Adenosine-to-inosine (A-to-I) RNA editing is a post-transcriptional, site-specific modification process that is catalyzed by Adenosine Deaminase Acting on RNA (ADAR) gene family members. Since ADARs act on double-stranded RNA, most A-to-I editing occurs within repetitive elements, particularly Alu elements, as the result of the inherent property of these sequences to fold and form double strands. ADAR1-mediated A-to-I RNA editing was recently implicated in the regulation of human embryonic stem cells (hESCs). Spontaneous and neuronal differentiation of hESC was shown to result in a decrease in A-to-I editing levels. Knockdown of ADAR1 in hESCs results in an elevation of the expression of differentiation-related genes. In addition, we found that hESCs over-expressing ADAR1 could not be generated. The current study shows that the editing levels of induced pluripotent stem cells (iPSCs) change throughout reprogramming, from a source cell level to a level similar to that of hESCs. Up- or down-regulation of the ADAR1 level in human foreskin fibroblast (HFF) cells before induction of reprogramming results in varied reprogramming efficiencies. Furthermore, HFF-iPSC early clones derived from source cells in which the ADAR1 level was down-regulated lose their iPSC properties shortly after iPSC colony formation and instead exhibit characteristics of cancer cells. Taken together, our results imply a role for ADAR1 in the regulation of pluripotency induction as well as in the maintenance of early iPSC properties.
Get full access to this article
View all access options for this article.
References
1.
BassBL. (1997). RNA editing and hypermutation by adenosine deamination. Trends Biochem Sci, 22:157–162.
2.
BassBL. (2002). RNA editing by adenosine deaminases that act on RNA. Annu Rev Biochem, 71:817–846.
3.
MelcherT, MaasS, HerbA, SprengelR, SeeburgPH and HiguchiM. (1996). A mammalian RNA editing enzyme. Nature, 379:460–464.
4.
LaiF, ChenCX, CarterKC and NishikuraK. (1997). Editing of glutamate receptor B subunit ion channel RNAs by four alternatively spliced DRADA2 double-stranded RNA adenosine deaminases. Mol Cell Biol, 17:2413–2424.
5.
ChenCX, ChoDS, WangQ, LaiF, CarterKC and NishikuraK. (2000). A third member of the RNA-specific adenosine deaminase gene family, ADAR3, contains both single- and double-stranded RNA binding domains. RNA, 6:755–767.
6.
PattersonJB and SamuelCE. (1995). Expression and regulation by interferon of a double-stranded-RNA-specific adenosine deaminase from human cells: evidence for two forms of the deaminase. Mol Cell Biol, 15:5376–5388.
7.
PalladinoMJ, KeeganLP, O'ConnellMA and ReenanRA. (2000). A-to-I pre-mRNA editing in Drosophila is primarily involved in adult nervous system function and integrity. Cell, 102:437–449.
8.
WangQ, MiyakodaM, YangW, KhillanJ, StachuraDL, WeissMJ and NishikuraK. (2004). Stress-induced apoptosis associated with null mutation of ADAR1 RNA editing deaminase gene. J Biol Chem, 279:4952–4961.
9.
HartnerJC, SchmittwolfC, KispertA, MullerAM, HiguchiM and SeeburgPH. (2004). Liver disintegration in the mouse embryo caused by deficiency in the RNA-editing enzyme ADAR1. J Biol Chem, 279:4894–4902.
10.
HiguchiM, MaasS, SingleFN, HartnerJ, RozovA, BurnashevN, FeldmeyerD, SprengelR and SeeburgPH. (2000). Point mutation in an AMPA receptor gene rescues lethality in mice deficient in the RNA-editing enzyme ADAR2. Nature, 406:78–81.
11.
KawaharaY, ItoK, SunH, AizawaH, KanazawaI and KwakS. (2004). Glutamate receptors: RNA editing and death of motor neurons. Nature, 427:801
12.
AkbarianS, SmithMA and JonesEG. (1995). Editing for an AMPA receptor subunit RNA in prefrontal cortex and striatum in Alzheimer's disease, Huntington's disease and schizophrenia. Brain Res, 699:297–304.
13.
GurevichI, TamirH, ArangoV, DworkAJ, MannJJ and SchmaussC. (2002). Altered editing of serotonin 2C receptor pre-mRNA in the prefrontal cortex of depressed suicide victims. Neuron, 34:349–356.
14.
LaxminarayanaD, O'RourkeKS, MaasS and OlorenshawI. (2007). Altered editing in RNA editing adenosine deaminase ADAR2 gene transcripts of systemic lupus erythematosus T lymphocytes. Immunology, 121:359–369.
15.
AthanasiadisA, RichA and MaasS. (2004). Widespread A-to-I RNA editing of Alu-containing mRNAs in the human transcriptome. PLoS Biol, 2:e391
16.
LevanonEY, EisenbergE, YelinR, NemzerS, HalleggerM, ShemeshR, FligelmanZY, ShoshanA, PollockSR, et al. (2004). Systematic identification of abundant A-to-I editing sites in the human transcriptome. Nat Biotechnol, 22:1001–1005.
17.
KimDD, KimTT, WalshT, KobayashiY, MatiseTC, BuyskeS and GabrielA. (2004). Widespread RNA editing of embedded alu elements in the human transcriptome. Genome Res, 14:1719–1725.
18.
OsenbergS, DominissiniD, RechaviG and EisenbergE. (2009). Widespread cleavage of A-to-I hyperediting substrates. RNA, 15:1632–1639.
19.
BlowMJ, GrocockRJ, van DongenS, EnrightAJ, DicksE, FutrealPA, WoosterR and StrattonMR. (2006). RNA editing of human microRNAs. Genome Biol, 7:R27
20.
PrasanthKV, PrasanthSG, XuanZ, HearnS, FreierSM, BennettCF, ZhangMQ and SpectorDL. (2005). Regulating gene expression through RNA nuclear retention. Cell, 123:249–263.
21.
WangQ, ZhangZ, BlackwellK and CarmichaelGG. (2005). Vigilins bind to promiscuously A-to-I-edited RNAs and are involved in the formation of heterochromatin. Curr Biol, 15:384–391.
22.
PazN, LevanonEY, AmariglioN, HeimbergerAB, RamZ, ConstantiniS, BarbashZS, AdamskyK, SafranM, et al. (2007). Altered adenosine-to-inosine RNA editing in human cancer. Genome Res, 17:1586–1595.
23.
CenciC, BarzottiR, GaleanoF, CorbelliS, RotaR, MassimiL, Di RoccoC, O'ConnellMA and GalloA. (2008). Down-regulation of RNA editing in pediatric astrocytomas: ADAR2 editing activity inhibits cell migration and proliferation. J Biol Chem, 283:7251–7260.
24.
ThomsonJA, Itskovitz-EldorJ, ShapiroSS, WaknitzMA, SwiergielJJ, MarshallVS and JonesJM. (1998). Embryonic stem cell lines derived from human blastocysts. Science, 282:1145–1147.
25.
YoungRA. (2011). Control of the embryonic stem cell state. Cell, 144:940–954.
26.
ChenLL and CarmichaelGG. (2009). Altered nuclear retention of mRNAs containing inverted repeats in human embryonic stem cells: functional role of a nuclear noncoding RNA. Mol Cell, 35:467–478.
27.
OsenbergS, Paz YaacovN, SafranM, MoshkovitzS, ShtrichmanR, SherfO, Jacob-HirschJ, KeshetG, AmariglioN, Itskovitz-EldorJ and RechaviG. (2010). Alu sequences in undifferentiated human embryonic stem cells display high levels of A-to-I RNA editing. PLoS One, 5:e11173
28.
ShtrichmanR, GermanguzI, MandelR, ZiskindA, NahorI, SafranM, OsenbergS, SherfO, RechaviG and Itskovitz-EldorJ (2012). Altered A-to-I RNA editing in human embryogenesis. PLoS One, 7:e41576
29.
TakahashiK, TanabeK, OhnukiM, NaritaM, IchisakaT, TomodaK and YamanakaS. (2007). Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell, 131:861–872.
30.
AmitM, CarpenterMK, InokumaMS, ChiuCP, HarrisCP, WaknitzMA, Itskovitz-EldorJ and ThomsonJA. (2000). Clonally derived human embryonic stem cell lines maintain pluripotency and proliferative potential for prolonged periods of culture. Dev Biol, 227:271–278.
31.
BrummelkampTR, BernardsR and AgamiR. (2002). A system for stable expression of short interfering RNAs in mammalian cells. Science, 296:550–553.
32.
NovakA, ShtrichmanR, GermanguzI, SegevH, Zeevi-LevinN, FishmanB, MandelYE, BaradL, DomevH, et al. (2010). Enhanced reprogramming and cardiac differentiation of human keratinocytes derived from plucked hair follicles, using a single excisable lentivirus. Cell Reprogram, 12:665–678.
33.
Koren-MichowitzM, ShimoniA, VivanteA, TrakhtenbrotL, RechaviG, AmariglioN, LoewenthalR, NaglerA and CohenY. (2008). A new MALDI-TOF-based assay for monitoring JAK2 V617F mutation level in patients undergoing allogeneic stem cell transplantation (allo SCT) for classic myeloproliferative disorders (MPD). Leuk Res, 32:421–427.
34.
HoR, ChronisC and PlathK. (2011). Mechanistic insights into reprogramming to induced pluripotency. J Cell Physiol, 226:868–878.
35.
GermanguzI, SedanO, Zeevi-LevinN, ShtrichmanR, BarakE, ZiskindA, EliyahuS, MeiryG, AmitM, Itskovitz-EldorJ and BinahO. (2011). Molecular characterization and functional properties of cardiomyocytes derived from human inducible pluripotent stem cells. J Cell Mol Med, 15:38–51.
36.
ViswanathanSR, DaleyGQ and GregoryRI. (2008). Selective blockade of microRNA processing by Lin28. Science, 320:97–100.
37.
NicholsJ, ZevnikB, AnastassiadisK, NiwaH, Klewe-NebeniusD, ChambersI, ScholerH and SmithA. (1998). Formation of pluripotent stem cells in the mammalian embryo depends on the POU transcription factor Oct4. Cell, 95:379–391.
LowryWE, RichterL, YachechkoR, PyleAD, TchieuJ, SridharanR, ClarkAT and PlathK. (2008). Generation of human induced pluripotent stem cells from dermal fibroblasts. Proc Natl Acad Sci U S A, 105:2883–2888.
40.
KurodaT, YasudaS, KusakawaS, HirataN, KandaY, SuzukiK, TakahashiM, NishikawaS, KawamataS and SatoY. (2012). Highly sensitive in vitro methods for detection of residual undifferentiated cells in retinal pigment epithelial cells derived from human iPS cells. PLoS One, 7:e37342
41.
ProkhorovaTA, HarknessLM, FrandsenU, DitzelN, SchroderHD, BurnsJS and KassemM. (2009). Teratoma formation by human embryonic stem cells is site dependent and enhanced by the presence of Matrigel. Stem Cells Dev, 18:47–54.
42.
LiJB, LevanonEY, YoonJK, AachJ, XieB, LeproustE, ZhangK, GaoY and ChurchGM. (2009). Genome-wide identification of human RNA editing sites by parallel DNA capturing and sequencing. Science, 324:1210–1213.
43.
JuYS, KimJI, KimS, HongD, ParkH, ShinJY, LeeS, LeeWC, YuSB, et al. (2011). Extensive genomic and transcriptional diversity identified through massively parallel DNA and RNA sequencing of eighteen Korean individuals. Nat Genet, 43:745–752.
44.
GreenbergerS, LevanonEY, Paz-YaacovN, BarzilaiA, SafranM, OsenbergS, AmariglioN, RechaviG and EisenbergE. (2010). Consistent levels of A-to-I RNA editing across individuals in coding sequences and non-conserved Alu repeats. BMC Genomics, 11:608
45.
MaheraliN, SridharanR, XieW, UtikalJ, EminliS, ArnoldK, StadtfeldM, YachechkoR, TchieuJ, et al. (2007). Directly reprogrammed fibroblasts show global epigenetic remodeling and widespread tissue contribution. Cell Stem Cell, 1:55–70.
46.
WernigM, MeissnerA, ForemanR, BrambrinkT, KuM, HochedlingerK, BernsteinBE and JaenischR. (2007). In vitro reprogramming of fibroblasts into a pluripotent ES-cell-like state. Nature, 448:318–324.
47.
ChinMH, MasonMJ, XieW, VoliniaS, SingerM, PetersonC, AmbartsumyanG, AimiuwuO, RichterL, et al. (2009). Induced pluripotent stem cells and embryonic stem cells are distinguished by gene expression signatures. Cell Stem Cell, 5:111–123.
48.
PoloJM, LiuS, FigueroaME, KulalertW, EminliS, TanKY, ApostolouE, StadtfeldM, LiY, et al. (2010). Cell type of origin influences the molecular and functional properties of mouse induced pluripotent stem cells. Nat Biotechnol, 28:848–855.
49.
DoiA, ParkIH, WenB, MurakamiP, AryeeMJ, IrizarryR, HerbB, Ladd-AcostaC, RhoJ, et al. (2009). Differential methylation of tissue- and cancer-specific CpG island shores distinguishes human induced pluripotent stem cells, embryonic stem cells and fibroblasts. Nat Genet, 41:1350–1353.
SmithZD, NachmanI, RegevA and MeissnerA. (2010). Dynamic single-cell imaging of direct reprogramming reveals an early specifying event. Nat Biotechnol, 28:521–526.
52.
RuizS, PanopoulosAD, HerreriasA, BissigKD, LutzM, BerggrenWT, VermaIM andIzpisua BelmonteJC (2011). A high proliferation rate is required for cell reprogramming and maintenance of human embryonic stem cell identity. Curr Biol, 21:45–52.
53.
YanC and BoydDD. (2006). Histone H3 acetylation and H3 K4 methylation define distinct chromatin regions permissive for transgene expression. Mol Cell Biol, 26:6357–6371.
54.
HartnerJC, WalkleyCR, LuJ and OrkinSH. (2009). ADAR1 is essential for the maintenance of hematopoiesis and suppression of interferon signaling. Nat Immunol, 10:109–115.
55.
EisenbergE, NemzerS, KinarY, SorekR, RechaviG and LevanonEY. (2005). Is abundant A-to-I RNA editing primate-specific?. Trends Genet, 21:77–81.
56.
ReyaT, MorrisonSJ, ClarkeMF and WeissmanIL. (2001). Stem cells, cancer, and cancer stem cells. Nature, 414:105–111.
57.
WongDJ, LiuH, RidkyTW, CassarinoD, SegalE and ChangHY. (2008). Module map of stem cell genes guides creation of epithelial cancer stem cells. Cell Stem Cell, 2:333–344.
58.
UtikalJ, PoloJM, StadtfeldM, MaheraliN, KulalertW, WalshRM, KhalilA, RheinwaldJG and HochedlingerK. (2009). Immortalization eliminates a roadblock during cellular reprogramming into iPS cells. Nature, 460:1145–1148.
59.
KawamuraT, SuzukiJ, WangYV, MenendezS, MoreraLB, RayaA, WahlGM and BelmonteJC. (2009). Linking the p53 tumour suppressor pathway to somatic cell reprogramming. Nature, 460:1140–1144.
60.
RiggsJW, BarrilleauxBL, VarlakhanovaN, BushKM, ChanV and KnoepflerPS. (2013). Induced pluripotency and oncogenic transformation are related processes. Stem Cells Dev, 22:37–50.
61.
HealeBS, KeeganLP, McGurkL, MichlewskiG, BrindleJ, StantonCM, CaceresJF and O'ConnellMA (2009). Editing independent effects of ADARs on the miRNA/siRNA pathways. EMBO J, 28:3145–3156.
62.
NieY, DingL, KaoPN, BraunR and YangJH. (2005). ADAR1 interacts with NF90 through double-stranded RNA and regulates NF90-mediated gene expression independently of RNA editing. Mol Cell Biol, 25:6956–6963.
63.
OtaH, SakuraiM, GuptaR, ValenteL, WulffBE, AriyoshiK, IizasaH, DavuluriRV and NishikuraK. (2013). ADAR1 forms a complex with Dicer to promote microRNA processing and RNA-induced gene silencing. Cell, 153:575–589.
Supplementary Material
Please find the following supplemental material available below.
For Open Access articles published under a Creative Commons License, all supplemental material carries the same license as the article it is associated with.
For non-Open Access articles published, all supplemental material carries a non-exclusive license, and permission requests for re-use of supplemental material or any part of supplemental material shall be sent directly to the copyright owner as specified in the copyright notice associated with the article.
