title: Derivation of novel human ground state naive pluripotent stem cells.
abstract: Mouse embryonic stem (ES) cells are isolated from the inner cell mass of blastocysts, and can be preserved in vitro in a naive inner-cell-mass-like configuration by providing exogenous stimulation with leukaemia inhibitory factor (LIF) and small molecule inhibition of ERK1/ERK2 and GSK3β signalling (termed 2i/LIF conditions). 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. Upon withdrawal of 2i/LIF, naive mouse ES cells can drift towards a primed pluripotent state resembling that of the post-implantation epiblast. Although human ES cells share several molecular features with naive mouse ES cells, they also share a variety of epigenetic properties with primed murine epiblast stem cells (EpiSCs). 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. 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. 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 in vitro and in vivo.
authors.affiliation: 1] The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel [2].
authors.forename: Ohad
authors.initials: O
authors.lastname: Gafni
authors.affiliation: 1] The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel [2].
authors.forename: Leehee
authors.initials: L
authors.lastname: Weinberger
authors.affiliation: 1] The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel [2].
authors.forename: Abed AlFatah
authors.initials: AA
authors.lastname: Mansour
authors.affiliation: 1] The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel [2].
authors.forename: Yair S
authors.initials: YS
authors.lastname: Manor
authors.affiliation: 1] The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel [2] The Department of Biological Regulation, Weizmann Institute of Science, Rehovot 76100, Israel [3] The Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 76100, Israel [4].
authors.forename: Elad
authors.initials: E
authors.lastname: Chomsky
authors.affiliation: 1] Wolfe PGD Stem Cell Lab, Racine IVF Unit, Lis Maternity Hospital, Tel Aviv Sourasky Medical Center, Tel-Aviv, Israel [2] The Department of Cell and Developmental Biology, Sackler Medical School, Tel-Aviv University, Israel.
authors.forename: Dalit
authors.initials: D
authors.lastname: Ben-Yosef
authors.affiliation: Wolfe PGD Stem Cell Lab, Racine IVF Unit, Lis Maternity Hospital, Tel Aviv Sourasky Medical Center, Tel-Aviv, Israel.
authors.forename: Yael
authors.initials: Y
authors.lastname: Kalma
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Sergey
authors.initials: S
authors.lastname: Viukov
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Itay
authors.initials: I
authors.lastname: Maza
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Asaf
authors.initials: A
authors.lastname: Zviran
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Yoach
authors.initials: Y
authors.lastname: Rais
authors.affiliation: 1] The Department of Biological Regulation, Weizmann Institute of Science, Rehovot 76100, Israel [2] The Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Zohar
authors.initials: Z
authors.lastname: Shipony
authors.affiliation: 1] The Department of Biological Regulation, Weizmann Institute of Science, Rehovot 76100, Israel [2] The Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Zohar
authors.initials: Z
authors.lastname: Mukamel
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Vladislav
authors.initials: V
authors.lastname: Krupalnik
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Mirie
authors.initials: M
authors.lastname: Zerbib
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Shay
authors.initials: S
authors.lastname: Geula
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Inbal
authors.initials: I
authors.lastname: Caspi
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Dan
authors.initials: D
authors.lastname: Schneir
authors.affiliation: Wolfe PGD Stem Cell Lab, Racine IVF Unit, Lis Maternity Hospital, Tel Aviv Sourasky Medical Center, Tel-Aviv, Israel.
authors.forename: Tamar
authors.initials: T
authors.lastname: Shwartz
authors.affiliation: The Israel National Center for Personalized Medicine (INCPM), Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Shlomit
authors.initials: S
authors.lastname: Gilad
authors.affiliation: The Israel National Center for Personalized Medicine (INCPM), Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Daniela
authors.initials: D
authors.lastname: Amann-Zalcenstein
authors.affiliation: The Israel National Center for Personalized Medicine (INCPM), Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Sima
authors.initials: S
authors.lastname: Benjamin
authors.affiliation: The Department of Immunology, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Ido
authors.initials: I
authors.lastname: Amit
authors.affiliation: 1] The Department of Biological Regulation, Weizmann Institute of Science, Rehovot 76100, Israel [2] The Department of Computer Science and Applied Mathematics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Amos
authors.initials: A
authors.lastname: Tanay
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Rada
authors.initials: R
authors.lastname: Massarwa
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Noa
authors.initials: N
authors.lastname: Novershtern
authors.affiliation: The Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel.
authors.forename: Jacob H
authors.initials: JH
authors.lastname: Hanna
mesh_terms: D000818:Animals; D001755:Blastocyst / Q000166:cytology; D065150:Cellular Reprogramming; D002678:Chimera / Q000196:embryology; D002843:Chromatin / Q000378:metabolism; D019175:DNA Methylation; D004622:Embryo, Mammalian / Q000166:cytology / Q000196:embryology; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism; D044127:Epigenesis, Genetic; D005260:Female; D005855:Germ Layers / Q000166:cytology; D006657:Histones / Q000378:metabolism; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism / Q000637:transplantation; D008297:Male; D051379:Mice; D009028:Morula / Q000166:cytology; D038081:Organogenesis; D011401:Promoter Regions, Genetic / Q000235:genetics; D044968:Regenerative Medicine; D015203:Reproducibility of Results; D015398:Signal Transduction; D049951:X Chromosome Inactivation
chemical_list: D002843:Chromatin; D006657:Histones