- Email: [email protected]
Chromatin modulators as facilitating factors in cellular reprogramming
Available online at www.sciencedirect.com
ScienceDirect Chromatin modulators as facilitating factors in cellular reprogramming Luis Luna-Zurita1 and Benoit G Bruneau1,2 In the last few years, cellular reprogramming has emerged as a means to alter cellular identity and generate diverse cell types for disease modeling, drug testing, and potential therapeutic use. Since each cell type is a result of a specific gene expression profile finely regulated by the activity of a repertoire of transcription factors (TFs), reprogramming approaches have, thus far, been relatively inefficient and based largely on the forced expression of selective cell type-specific TFs. TFs function within the confines of chromatin, and the chromatin states can in turn be modulated by TF activity. Therefore, the knowledge of how chromatin remodeling factors alter chromatin structure, control TF activity and gene expression has led to an improved reprogramming efficiency and extended the number of cellular types that can be generated by cellular reprogramming. Here we review recent insights into the role and mechanisms by which chromatin remodeling, histone modifications, and DNA methylation contribute to cellular differentiation and reprogramming. Addresses 1 Gladstone Institute of Cardiovascular Disease, San Francisco, CA 94158, USA 2 Department of Pediatrics, Cardiovascular Research Institute, University of California, San Francisco, CA 94158, USA Corresponding author: Bruneau, Benoit G ([email protected])
Current Opinion in Genetics & Development 2013, 23:556–561 This review comes from a themed issue on Cell reprogramming Edited by Huck Hui Ng and Patrick Tam For a complete overview see the Issue and the Editorial Available online 28th August 2013 0959-437X/$ – see front matter, # 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.gde.2013.07.002
dynamic, allowing context-dependent changes of gene expression [1]. Once established, chromatin structure remains relatively stable for a specific differentiated stage [3], and it was assumed that the genomic structural changes during cellular differentiation were irreversible. Studies involving the transfer of somatic cell nuclei to enucleated oocytes [4,5] in Xenopus and similar experiments later in mammals [6], including studies with postmitotic cells as donor cells [7], showed that oocytes provide a specific environment that can reverse the chromatin architecture associated with the differentiated stage. Similarly, heterokaryon fusion experiments showed that the nucleus from one cell type could be efficiently reprogrammed to express the transcriptome of a different somatic cell type [8]. Although these studies did not identify the nuclear factors that were in play, other reports [9,10] in which certain cells were induced into skeletal muscle by introducing the bHLH transcription factor MyoD, showed that a single defined factor could reprogram cellular identity. The ability of TFs to reprogram somatic cells to a pluripotent state was first demonstrated by pioneering work from Shinya Yamanaka’s laboratory [11,12]. They showed that a handful of defined factors was sufficient to completely reset the somatic nucleus’s differentiation program. Since then, similar approaches with tissuespecific TFs have been used to reprogram cells directly to other somatic cell types. Although there are still a lot of questions about how the reprogramming TFs effect these lineage conversions, due to multiple studies aimed at identifying modulators of this process, the role of chromatin-level regulation in facilitating cell reprogramming and TFs activity is becoming better understood.
Introduction
Histone modifications
TFs are the main effectors of cell differentiation and govern gene expression changes that will give rise to the diverse cellular types that emerge during development. While TFs regulate gene expression, TF–DNA interactions are influenced by the dynamic structural and biochemical properties of chromatin. Chromatin states can be regulated by different entities or processes that modify chromatin biochemical properties, chromatin conformation or nucleosome distribution. They include histone and DNA modifications, nucleosome positioning, exchange of histone variants, and chromatin remodeler proteins [1,2]. These chromatin modifications are
Chromatin can transition between ‘open’ (euchromatin) and ‘closed’ (heterochromatin) states. Posttranslational histone modifications, including methylation, acetylation and ubiquitination, determine the changes in status. These dynamic marks promote structural changes in chromatin conformation and are essential in the control of gene expression.
Current Opinion in Genetics & Development 2013, 23:556–561
These same modifications modulate the efficiency of induced pluripotent stem (iPS) cell reprogramming [13,14]. For example, small molecules have been found to alter histone acetylation (valproic acid) or methylation www.sciencedirect.com
Chromatin regulation of reprogramming Luna-Zurita and Bruneau 557
(BIX-01294). Members of the Polycomb group (PcG) and Trithorax group (TrxG) are key in controlling gene expression during cell differentiation and reprogramming. PcG and TrxG are protein complexes with opposing roles in cell differentiation. While PcG activity is associated with gene silencing and heterochromatin stabilization, TrxG maintains active transcriptional states (reviewed in [2,15,16]). In mammals, Polycomb repressor complex 2 (PRC2) activity depends on three subunits: Ezh2, Eed and Suz12. Although Ezh2 is the enzymatic subunit of PRC2, Suz12 and Eed are also crucial to methylate (monomethylation, dimethylation and trimethylation) lysine 27 on histone 3 (H3K27) [17]. In contrast to H3K27me1 and H3K27me2, which are broadly distributed across the genome, H3K27me3 is the mark most associated with the repressive function of this complex [17]. H3K27me3 acts as a docking site for the PcG complex PRC1 (the other main Polycomb repressor complex) [18] (and reviewed in [19]) and promotes the repressing mark H2AK119Ub. PRC1 and PRC2 colocalize on a large set of genes in ES cells, concomitant with an enrichment of the repressor mark H3K27me3 [20,21]. They also colocalize with a fraction of binding sites for essential factors for the pluripotency stage, such as Oct4, Nanog, Sox2 and Sall4 [22,23]. These results suggest that PRC1/2 acts through these TFs to maintain the pluripotency state. Interestingly, global loss of H3K27me3 after genetic deletion of Ezh2 does not significantly affect iPSC reprogramming [24]. This does not necessarily mean that PRC complex activity is not required for somatic reprogramming. Perhaps Ezh1 (the other known PRC2 enzymatic subunit) retains H3K27me3 at specific loci [24]. PRDM14, a PR domain-containing transcriptional regulator, interacts directly with PRC2 to establish H3K27me3 modifications in human ES cells. This observation supports the effect of PRDM14 repression to increase the efficiency of iPSC induction in human cells [25]. H3K27 demethylases, such as Utx or Jmjd3, have been implicated in modifying histones during cellular reprogramming independently of PcG/TrxG. Utx has specific roles during the re-establishment of pluripotency and in germ cell development. In conjunction with the pluripotency factors Oct4, Sox2 and Klf4, it removes the H3K27me3 repressive mark during the reprogramming of somatic cells [26] (Figure 1). Jmjd3 inhibits iPS cell reprogramming, partly via demethylase-dependent activity, but also via a demethylase-independent targeting of PHF20 for ubiquitination. PHF20 interacts with Wdr5 to regulate gene expression and is itself essential for iPS cell generation [27]. H3K9me3 is a constitutive heterochromatin mark in differentiated cells but this mark is not abundant in ES www.sciencedirect.com
cells or iPS cells [28]. On the basis of the effectiveness of inhibition of the H3K9me histone methyltransferase G9a in promoting iPS cell production, H3K9me3 has been proposed to be one of the barriers for iPS cell reprogramming [14]. Curiously, regions enriched in H3K9me3 overlap with ‘hot spots’ of aberrant epigenomic reprogramming [29,30]. Interestingly, the deficiency of Suv39h1, the enzyme catalyzing H3K9me3, also enhances reprogramming [31] (Figure 1). Thus, although H3K9me3 patterns are largely reset in iPS cells, these domains may present a barrier to reprogramming. The initial binding of pluripotency TFs Oct4, Klf4, and Sox2 to large genomic regions enriched in H3K9me3 loci is blocked, but this is likely to be overcome later in the reprogramming process and can be enhanced by loss of Suv39h1 and Suv39h2 [32]. Inhibition of another histone-modifying enzyme, Dot1L, also enhances cell reprogramming. Dot1L catalyzes the H3K79me2 modification, and this enhanced reprogramming might be due, at least in part, to the lack of this mark in genes that should be repressed during reprogramming [31].
Chromatin-remodeling complexes Nucleosome distribution can be altered at specific loci by chromatin remodeling complexes (CRCs), and this action provides a mechanism for gene regulation distinct from histone modifications. There are various families of CRCs, including ATP-dependent CRCs ISWI (imitation switch), CHD (chromodomain-helicase–DNA binding), INO80 (inositol requiring 80), and SWI/SNF (switching defective/sucrose nonfermenting) complexes [1]. The pivotal role of SWI/SNF complexes during cellular differentiation and reprogramming, where they act as proliferation repressors, is well described [33], and here we will focus mainly on this process. SWI/SNF complexes in vertebrates (Brm/Brg1-associated factor (BAF) complexes) function, even for the same gene, as both transcriptional activators and repressors at different developmental stages [1,34]. They comprise at least 12 protein subunits. The ATPase subunit of the BAF complex is encoded by one of the two highly homologous genes, Brg1 (Brahma-related gene 1) and Brm (Brahma). The subunit composition of BAF complexes also depends on the cellular context. For example, in ES cells, BAF complexes (known as esBAF) are defined by the presence of Brg1, BAF60a, and BAF155, and the absence of Brm, BAF60c, and BAF170. The esBAF complex directly interacts with key regulators of pluripotency, such as Oct4 and Sox2, and its activity is required to maintain the undifferentiated state of the ES cells [35] (Figure 1). BAF activity has been studied during the differentiation of several cellular types, such as Schwann cells [36] and Bcells, where differentiation is regulated by the balance between BAF and Mi-2/NuRD (CHD) activities [37,38]. In certain tissues, a BAF complex enrichment enhances Current Opinion in Genetics & Development 2013, 23:556–561
558 Cell reprogramming
Figure 1
DNMT BRG1 BAF60c
M
Baf155 Baf60a Brg1
PRDM14 PRC2
PRC1
Sox2 Oct4
TF
ES Cells
iPS Cells
Dedifferentiation Differentiation
G9a
BRG1 BAF155
Suv39h1
Utx
M
M
M
M
M
M Oct4
Differentiated Cell Tet1 Nanog Tet2
H3K9ac H3K4me3 H3K27me3 H3K9me3
H2AK119Ub
M
Cytosine Methylation
AA
Tet2
Parp1
Regulatory region of pluripotency genes Developmental genes Current Opinion in Genetics & Development
Cell reprogramming is regulated by changes in chromatin architecture and chemical modification of DNA. Activation histone marks (H3K4me3 and H3K9Ac) and hypomethylated pluripotency gene promoters allow a low chromatin-compaction status and high levels of gene expression. In contrast, repressive histone modifications (H3K27me3 and H3K9me3) and DNA methylation reduce RNA-PolII accessibility and gene expression. These modifications play a key role in maintaining the differentiated/undifferentiated state and represent strong barriers for cellular reprogramming. During ES/IPS cell differentiation, pluripotency gene promoters increase their methylation levels due to the activity of DNA methyltransferases (DNMTs). This modification prevents promoter interaction with TFs and inhibits gene expression. In contrast, TFs interactions required for cell differentiation are facilitated by different factors, such as the BAF-complex subunit Baf60c. The differentiation process requires the repression of factors implicated in the undifferentiated stage maintenance, such as the esBAF complex (characterized by the Brg1, BAF60a, and BAF155 subunits), that allows the interaction of pluripotency regulators, such as Sox2 and Oct4 to gene promoters, or repressive histone modification close to development promoters carried out by the PcG members PRC1/PRC2 or enzymes, such as PRDM14. During somatic cell reprogramming, the activity of histone demethylases, such as Utx, or BAF complexes characterized by the presence of BRG1 and BAF155 has been described as required for repressive histone marks removal and the acquisition of activation marks in regions where pluripotency genes are localized. Promoters of these genes must change their methylation status during this process. Thus, the activity of molecules, such as Tet1, Te2 or Parp1, reduces methylation levels, allowing the interaction of pluripotency factors, such as Nanog or Oct4. A similar demethylation effect can be achieved by adding ascorbic acid, which inhibits DNMTs activity and reduces methylation levels in gene promoters. Increased reprogramming efficiency can also be achieved by inhibiting molecules, such as the histone methyltranferases Suv39h1 or G9a, and removing the H3K9me3 repressive mark of plutipotency genes promoters.
Current Opinion in Genetics & Development 2013, 23:556–561
www.sciencedirect.com
Chromatin regulation of reprogramming Luna-Zurita and Bruneau 559
differentiation or reprogramming of cell type-specific programs. This is the case of Baf60c, a BAF complex subunit highly restricted to the myocardium and somites during mouse development [39]. During myocardial differentiation, Baf60c is essential for the function of DNA-binding transcription factors (Gata4, Tbx5), and Baf60c is part of a trio of factors sufficient to direct differentiation from mesoderm to myocardium. Baf60c facilitates the binding of Gata4 to its DNA targets and recruits the BAF complex to these loci [39–41]. Baf60c is also expressed in developing skeletal muscle and is essential for MyoD-dependent differentiation of human ES cells toward a skeletal muscle phenotype [42]. BAF complexes are also potent modulators of reprogramming to pluripotency. For the reprogramming of human fibroblasts into iPS cells, overexpression of BRG1 and the BAF155 subunit can replace c-Myc [43]. The mechanism by which BRG1 and BAF155 promote cell reprogramming is (presumably indirectly) by an increase of H3K4me3 and H3K9 acetylation (activation marks) on specific pluripotency-associated genes, and the reduction of the repressive mark H3K27me3 on OCT4 (Figure 1). Thus, BRG1 and BAF155 seem to promote a euchromatic state at the promoters of pluripotency genes, but importantly also induce OCT4 binding to the promoter of pluripotency-specific genes, including to its own promoter. BAF activity also promotes somatic cell reprogramming in fetal and adult liver progenitor cells (LPCs). LPCs are more efficiently reprogrammed than differentiated liver cells, when associated with endogenous expression of reprogramming factors (Klf4 and c-Myc) and BAF complex members Baf155 and Brg1, which mediate chromatin remodeling during reprogramming. Knockdown of BAF complex members negated the increased reprogramming efficiency of LPCs, compared with non-LPCs [44]. Altogether, these findings show that, for certain cellular programs, chromatin remodelers are not only permissive, but also in fact instructive.
DNA methylation and demethylation DNA methylation at cytosine residues is generally associated with silencing of gene expression. DNA methyltransferases (DNMTs) catalyze the covalent attachment of a methyl residue to the carbon 5 position of cytosine residues of CpG dinucleotides [2]. Several lines of evidence show the importance of DNA methylation in regulating gene expression during cell differentiation. Thus, while methylation of gene promoter regions is widely seen in differentiated cells, most of these genes are unmethylated in ES cells and sperm [45]. This is the case of Oct4 and Nanog, whose promoters are hypermethylated in somatic and trophoblast cells and hypomethylated at undifferentiated stages [46,47]. Modulators of DNA methylation, such as Tet1, Tet2 and Parp1, are essential for generating iPS cells. Interestingly, they are shown to facilitate the binding of Oct4 to Nanog and Esrrb www.sciencedirect.com
loci (Tet2 and Parp1) [48] or promote Nanog activity (Tet1/Tet2) [49]. Furthermore, recent work suggested a direct role of DNA methylation in cellular reprogramming. For example, many Klf4-binding sites that were demethylated in differentiated cells were methylated in ES cells, Klf4 has a particular affinity for a specific methylated motif [50], and methylcytosine interactors were enriched with DNA binding zinc fingers (such as Klf4) [51]. The authors hypothesize a role for DNA methylation in promoting Klf4 binding site recognition during early stages of somatic cell differentiation. Although DNA demethylation has been implicated as an integral part of cellular reprogramming, chemical treatments can overcome the imprinting that impairs iPS cell reprogramming. For example, ascorbic acid acts directly over the methylated DNA to prevent the function of Dnmt3 at the Dlk1-Dio3 locus [52], and histone deacetylase inhibitors reactivate gene expression without affecting methylation status [53].
Summary and outlook So far, cellular reprogramming has relied mainly on DNAbinding transcription factors, as these are thought to have the required instructive potential to alter cell fate. This potential relies on and can be greatly enhanced by regulators of chromatin state. The identification of general transcriptional machinery components, such as TFIID, as essential and potent enhancers of reprogramming [54] indicates that many levels of regulation can be deployed to modulate cellular reprogramming. As reprogramming approaches become more diverse and mature, clearer strategies based on underlying biology will be essential. Ongoing efforts to map the epigenomic landscapes of differentiating cells [55–57] will likely be useful to define general rules of chromatin-level regulation that can be deployed to reliably enhance cellular reprogramming.
References and recommended reading Papers of particular interest, published within the period of review, have been highlighted as: of special interest of outstanding interest
1.
Ho L, Crabtree GR: Chromatin remodelling during development. Nature 2010, 463:474-484.
2.
Katsuyama T, Paro R: Epigenetic reprogramming during tissue regeneration. FEBS Lett 2011, 585:1617-1624.
3.
Vierbuchen T, Wernig M: Molecular roadblocks for cellular reprogramming. Mol Cell 2012, 47:827-838.
4.
Briggs R, King TJ: Transplantation of living nuclei from blastula cells into enucleated frogs’ eggs. Proc Natl Acad Sci U S A 1952, 38:455-463.
5.
Gurdon JB: Adult frogs derived from the nuclei of single somatic cells. Dev Biol 1962, 4:256-273.
6.
Wilmut I, Schnieke AE, McWhir J, Kind AJ, Campbell KH: Viable offspring derived from fetal and adult mammalian cells. Nature 1997, 385:810-813. Current Opinion in Genetics & Development 2013, 23:556–561
560 Cell reprogramming
7.
Eggan K, Baldwin K, Tackett M, Osborne J, Gogos J, Chess A, Axel R, Jaenisch R: Mice cloned from olfactory sensory neurons. Nature 2004, 428:44-49.
25. Chan YS, Goke J, Lu X, Venkatesan N, Feng B, Su IH, Ng HH: A PRC2 dependent repressive role of PRDM14 in human embryonic stem cells and iPSC reprogramming. Stem Cells 2012, 31:682-692.
8.
Blau HM, Pavlath GK, Hardeman EC, Chiu CP, Silberstein L, Webster SG, Miller SC, Webster C: Plasticity of the differentiated state. Science 1985, 230:758-766.
9.
Davis RL, Weintraub H, Lassar AB: Expression of a single transfected cDNA converts fibroblasts to myoblasts. Cell 1987, 51:987-1000.
26. Mansour AA, Gafni O, Weinberger L, Zviran A, Ayyash M, Rais Y, Krupalnik V, Zerbib M, Amann-Zalcenstein D, Maza I et al.: The H3K27 demethylase Utx regulates somatic and germ cell epigenetic reprogramming. Nature 2012, 488:409-413.
10. Weintraub H, Tapscott SJ, Davis RL, Thayer MJ, Adam MA, Lassar AB, Miller AD: Activation of muscle-specific genes in pigment, nerve, fat, liver, and fibroblast cell lines by forced expression of MyoD. Proc Natl Acad Sci U S A 1989, 86: 5434-5438. 11. Takahashi K, Yamanaka S: Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 2006, 126:663-676. 12. Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, Yamanaka S: Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 2007, 131:861-872. 13. Huangfu D, Maehr R, Guo W, Eijkelenboom A, Snitow M, Chen AE, Melton DA: Induction of pluripotent stem cells by defined factors is greatly improved by small-molecule compounds. Nat Biotechnol 2008, 26:795-797. 14. Shi Y, Do JT, Desponts C, Hahm HS, Scholer HR, Ding S: A combined chemical and genetic approach for the generation of induced pluripotent stem cells. Cell Stem Cell 2008, 2:525528. 15. Schuettengruber B, Cavalli G: Recruitment of Polycomb group complexes and their role in the dynamic regulation of cell fate choice. Development 2009, 136:3531-3542. 16. Ringrose L, Paro R: Polycomb/Trithorax response elements and epigenetic memory of cell identity. Development 2007, 134:223-232. 17. Simon JA, Kingston RE: Mechanisms of Polycomb gene silencing: knowns and unknowns. Nat Rev Mol Cell Biol 2009, 10:697-708. 18. Min J, Zhang Y, Xu RM: Structural basis for specific binding of Polycomb chromodomain to histone H3 methylated at Lys 27. Genes Dev 2003, 17:1823-1828. 19. Pereira CF, Piccolo FM, Tsubouchi T, Sauer S, Ryan NK, Bruno L, Landeira D, Santos J, Banito A, Gil J et al.: ESCs require PRC2 to direct the successful reprogramming of differentiated cells toward pluripotency. Cell Stem Cell 2010, 6:547-556. This work highlighted essential role of Polycomb-group proteins during ES cells reprogramming. Using PRC1-depleted and PRC2-depleted ES cells, the authors showed the incapability of these cells to be reprogrammed, but also the pivotal role of PRC2 in the control of the chromatin conformation changes required for the ES cell reprogramming. 20. Azuara V, Perry P, Sauer S, Spivakov M, Jorgensen HF, John RM, Gouti M, Casanova M, Warnes G, Merkenschlager M et al.: Chromatin signatures of pluripotent cell lines. Nat Cell Biol 2006, 8:532-538. 21. Bernstein E, Duncan EM, Masui O, Gil J, Heard E, Allis CD: Mouse Polycomb proteins bind differentially to methylated histone H3 and RNA and are enriched in facultative heterochromatin. Mol Cell Biol 2006, 26:2560-2569. 22. Lee TI, Jenner RG, Boyer LA, Guenther MG, Levine SS, Kumar RM, Chevalier B, Johnstone SE, Cole MF, Isono K et al.: Control of developmental regulators by Polycomb in human embryonic stem cells. Cell 2006, 125:301-313. 23. Yang J, Chai L, Fowles TC, Alipio Z, Xu D, Fink LM, Ward DC, Ma Y: Genome-wide analysis reveals Sall4 to be a major regulator of pluripotency in murine-embryonic stem cells. Proc Natl Acad Sci U S A 2008, 105:19756-19761. 24. Fragola G, Germain PL, Laise P, Cuomo A, Blasimme A, Gross F, Signaroldi E, Bucci G, Sommer C, Pruneri G et al.: Cell reprogramming requires silencing of a core subset of Polycomb targets. PLoS Genet 2013, 9:e1003292. Current Opinion in Genetics & Development 2013, 23:556–561
27. Zhao W, Li Q, Ayers S, Gu Y, Shi Z, Zhu Q, Chen Y, Wang HY, Wang RF: Jmjd3 inhibits reprogramming by upregulating expression of INK4a/Arf and targeting PHF20 for ubiquitination. Cell 2013, 152:1037-1050. 28. Zhu J, Adli M, Zou JY, Verstappen G, Coyne M, Zhang X, Durham T, Miri M, Deshpande V, De Jager PL et al.: Genome-wide chromatin state transitions associated with developmental and environmental cues. Cell 2013, 152:642-654. 29. Lister R, Pelizzola M, Kida YS, Hawkins RD, Nery JR, Hon G, Antosiewicz-Bourget J, O’Malley R, Castanon R, Klugman S et al.: Hotspots of aberrant epigenomic reprogramming in human induced pluripotent stem cells. Nature 2011, 471:68-73. 30. Ruiz S, Diep D, Gore A, Panopoulos AD, Montserrat N, Plongthongkum N, Kumar S, Fung HL, Giorgetti A, Bilic J et al.: Identification of a specific reprogramming-associated epigenetic signature in human induced pluripotent stem cells. Proc Natl Acad Sci U S A 2012, 109:16196-16201. 31. Onder TT, Kara N, Cherry A, Sinha AU, Zhu N, Bernt KM, Cahan P, Marcarci BO, Unternaehrer J, Gupta PB et al.: Chromatinmodifying enzymes as modulators of reprogramming. Nature 2012, 483:598-602. By inhibiting different chromatin-modifying proteins with shRNAs, the authors of this work identified enzymes that reduce or enhance the efficiency of iPS cells generation and highlighted the importance of the chromatin-modifying elements in cell reprogramming. 32. Soufi A, Donahue G, Zaret KS: Facilitators and impediments of the pluripotency reprogramming factors’ initial engagement with the genome. Cell 2012, 151:994-1004. In this work, the authors identified the differential binding pattern of Oct4, Sox2, Klf4 and c-Myc in different stages of somatic differentiation and ES cells. This pattern is clearly affected by the distribution of histone modifications along the genome, which acts as a repressor of differentiation. This work also shows the pivotal role of c-Myc as a facilitating factor for the interaction of Oct4, Sox2 and Klf4 with the genome. 33. Nagl NG Jr, Wang X, Patsialou A, Van Scoy M, Moran E: Distinct mammalian SWI/SNF chromatin remodeling complexes with opposing roles in cell-cycle control. EMBO J 2007, 26:752-763. 34. Han P, Hang CT, Yang J, Chang CP: Chromatin remodeling in cardiovascular development and physiology. Circ Res 2011, 108:378-396. 35. Ho L, Ronan JL, Wu J, Staahl BT, Chen L, Kuo A, Lessard J, Nesvizhskii AI, Ranish J, Crabtree GR: An embryonic stem cell chromatin remodeling complex, esBAF, is essential for embryonic stem cell self-renewal and pluripotency. Proc Natl Acad Sci U S A 2009, 106:5181-5186. 36. Limpert AS, Bai S, Narayan M, Wu J, Yoon SO, Carter BD, Lu QR: NF-kappaB forms a complex with the chromatin remodeler BRG1 to regulate Schwann cell differentiation. J Neurosci 2013, 33:2388-2397. 37. Gao H, Lukin K, Ramirez J, Fields S, Lopez D, Hagman J: Opposing effects of SWI/SNF and Mi-2/NuRD chromatin remodeling complexes on epigenetic reprogramming by EBF and Pax5. Proc Natl Acad Sci U S A 2009, 106:11258-11263. 38. Barneda-Zahonero B, Roman-Gonzalez L, Collazo O, Mahmoudi T, Parra M: Epigenetic regulation of B lymphocyte differentiation, transdifferentiation, and reprogramming. Comp Funct Genomics 2012, 2012:564381. 39. Lickert H, Takeuchi JK, Von Both I, Walls JR, McAuliffe F, Adamson SL, Henkelman RM, Wrana JL, Rossant J, Bruneau BG: Baf60c is essential for function of BAF chromatin remodelling complexes in heart development. Nature 2004, 432:107-112. 40. Takeuchi JK, Bruneau BG: Directed transdifferentiation of mouse mesoderm to heart tissue by defined factors. Nature 2009, 459:708-711. www.sciencedirect.com
Chromatin regulation of reprogramming Luna-Zurita and Bruneau 561
41. Zaret KS, Carroll JS: Pioneer transcription factors: establishing competence for gene expression. Genes Dev 2011, 25: 2227-2241. 42. Albini S, Coutinho P, Malecova B, Giordani L, Savchenko A, Forcales SV, Puri PL: Epigenetic reprogramming of human embryonic stem cells into skeletal muscle cells and generation of contractile myospheres. Cell Rep 2013, 3: 661-670. 43. Singhal N, Graumann J, Wu G, Arauzo-Bravo MJ, Han DW, Greber B, Gentile L, Mann M, Scholer HR: Chromatin-remodeling components of the BAF complex facilitate reprogramming. Cell 2010, 141:943-955. In this work, the authors identified different components of the BAF chromatin-remodeling complex, able to increase the efficiency of cell reprogramming due to the activity of Oct4, Sox2, Klf4 and c-Myc in somatic cells. 44. Kleger A, Mahaddalkar PU, Katz SF, Lechel A, Joo JY, Loya K, Lin Q, Hartmann D, Liebau S, Kraus JM et al.: Increased reprogramming capacity of mouse liver progenitor cells, compared with differentiated liver cells, requires the BAF complex. Gastroenterology 2012, 142:907-917. 45. Isagawa T, Nagae G, Shiraki N, Fujita T, Sato N, Ishikawa S, Kume S, Aburatani H: DNA methylation profiling of embryonic stem cell differentiation into the three germ layers. PLoS ONE 2011, 6:e26052. 46. Hattori N, Nishino K, Ko YG, Hattori N, Ohgane J, Tanaka S, Shiota K: Epigenetic control of mouse Oct-4 gene expression in embryonic stem cells and trophoblast stem cells. J Biol Chem 2004, 279:17063-17069. 47. Hattori N, Imao Y, Nishino K, Hattori N, Ohgane J, Yagi S, Tanaka S, Shiota K: Epigenetic regulation of Nanog gene in embryonic stem and trophoblast stem cells. Genes Cells 2007, 12:387-396. 48. Doege CA, Inoue K, Yamashita T, Rhee DB, Travis S, Fujita R, Guarnieri P, Bhagat G, Vanti WB, Shih A et al.: Early-stage epigenetic modification during somatic cell reprogramming by Parp1 and Tet2. Nature 2012, 488:652-655. The authors of this work identified two factors, Parp1 and Tet2, as essentials for the OSKM-dependent somatic reprogramming. Parp1 and Tet2 interact with Nanog and Esrrb loci, promoting the transition between 5-methylcytosine (5mC), 5-hydroxyethylcytosine (5hmC) and cytosine (C). Further, the work establishes 5hmC as an epigenetic mark and not as a transition step during loci demethylation.
www.sciencedirect.com
49. Costa Y, Ding J, Theunissen TW, Faiola F, Hore TA, Shliaha PV, Fidalgo M, Saunders A, Lawrence M, Dietmann S et al.: NANOG dependent function of TET1 and TET2 in establishment of pluripotency. Nature 2013, 495:370-374. This work shows how NANOG, a transcription factor with a pivotal role in the maintaining of the pluripotency stage, interacts with the 5mC hydrolases Tet1 and Tet2 during somatic cell reprogramming, promoting the expression of pluripotency genes and increasing the efficiency of cell reprogramming. 50. Chen X, Xu H, Yuan P, Fang F, Huss M, Vega VB, Wong E, Orlov YL, Zhang W, Jiang J et al.: Integration of external signaling pathways with the core transcriptional network in embryonic stem cells. Cell 2008, 133:1106-1117. 51. Spruijt CG, Gnerlich F, Smits AH, Pfaffeneder T, Jansen PW, Bauer C, Munzel M, Wagner M, Muller M, Khan F et al.: Dynamic readers for 5-(hydroxy)methylcytosine and its oxidized derivatives. Cell 2013, 152:1146-1159. 52. Stadtfeld M, Apostolou E, Ferrari F, Choi J, Walsh RM, Chen T, Ooi SS, Kim SY, Bestor TH, Shioda T et al.: Ascorbic acid prevents loss of Dlk1-Dio3 imprinting and facilitates generation of all-iPS cell mice from terminally differentiated B cells. Nat Genet 2012, 44:S391-S392. 53. Raynal NJ, Si J, Taby RF, Gharibyan V, Ahmed S, Jelinek J, Estecio MR, Issa JP: DNA methylation does not stably lock gene expression but instead serves as a molecular mark for gene silencing memory. Cancer Res 2012, 72:1170-1181. 54. Pijnappel WW, Esch D, Baltissen MP, Wu G, Mischerikow N, Bergsma AJ, van der Wal E, Han DW, Bruch HV, Moritz S et al.: A central role for TFIID in the pluripotent transcription circuitry. Nature 2013, 495:516-519. 55. Wamstad JA, Alexander JM, Truty RM, Shrikumar A, Li F, Eilertson KE, Ding H, Wylie JN, Pico AR, Capra JA et al.: Dynamic and coordinated epigenetic regulation of developmental transitions in the cardiac lineage. Cell 2012, 151:206-220. 56. Beshiri ML, Holmes KB, Richter WF, Hess S, Islam AB, Yan Q, Plante L, Litovchick L, Gevry N, Lopez-Bigas N et al.: Coordinated repression of cell cycle genes by KDM5A and E2F4 during differentiation. Proc Natl Acad Sci U S A 2012, 109:18499-18504. 57. Li Z, Gadue P, Chen K, Jiao Y, Tuteja G, Schug J, Li W, Kaestner KH: Foxa2 and H2A.Z mediate nucleosome depletion during embryonic stem cell differentiation. Cell 2012, 151:1608-1616.
Current Opinion in Genetics & Development 2013, 23:556–561
ScienceDirect Chromatin modulators as facilitating factors in cellular reprogramming Luis Luna-Zurita1 and Benoit G Bruneau1,2 In the last few years, cellular reprogramming has emerged as a means to alter cellular identity and generate diverse cell types for disease modeling, drug testing, and potential therapeutic use. Since each cell type is a result of a specific gene expression profile finely regulated by the activity of a repertoire of transcription factors (TFs), reprogramming approaches have, thus far, been relatively inefficient and based largely on the forced expression of selective cell type-specific TFs. TFs function within the confines of chromatin, and the chromatin states can in turn be modulated by TF activity. Therefore, the knowledge of how chromatin remodeling factors alter chromatin structure, control TF activity and gene expression has led to an improved reprogramming efficiency and extended the number of cellular types that can be generated by cellular reprogramming. Here we review recent insights into the role and mechanisms by which chromatin remodeling, histone modifications, and DNA methylation contribute to cellular differentiation and reprogramming. Addresses 1 Gladstone Institute of Cardiovascular Disease, San Francisco, CA 94158, USA 2 Department of Pediatrics, Cardiovascular Research Institute, University of California, San Francisco, CA 94158, USA Corresponding author: Bruneau, Benoit G ([email protected])
Current Opinion in Genetics & Development 2013, 23:556–561 This review comes from a themed issue on Cell reprogramming Edited by Huck Hui Ng and Patrick Tam For a complete overview see the Issue and the Editorial Available online 28th August 2013 0959-437X/$ – see front matter, # 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.gde.2013.07.002
dynamic, allowing context-dependent changes of gene expression [1]. Once established, chromatin structure remains relatively stable for a specific differentiated stage [3], and it was assumed that the genomic structural changes during cellular differentiation were irreversible. Studies involving the transfer of somatic cell nuclei to enucleated oocytes [4,5] in Xenopus and similar experiments later in mammals [6], including studies with postmitotic cells as donor cells [7], showed that oocytes provide a specific environment that can reverse the chromatin architecture associated with the differentiated stage. Similarly, heterokaryon fusion experiments showed that the nucleus from one cell type could be efficiently reprogrammed to express the transcriptome of a different somatic cell type [8]. Although these studies did not identify the nuclear factors that were in play, other reports [9,10] in which certain cells were induced into skeletal muscle by introducing the bHLH transcription factor MyoD, showed that a single defined factor could reprogram cellular identity. The ability of TFs to reprogram somatic cells to a pluripotent state was first demonstrated by pioneering work from Shinya Yamanaka’s laboratory [11,12]. They showed that a handful of defined factors was sufficient to completely reset the somatic nucleus’s differentiation program. Since then, similar approaches with tissuespecific TFs have been used to reprogram cells directly to other somatic cell types. Although there are still a lot of questions about how the reprogramming TFs effect these lineage conversions, due to multiple studies aimed at identifying modulators of this process, the role of chromatin-level regulation in facilitating cell reprogramming and TFs activity is becoming better understood.
Introduction
Histone modifications
TFs are the main effectors of cell differentiation and govern gene expression changes that will give rise to the diverse cellular types that emerge during development. While TFs regulate gene expression, TF–DNA interactions are influenced by the dynamic structural and biochemical properties of chromatin. Chromatin states can be regulated by different entities or processes that modify chromatin biochemical properties, chromatin conformation or nucleosome distribution. They include histone and DNA modifications, nucleosome positioning, exchange of histone variants, and chromatin remodeler proteins [1,2]. These chromatin modifications are
Chromatin can transition between ‘open’ (euchromatin) and ‘closed’ (heterochromatin) states. Posttranslational histone modifications, including methylation, acetylation and ubiquitination, determine the changes in status. These dynamic marks promote structural changes in chromatin conformation and are essential in the control of gene expression.
Current Opinion in Genetics & Development 2013, 23:556–561
These same modifications modulate the efficiency of induced pluripotent stem (iPS) cell reprogramming [13,14]. For example, small molecules have been found to alter histone acetylation (valproic acid) or methylation www.sciencedirect.com
Chromatin regulation of reprogramming Luna-Zurita and Bruneau 557
(BIX-01294). Members of the Polycomb group (PcG) and Trithorax group (TrxG) are key in controlling gene expression during cell differentiation and reprogramming. PcG and TrxG are protein complexes with opposing roles in cell differentiation. While PcG activity is associated with gene silencing and heterochromatin stabilization, TrxG maintains active transcriptional states (reviewed in [2,15,16]). In mammals, Polycomb repressor complex 2 (PRC2) activity depends on three subunits: Ezh2, Eed and Suz12. Although Ezh2 is the enzymatic subunit of PRC2, Suz12 and Eed are also crucial to methylate (monomethylation, dimethylation and trimethylation) lysine 27 on histone 3 (H3K27) [17]. In contrast to H3K27me1 and H3K27me2, which are broadly distributed across the genome, H3K27me3 is the mark most associated with the repressive function of this complex [17]. H3K27me3 acts as a docking site for the PcG complex PRC1 (the other main Polycomb repressor complex) [18] (and reviewed in [19]) and promotes the repressing mark H2AK119Ub. PRC1 and PRC2 colocalize on a large set of genes in ES cells, concomitant with an enrichment of the repressor mark H3K27me3 [20,21]. They also colocalize with a fraction of binding sites for essential factors for the pluripotency stage, such as Oct4, Nanog, Sox2 and Sall4 [22,23]. These results suggest that PRC1/2 acts through these TFs to maintain the pluripotency state. Interestingly, global loss of H3K27me3 after genetic deletion of Ezh2 does not significantly affect iPSC reprogramming [24]. This does not necessarily mean that PRC complex activity is not required for somatic reprogramming. Perhaps Ezh1 (the other known PRC2 enzymatic subunit) retains H3K27me3 at specific loci [24]. PRDM14, a PR domain-containing transcriptional regulator, interacts directly with PRC2 to establish H3K27me3 modifications in human ES cells. This observation supports the effect of PRDM14 repression to increase the efficiency of iPSC induction in human cells [25]. H3K27 demethylases, such as Utx or Jmjd3, have been implicated in modifying histones during cellular reprogramming independently of PcG/TrxG. Utx has specific roles during the re-establishment of pluripotency and in germ cell development. In conjunction with the pluripotency factors Oct4, Sox2 and Klf4, it removes the H3K27me3 repressive mark during the reprogramming of somatic cells [26] (Figure 1). Jmjd3 inhibits iPS cell reprogramming, partly via demethylase-dependent activity, but also via a demethylase-independent targeting of PHF20 for ubiquitination. PHF20 interacts with Wdr5 to regulate gene expression and is itself essential for iPS cell generation [27]. H3K9me3 is a constitutive heterochromatin mark in differentiated cells but this mark is not abundant in ES www.sciencedirect.com
cells or iPS cells [28]. On the basis of the effectiveness of inhibition of the H3K9me histone methyltransferase G9a in promoting iPS cell production, H3K9me3 has been proposed to be one of the barriers for iPS cell reprogramming [14]. Curiously, regions enriched in H3K9me3 overlap with ‘hot spots’ of aberrant epigenomic reprogramming [29,30]. Interestingly, the deficiency of Suv39h1, the enzyme catalyzing H3K9me3, also enhances reprogramming [31] (Figure 1). Thus, although H3K9me3 patterns are largely reset in iPS cells, these domains may present a barrier to reprogramming. The initial binding of pluripotency TFs Oct4, Klf4, and Sox2 to large genomic regions enriched in H3K9me3 loci is blocked, but this is likely to be overcome later in the reprogramming process and can be enhanced by loss of Suv39h1 and Suv39h2 [32]. Inhibition of another histone-modifying enzyme, Dot1L, also enhances cell reprogramming. Dot1L catalyzes the H3K79me2 modification, and this enhanced reprogramming might be due, at least in part, to the lack of this mark in genes that should be repressed during reprogramming [31].
Chromatin-remodeling complexes Nucleosome distribution can be altered at specific loci by chromatin remodeling complexes (CRCs), and this action provides a mechanism for gene regulation distinct from histone modifications. There are various families of CRCs, including ATP-dependent CRCs ISWI (imitation switch), CHD (chromodomain-helicase–DNA binding), INO80 (inositol requiring 80), and SWI/SNF (switching defective/sucrose nonfermenting) complexes [1]. The pivotal role of SWI/SNF complexes during cellular differentiation and reprogramming, where they act as proliferation repressors, is well described [33], and here we will focus mainly on this process. SWI/SNF complexes in vertebrates (Brm/Brg1-associated factor (BAF) complexes) function, even for the same gene, as both transcriptional activators and repressors at different developmental stages [1,34]. They comprise at least 12 protein subunits. The ATPase subunit of the BAF complex is encoded by one of the two highly homologous genes, Brg1 (Brahma-related gene 1) and Brm (Brahma). The subunit composition of BAF complexes also depends on the cellular context. For example, in ES cells, BAF complexes (known as esBAF) are defined by the presence of Brg1, BAF60a, and BAF155, and the absence of Brm, BAF60c, and BAF170. The esBAF complex directly interacts with key regulators of pluripotency, such as Oct4 and Sox2, and its activity is required to maintain the undifferentiated state of the ES cells [35] (Figure 1). BAF activity has been studied during the differentiation of several cellular types, such as Schwann cells [36] and Bcells, where differentiation is regulated by the balance between BAF and Mi-2/NuRD (CHD) activities [37,38]. In certain tissues, a BAF complex enrichment enhances Current Opinion in Genetics & Development 2013, 23:556–561
558 Cell reprogramming
Figure 1
DNMT BRG1 BAF60c
M
Baf155 Baf60a Brg1
PRDM14 PRC2
PRC1
Sox2 Oct4
TF
ES Cells
iPS Cells
Dedifferentiation Differentiation
G9a
BRG1 BAF155
Suv39h1
Utx
M
M
M
M
M
M Oct4
Differentiated Cell Tet1 Nanog Tet2
H3K9ac H3K4me3 H3K27me3 H3K9me3
H2AK119Ub
M
Cytosine Methylation
AA
Tet2
Parp1
Regulatory region of pluripotency genes Developmental genes Current Opinion in Genetics & Development
Cell reprogramming is regulated by changes in chromatin architecture and chemical modification of DNA. Activation histone marks (H3K4me3 and H3K9Ac) and hypomethylated pluripotency gene promoters allow a low chromatin-compaction status and high levels of gene expression. In contrast, repressive histone modifications (H3K27me3 and H3K9me3) and DNA methylation reduce RNA-PolII accessibility and gene expression. These modifications play a key role in maintaining the differentiated/undifferentiated state and represent strong barriers for cellular reprogramming. During ES/IPS cell differentiation, pluripotency gene promoters increase their methylation levels due to the activity of DNA methyltransferases (DNMTs). This modification prevents promoter interaction with TFs and inhibits gene expression. In contrast, TFs interactions required for cell differentiation are facilitated by different factors, such as the BAF-complex subunit Baf60c. The differentiation process requires the repression of factors implicated in the undifferentiated stage maintenance, such as the esBAF complex (characterized by the Brg1, BAF60a, and BAF155 subunits), that allows the interaction of pluripotency regulators, such as Sox2 and Oct4 to gene promoters, or repressive histone modification close to development promoters carried out by the PcG members PRC1/PRC2 or enzymes, such as PRDM14. During somatic cell reprogramming, the activity of histone demethylases, such as Utx, or BAF complexes characterized by the presence of BRG1 and BAF155 has been described as required for repressive histone marks removal and the acquisition of activation marks in regions where pluripotency genes are localized. Promoters of these genes must change their methylation status during this process. Thus, the activity of molecules, such as Tet1, Te2 or Parp1, reduces methylation levels, allowing the interaction of pluripotency factors, such as Nanog or Oct4. A similar demethylation effect can be achieved by adding ascorbic acid, which inhibits DNMTs activity and reduces methylation levels in gene promoters. Increased reprogramming efficiency can also be achieved by inhibiting molecules, such as the histone methyltranferases Suv39h1 or G9a, and removing the H3K9me3 repressive mark of plutipotency genes promoters.
Current Opinion in Genetics & Development 2013, 23:556–561
www.sciencedirect.com
Chromatin regulation of reprogramming Luna-Zurita and Bruneau 559
differentiation or reprogramming of cell type-specific programs. This is the case of Baf60c, a BAF complex subunit highly restricted to the myocardium and somites during mouse development [39]. During myocardial differentiation, Baf60c is essential for the function of DNA-binding transcription factors (Gata4, Tbx5), and Baf60c is part of a trio of factors sufficient to direct differentiation from mesoderm to myocardium. Baf60c facilitates the binding of Gata4 to its DNA targets and recruits the BAF complex to these loci [39–41]. Baf60c is also expressed in developing skeletal muscle and is essential for MyoD-dependent differentiation of human ES cells toward a skeletal muscle phenotype [42]. BAF complexes are also potent modulators of reprogramming to pluripotency. For the reprogramming of human fibroblasts into iPS cells, overexpression of BRG1 and the BAF155 subunit can replace c-Myc [43]. The mechanism by which BRG1 and BAF155 promote cell reprogramming is (presumably indirectly) by an increase of H3K4me3 and H3K9 acetylation (activation marks) on specific pluripotency-associated genes, and the reduction of the repressive mark H3K27me3 on OCT4 (Figure 1). Thus, BRG1 and BAF155 seem to promote a euchromatic state at the promoters of pluripotency genes, but importantly also induce OCT4 binding to the promoter of pluripotency-specific genes, including to its own promoter. BAF activity also promotes somatic cell reprogramming in fetal and adult liver progenitor cells (LPCs). LPCs are more efficiently reprogrammed than differentiated liver cells, when associated with endogenous expression of reprogramming factors (Klf4 and c-Myc) and BAF complex members Baf155 and Brg1, which mediate chromatin remodeling during reprogramming. Knockdown of BAF complex members negated the increased reprogramming efficiency of LPCs, compared with non-LPCs [44]. Altogether, these findings show that, for certain cellular programs, chromatin remodelers are not only permissive, but also in fact instructive.
DNA methylation and demethylation DNA methylation at cytosine residues is generally associated with silencing of gene expression. DNA methyltransferases (DNMTs) catalyze the covalent attachment of a methyl residue to the carbon 5 position of cytosine residues of CpG dinucleotides [2]. Several lines of evidence show the importance of DNA methylation in regulating gene expression during cell differentiation. Thus, while methylation of gene promoter regions is widely seen in differentiated cells, most of these genes are unmethylated in ES cells and sperm [45]. This is the case of Oct4 and Nanog, whose promoters are hypermethylated in somatic and trophoblast cells and hypomethylated at undifferentiated stages [46,47]. Modulators of DNA methylation, such as Tet1, Tet2 and Parp1, are essential for generating iPS cells. Interestingly, they are shown to facilitate the binding of Oct4 to Nanog and Esrrb www.sciencedirect.com
loci (Tet2 and Parp1) [48] or promote Nanog activity (Tet1/Tet2) [49]. Furthermore, recent work suggested a direct role of DNA methylation in cellular reprogramming. For example, many Klf4-binding sites that were demethylated in differentiated cells were methylated in ES cells, Klf4 has a particular affinity for a specific methylated motif [50], and methylcytosine interactors were enriched with DNA binding zinc fingers (such as Klf4) [51]. The authors hypothesize a role for DNA methylation in promoting Klf4 binding site recognition during early stages of somatic cell differentiation. Although DNA demethylation has been implicated as an integral part of cellular reprogramming, chemical treatments can overcome the imprinting that impairs iPS cell reprogramming. For example, ascorbic acid acts directly over the methylated DNA to prevent the function of Dnmt3 at the Dlk1-Dio3 locus [52], and histone deacetylase inhibitors reactivate gene expression without affecting methylation status [53].
Summary and outlook So far, cellular reprogramming has relied mainly on DNAbinding transcription factors, as these are thought to have the required instructive potential to alter cell fate. This potential relies on and can be greatly enhanced by regulators of chromatin state. The identification of general transcriptional machinery components, such as TFIID, as essential and potent enhancers of reprogramming [54] indicates that many levels of regulation can be deployed to modulate cellular reprogramming. As reprogramming approaches become more diverse and mature, clearer strategies based on underlying biology will be essential. Ongoing efforts to map the epigenomic landscapes of differentiating cells [55–57] will likely be useful to define general rules of chromatin-level regulation that can be deployed to reliably enhance cellular reprogramming.
References and recommended reading Papers of particular interest, published within the period of review, have been highlighted as: of special interest of outstanding interest
1.
Ho L, Crabtree GR: Chromatin remodelling during development. Nature 2010, 463:474-484.
2.
Katsuyama T, Paro R: Epigenetic reprogramming during tissue regeneration. FEBS Lett 2011, 585:1617-1624.
3.
Vierbuchen T, Wernig M: Molecular roadblocks for cellular reprogramming. Mol Cell 2012, 47:827-838.
4.
Briggs R, King TJ: Transplantation of living nuclei from blastula cells into enucleated frogs’ eggs. Proc Natl Acad Sci U S A 1952, 38:455-463.
5.
Gurdon JB: Adult frogs derived from the nuclei of single somatic cells. Dev Biol 1962, 4:256-273.
6.
Wilmut I, Schnieke AE, McWhir J, Kind AJ, Campbell KH: Viable offspring derived from fetal and adult mammalian cells. Nature 1997, 385:810-813. Current Opinion in Genetics & Development 2013, 23:556–561
560 Cell reprogramming
7.
Eggan K, Baldwin K, Tackett M, Osborne J, Gogos J, Chess A, Axel R, Jaenisch R: Mice cloned from olfactory sensory neurons. Nature 2004, 428:44-49.
25. Chan YS, Goke J, Lu X, Venkatesan N, Feng B, Su IH, Ng HH: A PRC2 dependent repressive role of PRDM14 in human embryonic stem cells and iPSC reprogramming. Stem Cells 2012, 31:682-692.
8.
Blau HM, Pavlath GK, Hardeman EC, Chiu CP, Silberstein L, Webster SG, Miller SC, Webster C: Plasticity of the differentiated state. Science 1985, 230:758-766.
9.
Davis RL, Weintraub H, Lassar AB: Expression of a single transfected cDNA converts fibroblasts to myoblasts. Cell 1987, 51:987-1000.
26. Mansour AA, Gafni O, Weinberger L, Zviran A, Ayyash M, Rais Y, Krupalnik V, Zerbib M, Amann-Zalcenstein D, Maza I et al.: The H3K27 demethylase Utx regulates somatic and germ cell epigenetic reprogramming. Nature 2012, 488:409-413.
10. Weintraub H, Tapscott SJ, Davis RL, Thayer MJ, Adam MA, Lassar AB, Miller AD: Activation of muscle-specific genes in pigment, nerve, fat, liver, and fibroblast cell lines by forced expression of MyoD. Proc Natl Acad Sci U S A 1989, 86: 5434-5438. 11. Takahashi K, Yamanaka S: Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 2006, 126:663-676. 12. Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, Yamanaka S: Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 2007, 131:861-872. 13. Huangfu D, Maehr R, Guo W, Eijkelenboom A, Snitow M, Chen AE, Melton DA: Induction of pluripotent stem cells by defined factors is greatly improved by small-molecule compounds. Nat Biotechnol 2008, 26:795-797. 14. Shi Y, Do JT, Desponts C, Hahm HS, Scholer HR, Ding S: A combined chemical and genetic approach for the generation of induced pluripotent stem cells. Cell Stem Cell 2008, 2:525528. 15. Schuettengruber B, Cavalli G: Recruitment of Polycomb group complexes and their role in the dynamic regulation of cell fate choice. Development 2009, 136:3531-3542. 16. Ringrose L, Paro R: Polycomb/Trithorax response elements and epigenetic memory of cell identity. Development 2007, 134:223-232. 17. Simon JA, Kingston RE: Mechanisms of Polycomb gene silencing: knowns and unknowns. Nat Rev Mol Cell Biol 2009, 10:697-708. 18. Min J, Zhang Y, Xu RM: Structural basis for specific binding of Polycomb chromodomain to histone H3 methylated at Lys 27. Genes Dev 2003, 17:1823-1828. 19. Pereira CF, Piccolo FM, Tsubouchi T, Sauer S, Ryan NK, Bruno L, Landeira D, Santos J, Banito A, Gil J et al.: ESCs require PRC2 to direct the successful reprogramming of differentiated cells toward pluripotency. Cell Stem Cell 2010, 6:547-556. This work highlighted essential role of Polycomb-group proteins during ES cells reprogramming. Using PRC1-depleted and PRC2-depleted ES cells, the authors showed the incapability of these cells to be reprogrammed, but also the pivotal role of PRC2 in the control of the chromatin conformation changes required for the ES cell reprogramming. 20. Azuara V, Perry P, Sauer S, Spivakov M, Jorgensen HF, John RM, Gouti M, Casanova M, Warnes G, Merkenschlager M et al.: Chromatin signatures of pluripotent cell lines. Nat Cell Biol 2006, 8:532-538. 21. Bernstein E, Duncan EM, Masui O, Gil J, Heard E, Allis CD: Mouse Polycomb proteins bind differentially to methylated histone H3 and RNA and are enriched in facultative heterochromatin. Mol Cell Biol 2006, 26:2560-2569. 22. Lee TI, Jenner RG, Boyer LA, Guenther MG, Levine SS, Kumar RM, Chevalier B, Johnstone SE, Cole MF, Isono K et al.: Control of developmental regulators by Polycomb in human embryonic stem cells. Cell 2006, 125:301-313. 23. Yang J, Chai L, Fowles TC, Alipio Z, Xu D, Fink LM, Ward DC, Ma Y: Genome-wide analysis reveals Sall4 to be a major regulator of pluripotency in murine-embryonic stem cells. Proc Natl Acad Sci U S A 2008, 105:19756-19761. 24. Fragola G, Germain PL, Laise P, Cuomo A, Blasimme A, Gross F, Signaroldi E, Bucci G, Sommer C, Pruneri G et al.: Cell reprogramming requires silencing of a core subset of Polycomb targets. PLoS Genet 2013, 9:e1003292. Current Opinion in Genetics & Development 2013, 23:556–561
27. Zhao W, Li Q, Ayers S, Gu Y, Shi Z, Zhu Q, Chen Y, Wang HY, Wang RF: Jmjd3 inhibits reprogramming by upregulating expression of INK4a/Arf and targeting PHF20 for ubiquitination. Cell 2013, 152:1037-1050. 28. Zhu J, Adli M, Zou JY, Verstappen G, Coyne M, Zhang X, Durham T, Miri M, Deshpande V, De Jager PL et al.: Genome-wide chromatin state transitions associated with developmental and environmental cues. Cell 2013, 152:642-654. 29. Lister R, Pelizzola M, Kida YS, Hawkins RD, Nery JR, Hon G, Antosiewicz-Bourget J, O’Malley R, Castanon R, Klugman S et al.: Hotspots of aberrant epigenomic reprogramming in human induced pluripotent stem cells. Nature 2011, 471:68-73. 30. Ruiz S, Diep D, Gore A, Panopoulos AD, Montserrat N, Plongthongkum N, Kumar S, Fung HL, Giorgetti A, Bilic J et al.: Identification of a specific reprogramming-associated epigenetic signature in human induced pluripotent stem cells. Proc Natl Acad Sci U S A 2012, 109:16196-16201. 31. Onder TT, Kara N, Cherry A, Sinha AU, Zhu N, Bernt KM, Cahan P, Marcarci BO, Unternaehrer J, Gupta PB et al.: Chromatinmodifying enzymes as modulators of reprogramming. Nature 2012, 483:598-602. By inhibiting different chromatin-modifying proteins with shRNAs, the authors of this work identified enzymes that reduce or enhance the efficiency of iPS cells generation and highlighted the importance of the chromatin-modifying elements in cell reprogramming. 32. Soufi A, Donahue G, Zaret KS: Facilitators and impediments of the pluripotency reprogramming factors’ initial engagement with the genome. Cell 2012, 151:994-1004. In this work, the authors identified the differential binding pattern of Oct4, Sox2, Klf4 and c-Myc in different stages of somatic differentiation and ES cells. This pattern is clearly affected by the distribution of histone modifications along the genome, which acts as a repressor of differentiation. This work also shows the pivotal role of c-Myc as a facilitating factor for the interaction of Oct4, Sox2 and Klf4 with the genome. 33. Nagl NG Jr, Wang X, Patsialou A, Van Scoy M, Moran E: Distinct mammalian SWI/SNF chromatin remodeling complexes with opposing roles in cell-cycle control. EMBO J 2007, 26:752-763. 34. Han P, Hang CT, Yang J, Chang CP: Chromatin remodeling in cardiovascular development and physiology. Circ Res 2011, 108:378-396. 35. Ho L, Ronan JL, Wu J, Staahl BT, Chen L, Kuo A, Lessard J, Nesvizhskii AI, Ranish J, Crabtree GR: An embryonic stem cell chromatin remodeling complex, esBAF, is essential for embryonic stem cell self-renewal and pluripotency. Proc Natl Acad Sci U S A 2009, 106:5181-5186. 36. Limpert AS, Bai S, Narayan M, Wu J, Yoon SO, Carter BD, Lu QR: NF-kappaB forms a complex with the chromatin remodeler BRG1 to regulate Schwann cell differentiation. J Neurosci 2013, 33:2388-2397. 37. Gao H, Lukin K, Ramirez J, Fields S, Lopez D, Hagman J: Opposing effects of SWI/SNF and Mi-2/NuRD chromatin remodeling complexes on epigenetic reprogramming by EBF and Pax5. Proc Natl Acad Sci U S A 2009, 106:11258-11263. 38. Barneda-Zahonero B, Roman-Gonzalez L, Collazo O, Mahmoudi T, Parra M: Epigenetic regulation of B lymphocyte differentiation, transdifferentiation, and reprogramming. Comp Funct Genomics 2012, 2012:564381. 39. Lickert H, Takeuchi JK, Von Both I, Walls JR, McAuliffe F, Adamson SL, Henkelman RM, Wrana JL, Rossant J, Bruneau BG: Baf60c is essential for function of BAF chromatin remodelling complexes in heart development. Nature 2004, 432:107-112. 40. Takeuchi JK, Bruneau BG: Directed transdifferentiation of mouse mesoderm to heart tissue by defined factors. Nature 2009, 459:708-711. www.sciencedirect.com
Chromatin regulation of reprogramming Luna-Zurita and Bruneau 561
41. Zaret KS, Carroll JS: Pioneer transcription factors: establishing competence for gene expression. Genes Dev 2011, 25: 2227-2241. 42. Albini S, Coutinho P, Malecova B, Giordani L, Savchenko A, Forcales SV, Puri PL: Epigenetic reprogramming of human embryonic stem cells into skeletal muscle cells and generation of contractile myospheres. Cell Rep 2013, 3: 661-670. 43. Singhal N, Graumann J, Wu G, Arauzo-Bravo MJ, Han DW, Greber B, Gentile L, Mann M, Scholer HR: Chromatin-remodeling components of the BAF complex facilitate reprogramming. Cell 2010, 141:943-955. In this work, the authors identified different components of the BAF chromatin-remodeling complex, able to increase the efficiency of cell reprogramming due to the activity of Oct4, Sox2, Klf4 and c-Myc in somatic cells. 44. Kleger A, Mahaddalkar PU, Katz SF, Lechel A, Joo JY, Loya K, Lin Q, Hartmann D, Liebau S, Kraus JM et al.: Increased reprogramming capacity of mouse liver progenitor cells, compared with differentiated liver cells, requires the BAF complex. Gastroenterology 2012, 142:907-917. 45. Isagawa T, Nagae G, Shiraki N, Fujita T, Sato N, Ishikawa S, Kume S, Aburatani H: DNA methylation profiling of embryonic stem cell differentiation into the three germ layers. PLoS ONE 2011, 6:e26052. 46. Hattori N, Nishino K, Ko YG, Hattori N, Ohgane J, Tanaka S, Shiota K: Epigenetic control of mouse Oct-4 gene expression in embryonic stem cells and trophoblast stem cells. J Biol Chem 2004, 279:17063-17069. 47. Hattori N, Imao Y, Nishino K, Hattori N, Ohgane J, Yagi S, Tanaka S, Shiota K: Epigenetic regulation of Nanog gene in embryonic stem and trophoblast stem cells. Genes Cells 2007, 12:387-396. 48. Doege CA, Inoue K, Yamashita T, Rhee DB, Travis S, Fujita R, Guarnieri P, Bhagat G, Vanti WB, Shih A et al.: Early-stage epigenetic modification during somatic cell reprogramming by Parp1 and Tet2. Nature 2012, 488:652-655. The authors of this work identified two factors, Parp1 and Tet2, as essentials for the OSKM-dependent somatic reprogramming. Parp1 and Tet2 interact with Nanog and Esrrb loci, promoting the transition between 5-methylcytosine (5mC), 5-hydroxyethylcytosine (5hmC) and cytosine (C). Further, the work establishes 5hmC as an epigenetic mark and not as a transition step during loci demethylation.
www.sciencedirect.com
49. Costa Y, Ding J, Theunissen TW, Faiola F, Hore TA, Shliaha PV, Fidalgo M, Saunders A, Lawrence M, Dietmann S et al.: NANOG dependent function of TET1 and TET2 in establishment of pluripotency. Nature 2013, 495:370-374. This work shows how NANOG, a transcription factor with a pivotal role in the maintaining of the pluripotency stage, interacts with the 5mC hydrolases Tet1 and Tet2 during somatic cell reprogramming, promoting the expression of pluripotency genes and increasing the efficiency of cell reprogramming. 50. Chen X, Xu H, Yuan P, Fang F, Huss M, Vega VB, Wong E, Orlov YL, Zhang W, Jiang J et al.: Integration of external signaling pathways with the core transcriptional network in embryonic stem cells. Cell 2008, 133:1106-1117. 51. Spruijt CG, Gnerlich F, Smits AH, Pfaffeneder T, Jansen PW, Bauer C, Munzel M, Wagner M, Muller M, Khan F et al.: Dynamic readers for 5-(hydroxy)methylcytosine and its oxidized derivatives. Cell 2013, 152:1146-1159. 52. Stadtfeld M, Apostolou E, Ferrari F, Choi J, Walsh RM, Chen T, Ooi SS, Kim SY, Bestor TH, Shioda T et al.: Ascorbic acid prevents loss of Dlk1-Dio3 imprinting and facilitates generation of all-iPS cell mice from terminally differentiated B cells. Nat Genet 2012, 44:S391-S392. 53. Raynal NJ, Si J, Taby RF, Gharibyan V, Ahmed S, Jelinek J, Estecio MR, Issa JP: DNA methylation does not stably lock gene expression but instead serves as a molecular mark for gene silencing memory. Cancer Res 2012, 72:1170-1181. 54. Pijnappel WW, Esch D, Baltissen MP, Wu G, Mischerikow N, Bergsma AJ, van der Wal E, Han DW, Bruch HV, Moritz S et al.: A central role for TFIID in the pluripotent transcription circuitry. Nature 2013, 495:516-519. 55. Wamstad JA, Alexander JM, Truty RM, Shrikumar A, Li F, Eilertson KE, Ding H, Wylie JN, Pico AR, Capra JA et al.: Dynamic and coordinated epigenetic regulation of developmental transitions in the cardiac lineage. Cell 2012, 151:206-220. 56. Beshiri ML, Holmes KB, Richter WF, Hess S, Islam AB, Yan Q, Plante L, Litovchick L, Gevry N, Lopez-Bigas N et al.: Coordinated repression of cell cycle genes by KDM5A and E2F4 during differentiation. Proc Natl Acad Sci U S A 2012, 109:18499-18504. 57. Li Z, Gadue P, Chen K, Jiao Y, Tuteja G, Schug J, Li W, Kaestner KH: Foxa2 and H2A.Z mediate nucleosome depletion during embryonic stem cell differentiation. Cell 2012, 151:1608-1616.
Current Opinion in Genetics & Development 2013, 23:556–561