show that in the mouse, pluripotency is extinguished at the onset of somitogenesis, coincident with reduced expression and chromatin accessibility of Oct4 and Nanog regulatory regions.

Transcription factor heterogeneity and epiblast pluripotency.

Stem cells are defined by the simultaneous possession of the seemingly incongruent properties of self-renewal 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.