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Enhanced Green Fluorescent Protein Gene under the Regulation of Human Oct4 Promoter as a Marker to Identify Reprogramming of Human Fibroblasts  

Heo, Soon-Young (Department of Physiology, Dankook University School of Medicine)
Ahn, Kwang-Sung (Department of Physiology, Dankook University School of Medicine)
Kang, Jee-Hyun (Department of Physiology, Dankook University School of Medicine)
Shim, Ho-Sup (Department of Physiology, Dankook University School of Medicine)
Publication Information
Reproductive and Developmental Biology / v.32, no.2, 2008 , pp. 135-140 More about this Journal
Abstract
Recent studies on nuclear transfer and induced pluripotent stem cells have demonstrated that differentiated somatic cells can be returned to the undifferentiated state by reversing their developmental process. These epigenetically reprogrammed somatic cells may again be differentiated into various cell types, and used for cell replacement therapies through autologous transplantation to treat many degenerative diseases. To date, however, reprogramming of somatic cells into undifferentiated cells has been extremely inefficient. Hence, reliable markers to identify the event of reprogramming would assist effective selection of reprogrammed cells. In this study, a transgene construct encoding enhanced green fluorescent protein (EGFP) under the regulation of human Oct4 promoter was developed as a reporter for the reprogramming of somatic cells. Microinjection of the transgene construct into pronuclei of fertilized mouse eggs resulted in the emission of green fluorescence, suggesting that the undifferentiated cytoplasmic environment provided by fertilized eggs induces the expression of EGFP. Next, the transgene construct was introduced into human embryonic fibroblasts, and the nuclei from these cells were transferred into enucleated porcine oocytes. Along with their in vitro development, nuclear transfer embryos emitted green fluorescence, suggesting the reprogramming of donor nuclei in nuclear transfer embryos. The results of the present study demonstrate that expression of the transgene under the regulation of human Oct4 promoter coincides with epigenetic reprogramming, and may be used as a convenient marker that non-invasively reflects reprogramming of somatic cells.
Keywords
EGFP; Pluripotency marker; Epigenetic reprogramming; Nuclear transfer;
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1 Chesne P, Adenot PG, Viglietta C, Baratte M, Boulanger L, Renard JP (2002): Cloned rabbits produced by nuclear transfer from adult somatic cells. Nat Biotechnol 20:366-369   DOI   ScienceOn
2 Cibelli JB, Campbell KH, Seidel GE, West MD, Lanza RP (2002): The health profile of cloned animals. Nat Biotechnol 20:13-14   DOI   ScienceOn
3 Collas P, Gammelsaeter R (2007): Novel approaches to epigenetic reprogramming of somatic cells. Cloning Stem Cells 9:26-32   DOI   ScienceOn
4 Lee BC, Kim MK, Jang G, Oh HJ, Yuda F, Kim HJ, Hossein MS, Kim JJ, Kang SK, Schatten G, Hwang WS (2005): Dogs cloned from adult somatic cells. Nature 436:641   DOI   ScienceOn
5 Niwa H, Miyazaki J, Smith AG (2000): Quantitative expression of Oct-3/4 defines differentiation, dedifferentiation or self-renewal of ES cells. Nat Genet 24:372-376   DOI   ScienceOn
6 Okita K, Ichisaka T, Yamanaka S (2007): Generation of germline-competent induced pluripotent stem cells. Nature 448:313-317   DOI   ScienceOn
7 Scholer HR, Ruppert S, Suzuki N, Chowdhury K, Gruss P (1990): New type of POU domain in germ line-specific protein Oct-4. Nature 344:435-439   DOI   ScienceOn
8 Shamblott MJ, Axelman J, Wang S, Bugg EM, Littlefield JW, Donovan PJ, Blumenthal PD, Huggins GR, Gearhart JD (1998): Derivation of pluripotent stem cells from cultured human primordial germ cells. Proc Natl Acad Sci USA 95:13726-13731
9 Shim H, Jun SH, Kim YH, Lim JM, Lee HC, Cha KY (2000): Reprogramming of human embryonic and somatic cell nuclei in bovine oocytes. Fert Steril 74 (3 Suppl. 1):S195
10 Shin T, Kraemer D, Pryor J, Liu L, Rugila J, Howe L, Buck S, Murphy K, Lyons L, Westhusin M (2002): A cat cloned by nuclear transplantation. Nature 415:859   DOI   ScienceOn
11 Thomson JA, Itskovitz-Eldor J, Shapiro SS, Waknitz MA, Swiergiel JJ, Marshall VS, Jones JM (1998): Embryonic stem cell lines derived from human blastocysts. Science 282:1145-1147   DOI
12 Yamanaka S (2007): Strategies and new developments in the generation of patient-specific pluripotent stem cells. Cell Stem Cell 1:39-49   DOI   ScienceOn
13 Wakayama T, Perry AC, Zuccotti M, Johnson KR, Yanagimachi R (1998): Full-term development of mice from enucleated oocytes injected with cumulus cell nuclei. Nature 394:369-374   DOI   ScienceOn
14 Wilmut I, Schnieke AE, McWhir J, Kind AJ, Campbell KHS (1997): Viable offspring derived from fetal and adult mammalian cells. Nature 385:810-813   DOI   ScienceOn
15 Takahashi K, Tanabe K, Ohnuki M, Narita M, Ichisaka T, Tomoda K, Yamanaka S (2007): Induction of pluripotent stem cells from adult human fibroblasts by defined factors. Cell 131:861-872   DOI   ScienceOn
16 Yang X, Smith SL, Tian XC, Lewin HA, Renard JP, Wakayama T (2007): Nuclear reprogramming of cloned embryos and its implications for therapeutic cloning. Nat Genet 39:295-302   DOI   ScienceOn
17 Polejaeva IA, Chen SH, Vaught TD, Page RL, Mullins J, Ball S, Dai Y, Boone J, Walker S, Ayares DL, Colman A, Campbell KHS (2000): Cloned pigs produced by nuclear transfer from adult somatic cells. Nature 407:86-90   DOI   ScienceOn
18 Wernig M, Meissner A, Foreman R, Brambrink T, Ku M, Hochedlinger K, Bernstein BE, Jaenisch R (2007): In vitro reprogramming of fibroblasts into a pluripotent ES-cell-like state. Nature 448:318-324   DOI   ScienceOn
19 Galli C, Lagutina I, Crotti G, Colleoni S, Turini P, Ponderato N, Duchi R, Lazzari G (2003): Pregnancy: a cloned horse born to its dam twin. Nature 424:635
20 Mitsui K, Tokuzawa Y, Itoh H, Segawa K, Murakami M, Takahashi K, Maruyama M, Maeda M, Yamanaka S (2003): The homeoprotein Nanog is required for maintenance of pluripotency in mouse epiblast and ES cells. Cell 113:631-642   DOI   ScienceOn
21 Park IH, Zhao R, West JA, Yabuuchi A, Huo H, Ince TA, Lerou PH, Lensch MW, Daley GQ (2008): Reprogramming of human somatic cells to pluripotency with defined factors. Nature 451:141-146   DOI   ScienceOn
22 Takahashi K, Yamanaka S (2006): Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell 126: 663-676   DOI   ScienceOn
23 Cibelli JB, Stice SL, Golueke PJ, Kane JJ, Jerry J, Blackwell C, Ponce de Leon FA, Robl JM (1998): Cloned transgenic calves produced from nonquiescent fetal fibroblasts. Science 280:1256-1258   DOI   ScienceOn
24 Yang HM, Do HJ, Oh JH, Kim JH, Choi SY, Cha KY, Chung HM, Kim JH (2005): Characterization of putative cis-regulatory elements that control the transcriptional activity of the human Oct4 promoter. J Cell Biochem 96:821-830   DOI   ScienceOn
25 Yeom YI, Fuhrmann G, Ovitt CE, Brehm A, Ohbo K, Gross M, Hübner K, Scholer HR. (1996): Germline regulatory element of Oct-4 specific for the totipotent cycle of embryonal cells. Development 122: 881-894
26 Okamoto K, Okazawa H, Okuda A, Sakai M, Muramatsu M, Hamada H (1990): A novel octamer binding transcription factor is differentially expressed in mouse embryonic cells. Cell 60L:461-472
27 Yu J, Vodyanik MA, Smuga-Otto K, Antosiewicz- Bourget J, Frane JL, Tian S, Nie J, Jonsdottir GA, Ruotti V, Stewart R, Slukvin II, Thomson JA (2007): Induced pluripotent stem cell lines derived from human somatic cells. Science 318:1917-1920   DOI   ScienceOn
28 Baguisi A, Behboodi E, Melican DT, Pollock JS, Destrempes MM, Cammuso C, Williams JL, Nims SD, Porter CA, Midura P, Palacios MJ, Ayres SL, Denniston RS, Hayes ML, Ziomek CA, Meade HM, Godke RA, Gavin WG, Overstrom EW, Echelard Y (1999): Production of goats by somatic cell nuclear transfer. Nat Biotechnol 17:456-461   DOI   ScienceOn
29 Chambers I, Colby D, Robertson M, Nichols J, Lee S, Tweedie S, Smith A (2003): Functional expression cloning of Nanog, a pluripotency sustaining factor in embryonic stem cells. Cell 113:643-655   DOI   ScienceOn
30 Lowry WE, Richter L, Yachechko R, Pyle AD, Tchieu J, Sridharan R, Clark AT, Plath K (2008): Generation of human induced pluripotent stem cells from dermal fibroblasts. Proc Natl Acad Sci USA 105:2883-2888
31 Rosner MH, Vigano MA, Ozato K, Timmons PM, Poirier F, Rigby PW, Staudt LM (1990): A POUdomain transcription factor in early stem cells and germ cells of the mammalian embryo. Nature 345: 686-692   DOI   ScienceOn
32 Pesce M, Scholer HR (2001): Oct-4: gatekeeper in the beginnings of mammalian development. Stem Cells 19:271-278   DOI   ScienceOn
33 Zhou Q, Renard JP, Le Friec G, Brochard V, Beaujean N, Cherifi Y, Fraichard A, Cozzi J (2003): Generation of fertile cloned rats by regulating oocyte activation. Science 302:1179   DOI   ScienceOn
34 Ahn KS, Won JY, Heo SY, Kang JH, Yang HS, Shim H (2007): Transgenesis and nuclear transfer using porcine embryonic germ cells. Cloning Stem Cells 9:461-468   DOI   ScienceOn