title: Efficient endoderm induction from human pluripotent stem cells by logically directing signals controlling lineage bifurcations.
abstract: Human pluripotent stem cell (hPSC) differentiation typically yields heterogeneous populations. Knowledge of signals controlling embryonic lineage bifurcations could efficiently yield desired cell types through exclusion of alternate fates. Therefore, we revisited signals driving induction and anterior-posterior patterning of definitive endoderm to generate a coherent roadmap for endoderm differentiation. With striking temporal dynamics, BMP and Wnt initially specified anterior primitive streak (progenitor to endoderm), yet, 24 hr later, suppressed endoderm and induced mesoderm. At lineage bifurcations, cross-repressive signals separated mutually exclusive fates; TGF-β and BMP/MAPK respectively induced pancreas versus liver from endoderm by suppressing the alternate lineage. We systematically blockaded alternate fates throughout multiple consecutive bifurcations, thereby efficiently differentiating multiple hPSC lines exclusively into endoderm and its derivatives. Comprehensive transcriptional and chromatin mapping of highly pure endodermal populations revealed that endodermal enhancers existed in a surprising diversity of "pre-enhancer" states before activation, reflecting the establishment of a permissive chromatin landscape as a prelude to differentiation.
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore; Department of Developmental Biology, Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA. Electronic address: kyleloh@stanford.edu.
authors.forename: Kyle M
authors.initials: KM
authors.lastname: Loh
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore. Electronic address: anglt1@gis.a-star.edu.sg.
authors.forename: Lay Teng
authors.initials: LT
authors.lastname: Ang
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Jingyao
authors.initials: J
authors.lastname: Zhang
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Vibhor
authors.initials: V
authors.lastname: Kumar
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Jasmin
authors.initials: J
authors.lastname: Ang
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Jun Qiang
authors.initials: JQ
authors.lastname: Auyeong
authors.affiliation: Cancer Science Institute of Singapore, Centre for Translational Medicine, National University of Singapore, Singapore 117599, Singapore.
authors.forename: Kian Leong
authors.initials: KL
authors.lastname: Lee
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Siew Hua
authors.initials: SH
authors.lastname: Choo
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Christina Y Y
authors.initials: CY
authors.lastname: Lim
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Massimo
authors.initials: M
authors.lastname: Nichane
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Junru
authors.initials: J
authors.lastname: Tan
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Monireh Soroush
authors.initials: MS
authors.lastname: Noghabi
authors.affiliation: Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia.
authors.forename: Lisa
authors.initials: L
authors.lastname: Azzola
authors.affiliation: Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia; Murdoch Childrens Research Institute, The Royal Children's Hospital, Parkville, VIC 3052, Australia.
authors.forename: Elizabeth S
authors.initials: ES
authors.lastname: Ng
authors.affiliation: Department of Developmental Biology, Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
authors.forename: Jens
authors.initials: J
authors.lastname: Durruthy-Durruthy
authors.affiliation: Department of Developmental Biology, Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
authors.forename: Vittorio
authors.initials: V
authors.lastname: Sebastiano
authors.affiliation: Cancer Science Institute of Singapore, Centre for Translational Medicine, National University of Singapore, Singapore 117599, Singapore.
authors.forename: Lorenz
authors.initials: L
authors.lastname: Poellinger
authors.affiliation: Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia; Murdoch Childrens Research Institute, The Royal Children's Hospital, Parkville, VIC 3052, Australia.
authors.forename: Andrew G
authors.initials: AG
authors.lastname: Elefanty
authors.affiliation: Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC 3800, Australia; Murdoch Childrens Research Institute, The Royal Children's Hospital, Parkville, VIC 3052, Australia.
authors.forename: Edouard G
authors.initials: EG
authors.lastname: Stanley
authors.affiliation: Humanized Mouse Unit, Institute of Molecular and Cell Biology, A(∗)STAR, Singapore 138673, Singapore; Interdisciplinary Research Group in Infectious Diseases, Singapore-MIT Alliance for Research and Technology, Singapore 138602, Singapore; Department of Microbiology, Yong Yoo Lin School of Medicine, National University of Singapore, Singapore 119228, Singapore.
authors.forename: Qingfeng
authors.initials: Q
authors.lastname: Chen
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore.
authors.forename: Shyam
authors.initials: S
authors.lastname: Prabhakar
authors.affiliation: Department of Developmental Biology, Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA 94305, USA.
authors.forename: Irving L
authors.initials: IL
authors.lastname: Weissman
authors.affiliation: Stem Cell and Developmental Biology Group, Genome Institute of Singapore, Agency for Science, Technology and Research (A(∗)STAR), Singapore 138672, Singapore; Department of Medicine, Harvard Medical School, Boston, MA 02115, USA; Division of Hematology/Oncology, Beth Israel Deaconess Medical Center, Boston, MA 02115, USA. Electronic address: limb1@gis.a-star.edu.sg.
authors.forename: Bing
authors.initials: B
authors.lastname: Lim
mesh_terms: D000818:Animals; D001483:Base Sequence; D019521:Body Patterning / Q000187:drug effects; D019485:Bone Morphogenetic Proteins / Q000378:metabolism; D019070:Cell Lineage / Q000187:drug effects; D002843:Chromatin / Q000378:metabolism; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004064:Digestive System / Q000196:embryology; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism; D004707:Endoderm / Q000166:cytology / Q000187:drug effects / Q000196:embryology*; D004742:Enhancer Elements, Genetic / Q000235:genetics; D044127:Epigenesis, Genetic / Q000187:drug effects; D005346:Fibroblast Growth Factors / Q000378:metabolism; D006801:Humans; D008099:Liver / Q000196:embryology; D020935:MAP Kinase Signaling System / Q000187:drug effects; D051379:Mice; D008969:Molecular Sequence Data; D010179:Pancreas / Q000196:embryology; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000187:drug effects; D054240:Primitive Streak / Q000166:cytology / Q000196:embryology; D011485:Protein Binding / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects; D014158:Transcription, Genetic / Q000187:drug effects; D016212:Transforming Growth Factor beta / Q000378:metabolism; D051153:Wnt Proteins / Q000378:metabolism
chemical_list: D019485:Bone Morphogenetic Proteins; D002843:Chromatin; D016895:Culture Media, Serum-Free; D016212:Transforming Growth Factor beta; D051153:Wnt Proteins; D005346:Fibroblast Growth Factors