Role of the Murine Reprogramming Factors in the Induction of Pluripotency

Sridharan, Rupa; Tchieu, Jason; Mason, Mike J.; Yachechko, Robin; Kuoy, Edward; Horvath, Steve; Zhou, Qing; Plath, Kathrin

doi:10.1016/j.cell.2009.01.001

Cited by 600 publications

(800 citation statements)

References 27 publications

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“…This unexpected finding was consistent with our hypothesis that a stable intermediate phase between fibroblasts and iPSCs could be achieved through the expression of only three reprogramming factors (Oct4, Klf4, and c-Myc). In fact, similar colonies were present in the original discovery [24,32], although subsequently identified differences in potential for reprogramming set these apart [21][22][23].…”

Section: Confirmation Of Two-phase Ipsc Inductionmentioning

confidence: 79%

“…The same genome-wide mapping method has been applied to study the function of Yamanaka factors among ESCs, iPSCs, and what have been termed ''partially reprogrammed cells.'' The last group of cells have ES-like morphology without ES-like pluripotency, following transduction of all four Yamanaka factors [21][22][23]. Examination of these cells can enhance our understanding of reprogramming barriers, but identified molecular mechanisms from comparison of these partially reprogrammed Author Contributions: Z.L.…”

Section: Introductionmentioning

confidence: 99%

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Two-Phase Analysis of Molecular Pathways Underlying Induced Pluripotent Stem Cell Induction

Lin

Perez

Lei³

et al. 2011

Stem Cells

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Induced pluripotent stem cells (iPSCs) can be reprogrammed from adult somatic cells by transduction with Oct4, Sox2, Klf4, and c-Myc, but the molecular cascades initiated by these factors remain poorly understood. Impeding their elucidation is the stochastic nature of the iPS induction process, which results in heterogeneous cell populations. Here we have synchronized the reprogramming process by a two-phase induction: an initial stable intermediate phase following transduction with Oct4, Klf4, and c-Myc, and a final iPS phase following overexpression of Sox2. This approach has enabled us to examine temporal gene expression profiles, permitting the identification of Sox2 downstream genes critical for induction. Furthermore, we have validated the feasibility of our new approach by using it to confirm that downregulation of transforming growth factor b signaling by Sox2 proves essential to the reprogramming process. Thus, we present a novel means for dissecting the details underlying the induction of iPSCs, an approach with significant utility in this arena and the potential for wide-ranging implications in the study of other reprogramming mechanisms. STEM

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Section: Confirmation Of Two-phase Ipsc Inductionmentioning

confidence: 79%

Section: Introductionmentioning

confidence: 99%

Two-Phase Analysis of Molecular Pathways Underlying Induced Pluripotent Stem Cell Induction

Lin

Perez

Lei³

et al. 2011

Stem Cells

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“…ChIP-on-Chip studies indicate that common targets of Klf4, Oct4, and Sox2 are most differentially bound between fully reprogrammed and partially reprogrammed iPS cells, compared with other promoters bound by only one or two of them [19]. This interesting phenomenon suggests that binding of target promoters by Oct4, Sox2, and Klf4 may not be through individual binding but may require assembly of a functional complex containing all three factors.…”

Section: Discussionmentioning

confidence: 99%

Klf4 Interacts Directly with Oct4 and Sox2 to Promote Reprogramming

et al. 2009

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Somatic cells can be reprogrammed to induced pluripotent stem (iPS) cells by ectopic expression of specific sets of transcription factors. Oct4, Sox2, and Klf4, factors that share many target genes in embryonic stem (ES) cells, are critical components in various reprogramming protocols. Nevertheless, it remains unclear whether these factors function together or separately in reprogramming. Here we show that Klf4 interacts directly with Oct4 and Sox2 when expressed at levels sufficient to induce iPS cells. Endogenous Klf4 also interacts with Oct4 and Sox2 in iPS cells and in mouse ES cells. The Klf4 C terminus, which contains three tandem zinc fingers, is critical for this interaction and is required for activation of the target gene Nanog. In addition, Klf4 and Oct4 co-occupy the Nanog promoter. A dominant negative mutant of Klf4 can compete with wild-type Klf4 to form defective Oct4/Sox2/ Klf4 complexes and strongly inhibit reprogramming. In the absence of Klf4 overexpression, interaction of endogenous Klf4 with Oct4/Sox2 is also required for reprogramming. This study supports the idea that direct interactions between Klf4, Oct4, and Sox2 are critical for somatic cell reprogramming.

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“…For example, efficient downregulation of somatic gene expression was observed, as was widespread posttranslational modification of histones on the regulatory regions of many of the target genes of Yamanaka factors [59][60][61] .…”

Section: Deterministic or Stochastic?mentioning

confidence: 99%

Mechanisms of nuclear reprogramming by eggs and oocytes: a deterministic process?

Jullien

Pasque²,

Halley-Stott³

et al. 2011

Nat Rev Mol Cell Biol

106

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Differentiated cells can be experimentally reprogrammed back to pluripotency by nuclear transfer, cell fusion or induced pluripotent stem cell technology. Nuclear transfer and cell fusion can lead to efficient reprogramming of gene expression. The egg and oocyte reprogramming process includes the exchange of somatic proteins for oocyte proteins, the post-translational modification of histones and the demethylation of DNA. These events occur in an ordered manner and on a defined timescale, indicating that reprogramming by nuclear transfer and by cell fusion rely on deterministic processes.The remarkable stability of cell differentiation under normal conditions can be reversed experimentally by nuclear transfer, cell fusion and induced pluripotent stem (iPS) cell technology [1][2][3][4][5] . This provides an opportunity to generate pluripotent embryonic cells from adult cells of the same individual and hence opens the possibilit y of cell replacement without the need for immunosuppression.iPS cell technology makes use of the overexpression of transcription factors (FIG. 1a) and has been extensively reviewed, so it is not discussed in detail [6][7][8][9] . To generate entirely unrelated cell types by iPS cell technology, treated cells must be grown for multiple cell divisions over a long period of time (up to 3 weeks). By contrast, nuclear transfer and cell fusion do not involve the overexpression of new genes, and instead make use of natura components present in eggs and some early embryos to initiate new transcription.There are two kinds of nuclear transfer experiments: egg-NT involves the transfer of a single somatic nucleus to an unfertilize d enucleated egg (in both mammals and amphibians) 1 (FIG. 1b); and ooc-NT involves the transplantation of multiple somatic cell nuclei into the germinal vesicle (the nucleus) of a growing meiotic prophase amphibian oocyte (an immature egg) 10 (FIG. 1c). Note that the terms egg and oocyte refer to different developmental stages in amphibians and mammals: amphibian eggs are in metaphase II of meiosis, which is equivalent to mouse metaphase II stage oocytes, whereas their immediate precursors, oocytes, are blocked in meiotic prophase I, which is equivalent to mouse germinal vesicle stage oocytes.© 2011 Macmillan Publishers Limited. All rights reserved Correspondence to J.B.G.j.gurdon@gurdon.cam.ac.uk. Competing interests statementThe authors declare no competing financial interests. There are important differences between the two types of nuclear transfer experimen t. Extensive cell division takes place in egg-NT experiments, and functional new cell types appear as the nuclear transplant embryo develops. By contrast, in ooc-NT experiments, no new cell types are formed, and neither the oocyte nor the introduced nuclei divide, but there is a direct transition from nuclei of differentiated cells to reprogrammed nuclei that transcribe pluripotency genes. Analysis of the mechanism of reprogramming (which involves transcription of pluripotency and other genes) in egg-NT expe...

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Role of the Murine Reprogramming Factors in the Induction of Pluripotency

Cited by 600 publications

References 27 publications

Two-Phase Analysis of Molecular Pathways Underlying Induced Pluripotent Stem Cell Induction

Two-Phase Analysis of Molecular Pathways Underlying Induced Pluripotent Stem Cell Induction

Klf4 Interacts Directly with Oct4 and Sox2 to Promote Reprogramming

Mechanisms of nuclear reprogramming by eggs and oocytes: a deterministic process?

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