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Author(s): Alejandro De Los Angeles [1, 2, 3]; Francesco Ferrari [4]; Yuko Fujiwara [1, 2, 3]; Ronald Mathieu [5]; Soohyun Lee [4]; Semin Lee [4]; Ho-Chou Tu [1, 2, 3]; [...]

Mouse embryonic stem (ES) cells are isolated from the inner cell mass of blastocysts, and can be preserved in vitro in a naive innercell-mass-like configuration by providing exogenous stimulation with leukaemia inhibitory factor (LIF) and small molecule inhibition of ERK1/ERK2 and GSKβ signalling (termed 2i/LIF conditions) (1,2). Hallmarks of naive pluripotency include driving Oct4 (also known as Pou5f1) transcription by its distal enhancer, retaining a pre-inactivation X chromosome state, and global reduction in DNA methylation and in H3K27me3 repressive chromatin mark deposition on developmental regulatory gene promoters (3). Upon withdrawal of 2i/LIF, naive mouse ES cells can drift towards a primed pluripotent state resembling that of the post-implantation epiblast (4). Although human ES cells share several molecular features with naive mouse ES cells (5), they also share a variety of epigenetic properties with primed murine epiblast stem cells (EpiSCs) (6,7). These include predominant use of the proximal enhancer element to maintain OCT4 expression, pronounced tendency for X chromosome inactivation in most female human ES cells, increase in DNA methylation and prominent deposition of H3K27me3 and bivalent domain acquisition on lineage regulatory genes (7). The feasibility of establishing human ground state naive pluripotency in vitro with equivalent molecular and functional features to those characterized in mouse ES cells remains to be defined (1). Here we establish defined conditions that facilitate the derivation of genetically unmodified human naive pluripotent stem cells from already established primed human ES cells, from somatic cells through induced pluripotent stem (iPS) cell reprogramming or directly from blastocysts. The novel naive pluripotent cells validated herein retain molecular characteristics and functional properties that are highly similar to mouse naive ES cells, and distinct from conventional primed human pluripotent cells. This includes competence in the generation of cross-species chimaeric mouse embryos that underwent organogenesis following microinjection of human naive iPS cells into mouse morulas. Collectively, our findings establish new avenues for regenerative medicine, patient-specific iPS cell disease modelling and the study of early human development invitro and invivo., Naive 2i/LIF conditions are not sufficient to elicit a self-renewal response in human ES cells/human iPS cells and result in their differentiation (7). Nevertheless, the proof of concept for the [...]

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. This study shows that the combination of naive pluripotency growth conditions, Oct4, Sox2, Klf4 and Myc (OSKM) overexpression, and depleting the Mbd3/NuRD co-repressor results in deterministic and synchronized reprogramming to pluripotency. Efficient and near-complete conversion to iPS cells Somatic cells can be reprogrammed to pluripotency by expression of exogenous factors,classically Oct4, Sox2,Klf4 and c-Myc (OSKM). During reprogramming, only a fraction of the cells converts into induced pluripotent stem (iPS) cells. The nature of rate limiting barrier(s) that prevent the majority of cells to convert into iPS cells remains elusive and it is unknown whether iPS cell reprogramming can be rendered deterministic and very efficient. Jacob Hanna and colleagues now show that the combination of 2i/LIF growth conditions, OSKM overexpression and neutralizing the Mbd3/NURD co-repressor results in deterministic and synchronized reprogramming to pluripotency. Using this approach, they found that almost 100% of mouse and human somatic cells convert into naive iPS cells after only seven days of OSKM induction., Author(s): Yoach Rais [sup.1] , Asaf Zviran [sup.1] , Shay Geula [sup.1] , Ohad Gafni [sup.1] , Elad Chomsky [sup.1] , Sergey Viukov [sup.1] , Abed AlFatah Mansour [sup.1] , [...]

Induced pluripotent stem cells (iPSCs) can be derived from somatic cells by ectopic expression of different transcription factors, classically Oct4 (also known as Pou5f1), Sox2, Klf4 and Myc (abbreviated as [...]

There are two methods for reprogramming mature cells to pluripotent stem cells, which can give rise to all cells of the body. The first direct comparison of the methods reveals that both can cause subtle molecular defects. See Article p.177 Stem cell types compared This study compares the distinct genetic, epigenetic and transcriptional signatures of human pluripotent stem cells produced by somatic cell nuclear transfer (SCNT) with induced pluripotent stem (iPS) cells produced by transcription-factor-mediated reprogramming. Both cell types were produced from the same pool of somatic donor cells to ensure a genetic match. iPS cells and SCNT-derived embryonic stem cells contained comparable de novo copy number variations. The abnormalities previously reported -- residual DNA methylation patterns typical of parental somatic cells -- were observed as anticipated in the iPS cells, but not in the SCNT-derived cells. This suggests that human somatic cells can be faithfully reprogrammed to pluripotency by SCNT and therefore might be more appropriate than iPS cells for use in cell replacement therapies., Author(s): Vladislav Krupalnik [sup.1] , Jacob H. Hanna [sup.1] Author Affiliations: (1) Vladislav Krupalnik and Jacob H. Hanna are in the Department of Molecular Genetics, The Weizmann Institute of Science, [...]

There are two methods for reprogramming mature cells to pluripotent stem cells, which can give rise to all cells of the body. The first direct comparison of the methods reveals [...]

Author(s): Ohad Gafni; Leehee Weinberger; Abed AlFatah Mansour; Yair S. Manor; Elad Chomsky; Dalit Ben-Yosef; Yael Kalma; Sergey Viukov; Itay Maza; Asaf Zviran; Yoach Rais; Zohar Shipony; Zohar Mukamel; Vladislav [...]

Author(s): Yoach Rais; Asaf Zviran; Shay Geula; Ohad Gafni; Elad Chomsky; Sergey Viukov; Abed AlFatah Mansour; Inbal Caspi; Vladislav Krupalnik; Mirie Zerbib; Itay Maza; Nofar Mor; Dror Baran; Leehee Weinberger; [...]