Article

Early-stage epigenetic modification during somatic cell reprogramming by Parp1 and Tet2

Department of Pathology, Taub Institute for Aging, Columbia University, New York, New York 10032, USA.
Nature (Impact Factor: 42.35). 08/2012; 488(7413):652-5. DOI: 10.1038/nature11333
Source: PubMed

ABSTRACT Somatic cells can be reprogrammed into induced pluripotent stem cells (iPSCs) by using the pluripotency factors Oct4, Sox2, Klf4 and c-Myc (together referred to as OSKM). iPSC reprogramming erases somatic epigenetic signatures—as typified by DNA methylation or histone modification at silent pluripotency loci—and establishes alternative epigenetic marks of embryonic stem cells (ESCs). Here we describe an early and essential stage of somatic cell reprogramming, preceding the induction of transcription at endogenous pluripotency loci such as Nanog and Esrrb. By day 4 after transduction with OSKM, two epigenetic modification factors necessary for iPSC generation, namely poly(ADP-ribose) polymerase-1 (Parp1) and ten-eleven translocation-2 (Tet2), are recruited to the Nanog and Esrrb loci. These epigenetic modification factors seem to have complementary roles in the establishment of early epigenetic marks during somatic cell reprogramming: Parp1 functions in the regulation of 5-methylcytosine (5mC) modification, whereas Tet2 is essential for the early generation of 5-hydroxymethylcytosine (5hmC) by the oxidation of 5mC (refs 3,4). Although 5hmC has been proposed to serve primarily as an intermediate in 5mC demethylation to cytosine in certain contexts, our data, and also studies of Tet2-mutant human tumour cells, argue in favour of a role for 5hmC as an epigenetic mark distinct from 5mC. Consistent with this, Parp1 and Tet2 are each needed for the early establishment of histone modifications that typify an activated chromatin state at pluripotency loci, whereas Parp1 induction further promotes accessibility to the Oct4 reprogramming factor. These findings suggest that Parp1 and Tet2 contribute to an epigenetic program that directs subsequent transcriptional induction at pluripotency loci during somatic cell reprogramming.

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    • "PARP1, localized primarily to the nucleus, is the most abundant family member in humans (Vyas et al., 2013; Wang et al., 2012) and has been mainly examined in the context of base excision repair (Sousa et al., 2012). Recently PARP1 was implicated in other DNA repair pathways as well as in pathways outside of DNA repair such as transcription (Ji and Tulin, 2013; Krishnakumar and Kraus, 2010b) and stem cell identity (Chiou et al., 2013; Doege et al., 2012; Ogino et al., 2007). The details of its involvement in any of these pathways remain poorly understood. "
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    ABSTRACT: Poly(ADP-ribose) polymerases (PARPs) catalyze poly(ADP-ribose) addition onto proteins, an important posttranslational modification involved in transcription, DNA damage repair, and stem cell identity. Previous studies established the activation of PARP1 in response to DNA damage, but little is known about PARP1 regulation outside of DNA repair. We developed an assay for measuring PARP activity in cell lysates and found that the basal activity of PARP1 was highly variable across breast cancer cell lines, independent of DNA damage. Sucrose gradient fractionation demonstrated that PARP1 existed in at least three biochemically distinct states in both high- and low-activity lines. A discovered complex containing the NuA4 chromatin-remodeling complex and PARP1 was responsible for high basal PARP1 activity, and NuA4 subunits were required for this activity. These findings present a pathway for PARP1 activation and a direct link between PARP1 and chromatin remodeling outside of the DNA damage response.
    Cell Reports 09/2014; 8(6). DOI:10.1016/j.celrep.2014.08.009 · 7.21 Impact Factor
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    • "In both experimental and natural reprogramming models, DNA demethylation has been intimately linked to the activation of pluripotency loci. In particular, cell fusion and iPSC induction experiments have implicated Tet-mediated hydroxylation in the epigenetic reactivation of silent pluripotency genes, a perceived bottleneck in the path toward the establishment of pluripotency (Doege et al., 2012; Piccolo et al., 2013). Oocyte Tet3 provides a reprogramming activity for pluripotency gene reactivation during the early embryonic development after nuclear transfer and natural fertilization (Gu et al., 2011). "
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    ABSTRACT: Tet-mediated DNA oxidation is a recently identified mammalian epigenetic modification, and its functional role in cell-fate transitions remains poorly understood. Here, we derive mouse embryonic fibroblasts (MEFs) deleted in all three Tet genes and examine their capacity for reprogramming into induced pluripotent stem cells (iPSCs). We show that Tet-deficient MEFs cannot be reprogrammed because of a block in the mesenchymal-to-epithelial transition (MET) step. Reprogramming of MEFs deficient in TDG is similarly impaired. The block in reprogramming is caused at least in part by defective activation of key miRNAs, which depends on oxidative demethylation promoted by Tet and TDG. Reintroduction of either the affected miRNAs or catalytically active Tet and TDG restores reprogramming in the knockout MEFs. Thus, oxidative demethylation to promote gene activation appears to be functionally required for reprogramming of fibroblasts to pluripotency. These findings provide mechanistic insight into the role of epigenetic barriers in cell-lineage conversion.
    Cell Stem Cell 04/2014; 14(4):512-522. DOI:10.1016/j.stem.2014.01.001 · 22.15 Impact Factor
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    • "Somatic cells can be reprogrammed into induced pluripotent stem cells through the overexpression of 'stemness' genes OCT4, SOX2, KLF4 and c-MYC (Takahashi and Yamanaka, 2006). The overexpression of these four genes is known to lead to the DNA demethylation of other downstream stemness genes, such as NANOG and ESRRB (Doege et al., 2012; Hajkova et al., 2008; Kafri et al., 1992). In nondividing heterokaryons consisting of fused mouse embryonic stem cells with human fibroblasts rapid DNA demethylation occur at the OCT4 and NANOG promoters, accompanied by their transcriptional induction (Bhutani et al., 2010). "
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    ABSTRACT: Over the last several years proteins involved in base excision repair (BER) have been implicated in active DNA demethylation. We review the literature supporting BER as a means of active DNA demethylation, and explain how the various components function and cooperate to remove the potentially most enduring means of epigenetic gene regulation. Recent evidence indicates that the same pathways implicated during periods of widespread DNA demethylation, such as the erasure of methyl marks in the paternal pronucleus soon after fertilization, are operational in post-mitotic neurons. Neuronal functional identities, defined here as the result of a combination of neuronal subtype, location, and synaptic connections are largely maintained through DNA methylation. Chronic mental illnesses, such as schizophrenia, may be the result of both altered neurotransmitter levels and neurons that have assumed dysfunctional neuronal identities. A limitation of most current psychopharmacological agents is their focus on the former, while not addressing the more profound latter pathophysiological process. Previously, it was believed that active DNA demethylation in post-mitotic neurons was rare if not impossible. If this were the case, then reversing the factors that maintain neuronal identity, would be highly unlikely. The emergence of an active DNA demethylation pathway in the brain is a reason for great optimism in psychiatry as it provides a means by which previously pathological neurons may be reprogrammed into a more favorable role. Agents targeting epigenetic processes have shown much promise in this regard, and may lead to substantial gains over traditional pharmacological approaches.
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