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: 41.46). 08/2012; 488(7413):652-5. DOI: 10.1038/nature11333
Source: PubMed


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 · 8.36 Impact Factor
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    • "The recent discovery of cytosine hydroxymethylation raises the possibility that methylation at CGIs can be reversed via active cytosine demethylation involving TET proteins leading to the reactivation of transcriptionally silenced promoters [9, 10]. Furthermore new evidence on induced pluripotent stem cells (iPSCs) suggests that the 5hmC base modification not only is an intermediate within cytosine demethylation, but acts as an epigenetic mark in itself, possibly aiding in recruitment of factors that promote chromatin remodeling [25, 26]. "
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    ABSTRACT: TET2 is a methylcytosine dioxygenase that is frequently mutated in myeloid malignancies, notably myelodysplasia and acute myeloid leukemia. TET2 catalyses the conversion of 5'-methylcytosine to 5'-hydroxymethylcytosine within DNA and has been implicated in the process of genomic demethylation. However, the mechanism by which TET2 loss of function results in hematopoietic dysplasia and leukemogenesis is poorly understood. Here, we show that TET2 is expressed in undifferentiated embryonic stem cells and that its knockdown results in reduction of 5'-hydroxymethylcytosine in genomic DNA. We also present DNA methylation data from bone marrow samples obtained from patients with TET2-mutated myelodysplasia. Based on these findings, we sought to identify the role of TET2 in regulating pluripotency and differentiation. We show that overexpression of TET2 in a stably integrated transgene leads to increased alkaline phosphatase expression in differentiating ES cells and impaired differentiation in methylcellulose culture. We speculate that this effect is due to TET2-mediated expression of stem cell genes in ES cells via hydroxylation of 5'-methylcytosines at key promoter sequences within genomic DNA. This leads to relative hypomethylation of gene promoters as 5'-hydroxymethylcytosine is not a substrate for DNMT1-mediated maintenance methylation. We sought to test this hypothesis by cotransfecting the TET2 gene with methylated reporter genes. The results of these experiments are presented.
    08/2014; 2014:986571. DOI:10.1155/2014/986571
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    • "Initially, different cocktails of transcription factors were described for pluripotency induction [2], [3], [4]. Subsequently, certain epigenetic regulators capable of altering chromatin state through histone modifications or DNA methylation were found to participate in somatic cell reprogramming [5], [6]. More recently, several studies have shown that mouse fibroblasts can be reprogrammed into iPSCs using nuclear factors that control lineage specification [7], [8]. "
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    ABSTRACT: Induction of pluripotent stem cells (iPSC) by defined transcription factors is the recognized canonical means for somatic reprogramming, however, it remains incompletely understood how individual transcription factors affect cell fate decisions during the reprogramming process. Here, we report induction of fibroblast reprogramming by various transcriptional factors is mediated by a miR19a/b-PTEN axis. cMyc, one of the four Yamanaka factors known to stimulate both somatic cell reprogramming and tumorigenesis, induced the expression of multiple mircoRNAs, miR-17∼92 cluster in particular, in the early stage of reprogramming of human fibroblasts. Importantly, miR-17∼92 cluster could greatly enhance human fibroblast reprogramming induced by either the four Yamanaka factors (Oct4, Sox2, Klf4, and cMyc, or 4F) or the first three transcriptional factors (Oct4, Sox2, and Klf4, or 3F). Among members of this microRNA cluster, miR-19a/b exhibited the most potent effect on stimulating fibroblst reprogramming to iPSCs. Additional studies revealed that miR-19a/b enhanced iPSC induction efficiency by targeted inhibition of phosphatase and tensin homolog (PTEN), a renowned tumor suppressor whose loss-of-function mutations were found in multiple human malignancies. Our results thus demonstrate an important role of miR-19a/b-PTEN axis in the reprogramming of human fibroblasts, illustrating that the somatic reprogramming process and its underlying regulation pathways are intertwined with oncogenic signaling in human malignancies.
    PLoS ONE 04/2014; 9(4):e95213. DOI:10.1371/journal.pone.0095213 · 3.23 Impact Factor
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