Systems-level dynamic analyses of fate change in murine embryonic stem cells. Nature

Department of Molecular Biology, Princeton University, Princeton, New Jersey 08544, USA.
Nature (Impact Factor: 42.35). 11/2009; 462(7271):358-62. DOI: 10.1038/nature08575
Source: PubMed

ABSTRACT Molecular regulation of embryonic stem cell (ESC) fate involves a coordinated interaction between epigenetic, transcriptional and translational mechanisms. It is unclear how these different molecular regulatory mechanisms interact to regulate changes in stem cell fate. Here we present a dynamic systems-level study of cell fate change in murine ESCs following a well-defined perturbation. Global changes in histone acetylation, chromatin-bound RNA polymerase II, messenger RNA (mRNA), and nuclear protein levels were measured over 5 days after downregulation of Nanog, a key pluripotency regulator. Our data demonstrate how a single genetic perturbation leads to progressive widespread changes in several molecular regulatory layers, and provide a dynamic view of information flow in the epigenome, transcriptome and proteome. We observe that a large proportion of changes in nuclear protein levels are not accompanied by concordant changes in the expression of corresponding mRNAs, indicating important roles for translational and post-translational regulation of ESC fate. Gene-ontology analysis across different molecular layers indicates that although chromatin reconfiguration is important for altering cell fate, it is preceded by transcription-factor-mediated regulatory events. The temporal order of gene expression alterations shows the order of the regulatory network reconfiguration and offers further insight into the gene regulatory network. Our studies extend the conventional systems biology approach to include many molecular species, regulatory layers and temporal series, and underscore the complexity of the multilayer regulatory mechanisms responsible for changes in protein expression that determine stem cell fate.

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    • "While the transcriptional, signaling, and epigenetic regulation of these cells have been the primary focus of research efforts in recent years (reviewed in Ng and Surani, 2011; Young, 2011; Watanabe et al., 2013), posttranscriptional and translational mechanisms of control remain relatively unexplored, despite evidence that they play a dominant role in driving ESC fate decisions . Indeed, posttranscriptional regulation has been reported to account for nearly 75% of the changes in protein levels after differentiation induced by knockdown of the transcription factor Nanog (Lu et al., 2009), and it was recently demonstrated that control over translational initiation by the eIF4e binding proteins dramatically influences the efficiency of reprogramming somatic cells to induced pluripotent stem cells (iPSCs) (Tahmasebi et al., 2014). The cell controls protein levels posttranscriptionally using a large collection of tools that includes noncoding RNAs and RNA binding proteins (RBPs). "
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    ABSTRACT: Establishment, maintenance, and exit from pluripotency require precise coordination of a cell’s molecular machinery. Substantial headway has been made in deciphering many aspects of this elaborate system, particularly with respect to epigenetics, transcription, and noncoding RNAs. Less attention has been paid to posttranscriptional regulatory processes such as alternative splicing, RNA processing and modification, nuclear export, regulation of transcript stability, and translation. Here, we introduce the RNA binding proteins that enable the posttranscriptional regulation of gene expression, summarizing current and ongoing research on their roles at different regulatory points and discussing how they help script the fate of pluripotent stem cells.
    Cell Stem Cell 09/2014; 15(3):271–280. DOI:10.1016/j.stem.2014.08.010 · 22.15 Impact Factor
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    • "This finding is reminiscent of a transcriptionally poised state of the developmental genes in ESCs, which repress the transcription of differentiationinducing genes (Bernstein et al., 2006). The genome-wide crosstalk between transcription and translation during reprogramming and differentiation remains to be examined (Lu et al., 2009). Our results also reveal a functional interplay between 4E-BPs and the p53-p21 pathway, which affects the synchronization of the transcriptional and translational programs and is essential for reprogramming (Figure 5F). "
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    ABSTRACT: Translational control plays a pivotal role in the regulation of the pluripotency network in embryonic stem cells, but its effect on reprogramming somatic cells to pluripotency has not been explored. Here, we show that eukaryotic translation initiation factor 4E (eIF4E) binding proteins (4E-BPs), which are translational repressors, have a multifaceted effect on the reprogramming of mouse embryonic fibroblasts (MEFs) into induced pluripotent stem cells (iPSCs). Loss of 4E-BP expression attenuates the induction of iPSCs at least in part through increased translation of p21, a known inhibitor of somatic cell reprogramming. However, MEFs lacking both p53 and 4E-BPs show greatly enhanced reprogramming resulting from a combination of reduced p21 transcription and enhanced translation of endogenous mRNAs such as Sox2 and Myc and can be reprogrammed through the expression of only exogenous Oct4. Thus, 4E-BPs exert both positive and negative effects on reprogramming, highlighting the key role that translational control plays in regulating this process.
    Cell stem cell 03/2014; 14(5). DOI:10.1016/j.stem.2014.02.005 · 22.15 Impact Factor
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    • "In addition to the transcriptional network, recent studies have begun to reveal an important role of posttranscriptional regulation in the maintenance of ESC pluripotency and self-renewal. For example, alternative splicing and posttranslational modification of key pluripotency factors have been shown to modulate their function in ESCs (Buckley et al., 2012; Han et al., 2013; Lu et al., 2009). In addition, noncoding RNAs control ESC fate by regulating pluripotency gene mRNA decay and translation (Guttman et al., 2011; Melton et al., 2010; Tay et al., 2008; Wang et al., 2008). "
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    ABSTRACT: Embryonic stem cell (ESC) self-renewal and differentiation are governed by a broad-ranging regulatory network. Although the transcriptional regulatory mechanisms involved have been investigated extensively, posttranscriptional regulation is still poorly understood. Here we describe a critical role of the THO complex in ESC self-renewal and differentiation. We show that THO preferentially interacts with pluripotency gene transcripts through Thoc5 and is required for self-renewal at least in part by regulating their export and expression. During differentiation, THO loses its interaction with those transcripts due to reduced Thoc5 expression, leading to decreased expression of pluripotency proteins that facilitates exit from self-renewal. THO is also important for the establishment of pluripotency, because its depletion inhibits somatic cell reprogramming and blastocyst development. Together, our data indicate that THO regulates pluripotency gene mRNA export to control ESC self-renewal and differentiation, and therefore uncover a role for this aspect of posttranscriptional regulation in stem cell fate specification.
    Cell stem cell 12/2013; 13(6):676-90. DOI:10.1016/j.stem.2013.10.008 · 22.15 Impact Factor
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