Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transition of embryonic stem cells from ground state pluripotency (RNA-seq)

ABSTRACT: Mouse embryonic stem cells were maintained in minimal and chemically-defined culture conditions supporting naive pluripotency. Inhibitors of the Gsk3 (CHIR99021) and Mek/Erk (PD0325901) pathways were withdrawn, cultures maintained for 24 hours, and subsequently sorted by flow cytometry based on fluorescence of a short-half-life Rex1(Zfp42)::GFP reporter into two populations: Rex1-high cells, functionally capable of reversion to naive pluripotency, and Rex1-low cells that have exited the naive state.

INSTRUMENT(S): Illumina Genome Analyzer IIx

ORGANISM(S): Mus musculus

SUBMITTER: Paul Bertone

PROVIDER: E-MTAB-5305 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Tracking the embryonic stem cell transition from ground state pluripotency.

Kalkan Tüzer T Olova Nelly N Roode Mila M Mulas Carla C Lee Heather J HJ Nett Isabelle I Marks Hendrik H Walker Rachael R Stunnenberg Hendrik G HG Lilley Kathryn S KS Nichols Jennifer J Reik Wolf W Bertone Paul P Smith Austin A

Development (Cambridge, England) 20170207 7

Mouse embryonic stem (ES) cells are locked into self-renewal by shielding from inductive cues. Release from this ground state in minimal conditions offers a system for delineating developmental progression from naïve pluripotency. Here, we examine the initial transition process. The ES cell population behaves asynchronously. We therefore exploited a short-half-life <i>Rex1::GFP</i> reporter to isolate cells either side of exit from naïve status. Extinction of ES cell identity in single cells is ...[more]

PMID: 28174249

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Project description:Nanog, a core pluripotency factor in the inner cell mass of blastocysts, is also expressed in unipotent primordial germ cells (PGC) in mice1, where its precise role is yet unclear2-4. We investigated this in an in vitro model, where naÃ¯ve pluripotent embryonic stem cells (ESCs) cultured in bFGF/ActivinA develop as epiblast-like cells (EpiLCs), and gain competence for PGC-like fate5. Consequently, bone morphogenetic protein (BMP4), or ectopic expression of key germline transcription factors Prdm1/ Prdm14/ Tfap2c, directly induce PGC-like cells (PGCLCs) in EpiLCs, but not in ESCs6-8. Here we report an unexpected discovery that Nanog alone can induce PGCLCs in EpiLCs, independently of BMP4. We propose that following the dissolution of the naÃ¯ve ESC pluripotency network during establishment of EpiLCs9,10, the epigenome is reset for cell fate determination. Indeed, we found genome-wide changes in NANOG binding pattern between ESCs and EpiLCs, indicating epigenetic resetting of regulatory elements. Accordingly, we show that NANOG can bind and activate enhancers of Prdm1 and Prdm14 in EpiLCs in vitro; BLIMP1 (encoded by Prdm1) then directly induces Tfap2c. Furthermore, while SOX2 and NANOG promote the pluripotent state in ESCs, they show contrasting roles in EpiLCs since Sox2 specifically represses PGCLC induction by Nanog. This study demonstrates a broadly applicable mechanistic principle for how cells acquire competence for cell fate determination, resulting in the context-dependent roles of key transcription factors during development. To characterize Nanog-induced Primordial Germ cell-like cells (PGCLCs), we performed expression analysis of Nanog-induced Day4 PGCLCs compared to male mouse Embryonic Stem Cells (mESCs) and male Day4 PGCLCs which were induced by cytokines. mESCs were maintained in N2B27 2i(CHIR99021 3 ÂµM, PD0325901 1 ÂµM) LIF(1000 U/ml) medium and Day4 PGCLCs were induced in GK15 medium with Nanog induction (0.7 Âµg/ml) or cytokines (BMP4 500 ng/ml, BMP8A 500 ng/ml, SCF 100 ng/ml, EGF 50 ng/ml and LIF 1000U/ml) as previously reported (Hayashi K et al., Cell, 2011).

Dataset Information

Transition of embryonic stem cells from ground state pluripotency (RNA-seq)

Publications

Tracking the embryonic stem cell transition from ground state pluripotency.

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