Oct4 dependence of chromatin structure within the extended Nanog locus in ES cells

Division of Hematology-Oncology, Children's Hospital and Dana Farber Cancer Institute, Harvard Medical School, Harvard Stem Cell Institute, Boston, MA 02115, USA.
Genes & Development (Impact Factor: 10.8). 04/2008; 22(5):575-80. DOI: 10.1101/gad.1606308
Source: PubMed


Embryonic stem (ES) cells offer insight into early developmental fate decisions, and their controlled differentiation may yield vast regenerative potential. The molecular determinants supporting ES cell self-renewal are incompletely understood. The homeodomain proteins Nanog and Oct4 are essential for mouse ES cell self-renewal. Using a high-throughput approach, we discovered DNaseI hypersensitive sites and potential regulatory elements along a 160-kb region of the genome that includes GDF3, Dppa3, and Nanog. We analyzed gene expression, chromatin occupancy, and higher-order chromatin structure throughout this gene locus and found that expression of the reprogramming factor Oct4 is required to maintain its integrity.

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    • "were previously demonstrated to be engaged in enhancer-promoter contacts that are lost during differentiation [31] [130]. Although this illustrates that the extent to which key pluripotency genes and super-enhancers adopt tissue-invariant or tissue-specific chromatin configurations remains to be elucidated in more detail, the latter stories clearly demonstrated the existence of PSC-specific regulatory chromatin loops that are often dependent on the binding of PSC-specific transcription factors. "
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    ABSTRACT: Pluripotent stem cells (PSCs) have the ability to self-renew and are capable of generating all embryonic germ layers (Evans and Kaufman, 1981; Thomson et al., 1998). PSCs can be isolated from early embryos or may be induced via overexpression of pluripotency transcription factors in differentiated cells (Takahashi and Yamanaka, 2006). As PSCs hold great promise for regenerative medicine, the mechanisms underlying pluripotency and induction thereof are studied intensively. Pluripotency is characterized by a unique transcriptional program that is in part controlled by an exceptionally plastic regulatory chromatin landscape. In recent years, 3D genome configuration emerged as an important regulator of transcriptional control and cellular identity (Taddei et al., 2004 [4]; Lanctot et al., 2007 [5]; Gibcus and Dekker, 2013; Misteli, 2009 [7]). Here we provide an overview of recent findings on the 3D genome organization in PSCs and discuss its putative functional role in regulation of the pluripotent state. Copyright © 2015. Published by Elsevier B.V.
    Full-text · Article · May 2015 · FEBS letters
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    • "Moreover, non-histone mechanisms in ESCs may play additional roles in chromatin structural maintenance, thereby allowing the redeployment of H1 linker histone proteins to other later developmental functions. One mechanism, in particular, encompasses the binding of Oct4, Nanog and other pluripotency factors to DNase I hypersensitivity sites (HS sites) within the Nanog locus, resulting in chromatin restructuring and modulation of Nanog-associated genes linked to pluripotency and early development, including Dpp3 and GDF3 [35], [37]. "
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    ABSTRACT: H1 linker histone proteins are essential for the structural and functional integrity of chromatin and for the fidelity of additional epigenetic modifications. Deletion of H1c, H1d and H1e in mice leads to embryonic lethality by mid-gestation with a broad spectrum of developmental alterations. To elucidate the cellular and molecular mechanisms underlying H1 linker histone developmental functions, we analyzed embryonic stem cells (ESCs) depleted of H1c, H1d and H1e subtypes (H1-KO ESCs) by utilizing established ESC differentiation paradigms. Our study revealed that although H1-KO ESCs continued to express core pluripotency genes and the embryonic stem cell markers, alkaline phosphatase and SSEA1, they exhibited enhanced cell death during embryoid body formation and during specification of mesendoderm and neuroectoderm. In addition, we demonstrated deregulation in the developmental programs of cardiomyocyte, hepatic and pancreatic lineage elaboration. Moreover, ectopic neurogenesis and cardiomyogenesis occurred during endoderm-derived pancreatic but not hepatic differentiation. Furthermore, neural differentiation paradigms revealed selective impairments in the specification and maturation of glutamatergic and dopaminergic neurons with accelerated maturation of glial lineages. These impairments were associated with deregulation in the expression profiles of pro-neural genes in dorsal and ventral forebrain-derived neural stem cell species. Taken together, these experimental observations suggest that H1 linker histone proteins are critical for the specification, maturation and fidelity of organ-specific cellular lineages derived from the three cardinal germ layers.
    Full-text · Article · May 2014 · PLoS ONE
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    • "Similarly, binding sites in differentiated cells were considered either constitutive or gained depending on whether they could be matched to a site in undifferentiated cells (7102 and 1914 sites, respectively). Examples of sites where CTCF binding is lost, gained or constitutive, some of which have been previously characterized, are shown in Figure 1A–C (39,40). As expected, LowOc sites comprised a larger proportion of CBSs where binding was lost or gained as compared with sites where binding was constitutive (Figure 1D). "
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    ABSTRACT: CTCF (CCCTC-binding factor) is a highly conserved multifunctional DNA-binding protein with thousands of binding sites genome-wide. Our previous work suggested that differences in CTCF's binding site sequence may affect the regulation of CTCF recruitment and its function. To investigate this possibility, we characterized changes in genome-wide CTCF binding and gene expression during differentiation of mouse embryonic stem cells. After separating CTCF sites into three classes (LowOc, MedOc and HighOc) based on similarity to the consensus motif, we found that developmentally regulated CTCF binding occurs preferentially at LowOc sites, which have lower similarity to the consensus. By measuring the affinity of CTCF for selected sites, we show that sites lost during differentiation are enriched in motifs associated with weaker CTCF binding in vitro. Specifically, enrichment for T at the 18(th) position of the CTCF binding site is associated with regulated binding in the LowOc class and can predictably reduce CTCF affinity for binding sites. Finally, by comparing changes in CTCF binding with changes in gene expression during differentiation, we show that LowOc and HighOc sites are associated with distinct regulatory functions. Our results suggest that the regulatory control of CTCF is dependent in part on specific motifs within its binding site.
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