Reprogramming towards pluripotency requires AID-dependent DNA demethylation (original) (raw)
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Stem Cells, the Molecular Circuitry of Pluripotency and Nuclear Reprogramming
Cell, 2008
Reprogramming of somatic cells to a pluripotent embryonic stem cell-like state has been achieved by nuclear transplantation of a somatic nucleus into an enucleated egg and most recently by introducing defined transcription factors into somatic cells. Nuclear reprogramming is of great medical interest, as it has the potential to generate a source of patient-specific cells. Here, we review strategies to reprogram somatic cells to a pluripotent embryonic state and discuss our understanding of the molecular mechanisms of reprogramming based on recent insights into the regulatory circuitry of the pluripotent state.
Stem cells, pluripotency and nuclear reprogramming
Journal of Thrombosis and Haemostasis, 2009
Reprogramming of somatic cells to a pluripotent embryonic stem cell-like state has been achieved by nuclear transplantation of a somatic nucleus into an enucleated egg and most recently by introducing defined transcription factors into somatic cells. Nuclear reprogramming is of great medical interest as it has the potential to generate a source of patientspecific cells. This short review summarizes strategies to reprogram somatic cells to a pluripotent embryonic state and discuss the implications of this technology for transplantation medicine.
Advancements in reprogramming strategies for the generation of induced pluripotent stem cells
Journal of Assisted Reproduction and Genetics, 2011
Direct reprogramming of somatic cells into induced pluripotent stem (iPS) cells has emerged as an invaluable method for generating patient-specific stem cells of any lineage without the use of embryonic materials. Following the first reported generation of iPS cells from murine fibroblasts using retroviral transduction of a defined set of transcription factors, various new strategies have been developed to improve and refine the reprogramming technology. Recent developments provide optimism that the generation of safe iPS cells without any genomic modification could be derived in the near future for the use in clinical settings. This review summarizes current and evolving strategies in the generation of iPS cells, including types of somatic cells for reprogramming, variations of reprogramming genes, reprogramming methods, and how the advancement iPS cells technology can lead to the future success of reproductive medicine.
Advances in Reprogramming Somatic Cells to Induced Pluripotent Stem Cells
Stem Cell Reviews and Reports, 2010
Traditionally, nuclear reprogramming of cells has been performed by transferring somatic cell nuclei into oocytes, by combining somatic and pluripotent cells together through cell fusion and through genetic integration of factors through somatic cell chromatin. All of these techniques changes gene expression which further leads to a change in cell fate. Here we discuss recent advances in generating induced pluripotent stem cells, different reprogramming methods and clinical applications of iPS cells. Viral vectors have been used to transfer transcription factors (Oct4, Sox2, c-myc, Klf4, and nanog) to induce reprogramming of mouse fibroblasts, neural stem cells, neural progenitor cells, keratinocytes, B lymphocytes and meningeal membrane cells towards pluripotency. Human fibroblasts, neural cells, blood and keratinocytes have also been reprogrammed towards pluripotency. In this review we have discussed the use of viral vectors for reprogramming both animal and human stem cells. Currently, many studies are also involved in finding alternatives to using viral vectors carrying transcription factors for reprogramming cells. These include using plasmid transfection, piggyback transposon system and piggyback transposon system combined with a non viral vector system. Applications of these techniques have been discussed in detail including its advantages and disadvantages. Finally, current clinical applications of induced pluripotent stem cells and its limitations have also been reviewed. Thus, this review is a summary of current research advances in reprogramming cells into induced pluripotent stem cells.
Deterministic direct reprogramming of somatic cells to pluripotency
Somatic cells can be inefficiently and stochastically reprogrammed into induced pluripotent stem (iPS) cells by exogenous expression of Oct4 (also called Pou5f1), Sox2, Klf4 and Myc (hereafter referred to as OSKM). The nature of the predominant rate-limiting barrier(s) preventing the majority of cells to successfully and synchronously reprogram remains to be defined. Here we show that depleting Mbd3, a core member of the Mbd3/NuRD (nucleosome remodelling and deacetylation) repressor complex, together with OSKM transduction and reprogramming in naive pluripotency promoting conditions, result in deterministic and synchronized iPS cell reprogramming (near 100% efficiency within seven days from mouse and human cells). Our findings uncover a dichotomous molecular function for the reprogramming factors, serving to reactivate endogenous pluripotency networks while simultaneously directly recruiting the Mbd3/NuRD repressor complex that potently restrains the reactivation of OSKM downstream target genes. Subsequently, the latter interactions, which are largely depleted during early pre-implantation development in vivo, lead to a stochastic and protracted reprogramming trajectory towards pluripotency in vitro. The deterministic reprogramming approach devised here offers a novel platform for the dissection of molecular dynamics leading to establishing pluripotency at unprecedented flexibility and resolution.
Transcriptional and epigenetic mechanisms of cellular reprogramming to induced pluripotency
Epigenomics, 2016
Enforced ectopic expression of a cocktail of pluripotency-associated genes such as Oct4, Sox2, Klf4 and c-Myc can reprogram somatic cells into induced pluripotent stem cells (iPSCs). The remarkable proliferation ability of iPSCs and their aptitude to redifferentiate into any cell lineage makes these cells a promising tool for generating a variety of human tissue in vitro. Yet, pluripotency induction is an inefficient process, as cells undergoing reprogramming need to overcome developmentally imposed epigenetic barriers. Recent work has shed new light on the molecular mechanisms that drive the reprogramming of somatic cells to iPSCs. Here, we present current knowledge on the transcriptional and epigenetic regulation of pluripotency induction and discuss how variability in epigenetic states impacts iPSCs' inherent biological properties.
Revista da Associacao Medica Brasileira (1992), 2017
Induced pluripotent stem cells (iPSCs) are somatic cells reprogrammed into an embryonic-like pluripotent state by the expression of specific transcription factors. iPSC technology is expected to revolutionize regenerative medicine in the near future. Despite the fact that these cells have the capacity to self-renew, they present low efficiency of reprogramming. Recent studies have demonstrated that the previous somatic epigenetic signature is a limiting factor in iPSC performance. Indeed, the process of effective reprogramming involves a complete remodeling of the existing somatic epigenetic memory, followed by the establishment of a "new epigenetic signature" that complies with the new type of cell to be differentiated. Therefore, further investigations of epigenetic modifications associated with iPSC reprogramming are required in an attempt to improve their self-renew capacity and potency, as well as their application in regenerative medicine, with a new strategy to redu...
Bressan et al SBTE 2011 Reprogramming somatic cells trough gene induction and nuclear reprogramming
The understanding of nuclear reprogramming pathways provides important contributions to applied and basic sciences such as the development of autologous cellular therapies for the treatment of numerous diseases, the improved efficiency of animal-based biotechnology or the generation of functional gametes in vitro. Strategies such as nuclear transfer and induced reprogramming have been used to induce somatic cells into an embryonic-like pluripotent state. Both techniques have been routinely performed worldwide, and live offspring have been successfully derived from them, resulting in a proof of efficacy of both techniques. Detailed studies on cellular and molecular mechanisms that mediate reprogramming, however, still require further investigation to develop practical applications in veterinary and human medicine. Review: Studies on cell reprogramming, differentiation and proliferation have revealed that a core of transcription factors, as for example, OCT4, SOX2 and NANOG, act together promoting cell commitment or pluripotency. Mechanisms of induced reprogramming by pluripotency-related transcription factors forced expression or nuclear transfer seems to be mediated by the same pathways observed in fertilization, eliciting nuclear remodeling and modulating gene expression. However, abnormal chromatin conformation, often leading to disrupted imprinting and atypical gene expression patterns are frequently observed on in vitro reprogramming. Strategies used to facilitate nuclear remodeling, such as chromatin modifying agents, as for example, histone deacetilases inhibitors or DNA methyltransferases; or chemicals responsible for the inhibition of developmentrelated pathways, as for example, MEK and GSK3 inhibitors, when used in the in vitro culture of cells or embryos, have proved to favors transcriptional regulation and improve reprogramming. Such alternatives are highly prone to enable the routine use of in vitro reprogramming in animal production and medical sciences, for example, by promoting the generation of functional male and female functional gametes capable of producing viable offspring. Thus, the properties, deficiencies and implications of induced reprogramming and nuclear transfer techniques in somatic cells were discussed in this review, as well as its probable outcomes.
Sequential Expression of Pluripotency Markers during Direct Reprogramming of Mouse Somatic Cells
Cell Stem Cell, 2008
Pluripotency can be induced in differentiated murine and human cells by retroviral transduction of Oct4, Sox2, Klf4 and c-Myc. We have devised a reprogramming strategy in which these four transcription factors are expressed from doxycycline (dox) inducible lentiviral vectors. Using these inducible constructs, we derived induced pluripotent stem (iPS) cells from mouse embryonic fibroblasts (MEFs) and found that transgene silencing is a prerequisite for normal cell differentiation. We have analyzed the timing of known pluripotency marker activation during mouse iPS cell derivation and observed that alkaline phosphatase (AP) was activated first, followed by stage specific embryonic antigen 1 (SSEA1). Expression of Nanog and the endogenous Oct4 gene, marking fully reprogrammed cells, were only observed late in the process. Importantly, the virally transduced cDNAs needed to be expressed for at least 12 days in order to generate iPS cells. Our results are a step towards understanding some of the molecular events governing epigenetic reprogramming.