title: Identification of novel regulators of developmental hematopoiesis using Endoglin regulatory elements as molecular probes.
abstract: Enhancers are the primary determinants of cell identity, and specific promoter/enhancer combinations of Endoglin (ENG) have been shown to target blood and endothelium in the embryo. Here, we generated a series of embryonic stem cell lines, each targeted with reporter constructs driven by specific promoter/enhancer combinations of ENG, to evaluate their discriminative potential and value as molecular probes of the corresponding transcriptome. The Eng promoter (P) in combination with the -8/+7/+9-kb enhancers, targeted cells in FLK1 mesoderm that were enriched for blast colony forming potential, whereas the P/-8-kb enhancer targeted TIE2+/c-KIT+/CD41- endothelial cells that were enriched for hematopoietic potential. These fractions were isolated using reporter expression and their transcriptomes profiled by RNA-seq. There was high concordance between our signatures and those from embryos with defects at corresponding stages of hematopoiesis. Of the 6 genes that were upregulated in both hemogenic mesoderm and hemogenic endothelial fractions targeted by the reporters, LRP2, a multiligand receptor, was the only gene that had not previously been associated with hematopoiesis. We show that LRP2 is indeed involved in definitive hematopoiesis and by doing so validate the use of reporter gene-coupled enhancers as probes to gain insights into transcriptional changes that facilitate cell fate transitions.
authors.affiliation: Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia.
authors.forename: Rabab
authors.initials: R
authors.lastname: Nasrallah
authors.affiliation: Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA; Howard Hughes Medical Institute, Stem Cell Program and Division of Pediatric Hematology/Oncology, Boston Children's Hospital, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA.
authors.forename: Eva M
authors.initials: EM
authors.lastname: Fast
authors.affiliation: Erasmus Medical Center Stem Cell Institute, Department of Cell Biology, Erasmus University Medical Center, Rotterdam, The Netherlands.
authors.forename: Parham
authors.initials: P
authors.lastname: Solaimani
authors.affiliation: Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia.
authors.forename: Kathy
authors.initials: K
authors.lastname: Knezevic
authors.affiliation: Cancer Research United Kingdom Manchester Institute, The University of Manchester, Manchester, United Kingdom.
authors.forename: Alexia
authors.initials: A
authors.lastname: Eliades
authors.affiliation: Cancer Research United Kingdom Manchester Institute, The University of Manchester, Manchester, United Kingdom.
authors.forename: Rahima
authors.initials: R
authors.lastname: Patel
authors.affiliation: Cancer Research United Kingdom Manchester Institute, The University of Manchester, Manchester, United Kingdom.
authors.forename: Roshana
authors.initials: R
authors.lastname: Thambyrajah
authors.affiliation: Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia.
authors.forename: Ashwin
authors.initials: A
authors.lastname: Unnikrishnan
authors.affiliation: Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia.
authors.forename: Julie
authors.initials: J
authors.lastname: Thoms
authors.affiliation: Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia.
authors.forename: Dominik
authors.initials: D
authors.lastname: Beck
authors.affiliation: Erasmus Medical Center Stem Cell Institute, Department of Cell Biology, Erasmus University Medical Center, Rotterdam, The Netherlands; Medical Research Council Centre for Inflammation Research, University of Edinburgh, Edinburgh, United Kingdom.
authors.forename: Chris S
authors.initials: CS
authors.lastname: Vink
authors.affiliation: Department of Haematology, Wellcome Trust and Medical Research Council Cambridge Stem Cell Institute and Cambridge Institute for Medical Research, Cambridge University, Cambridge, United Kingdom.
authors.forename: Aileen
authors.initials: A
authors.lastname: Smith
authors.affiliation: Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia.
authors.forename: Jason
authors.initials: J
authors.lastname: Wong
authors.affiliation: Department of Haematology, Wellcome Trust and Medical Research Council Cambridge Stem Cell Institute and Cambridge Institute for Medical Research, Cambridge University, Cambridge, United Kingdom.
authors.forename: Mairi
authors.initials: M
authors.lastname: Shepherd
authors.affiliation: Department of Haematology, Wellcome Trust and Medical Research Council Cambridge Stem Cell Institute and Cambridge Institute for Medical Research, Cambridge University, Cambridge, United Kingdom.
authors.forename: David
authors.initials: D
authors.lastname: Kent
authors.affiliation: The Babraham Institute, Babraham Research Campus, Cambridge, United Kingdom.
authors.forename: Rahul
authors.initials: R
authors.lastname: Roychoudhuri
authors.affiliation: Max Delbrueck Center for Molecular Medicine, Berlin, Germany; and.
authors.forename: Fabian
authors.initials: F
authors.lastname: Paul
authors.affiliation: Max Delbrueck Center for Molecular Medicine, Berlin, Germany; and.
authors.forename: Julia
authors.initials: J
authors.lastname: Klippert
authors.affiliation: Max Delbrueck Center for Molecular Medicine, Berlin, Germany; and.
authors.forename: Annette
authors.initials: A
authors.lastname: Hammes
authors.affiliation: Max Delbrueck Center for Molecular Medicine, Berlin, Germany; and.
authors.forename: Thomas
authors.initials: T
authors.lastname: Willnow
authors.affiliation: Department of Haematology, Wellcome Trust and Medical Research Council Cambridge Stem Cell Institute and Cambridge Institute for Medical Research, Cambridge University, Cambridge, United Kingdom.
authors.forename: Bertie
authors.initials: B
authors.lastname: Göttgens
authors.affiliation: Erasmus Medical Center Stem Cell Institute, Department of Cell Biology, Erasmus University Medical Center, Rotterdam, The Netherlands; Medical Research Council Centre for Inflammation Research, University of Edinburgh, Edinburgh, United Kingdom.
authors.forename: Elaine
authors.initials: E
authors.lastname: Dzierzak
authors.affiliation: Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA; Howard Hughes Medical Institute, Stem Cell Program and Division of Pediatric Hematology/Oncology, Boston Children's Hospital, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA.
authors.forename: Leonard I
authors.initials: LI
authors.lastname: Zon
authors.affiliation: Cancer Research United Kingdom Manchester Institute, The University of Manchester, Manchester, United Kingdom.
authors.forename: George
authors.initials: G
authors.lastname: Lacaud
authors.affiliation: Cancer Research United Kingdom Manchester Institute, The University of Manchester, Manchester, United Kingdom.
authors.forename: Valerie
authors.initials: V
authors.lastname: Kouskoff
authors.affiliation: Lowy Cancer Research Centre and the Prince of Wales Clinical School, University of New South Wales Australia, Sydney, Australia; Department of Haematology, The Prince of Wales Hospital, Sydney, Australia.
authors.forename: John E
authors.initials: JE
authors.lastname: Pimanda
mesh_terms: D000818:Animals; D002460:Cell Line; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism*; D000071063:Endoglin / Q000235:genetics / Q000378:metabolism*; D042783:Endothelial Cells / Q000166:cytology / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000502:physiology*; D006410:Hematopoiesis / Q000502:physiology*; D026561:Low Density Lipoprotein Receptor-Related Protein-2 / Q000235:genetics / Q000378:metabolism; D008648:Mesoderm / Q000166:cytology / Q000378:metabolism; D051379:Mice; D015335:Molecular Probes / Q000235:genetics / Q000378:metabolism*; D000066450:Mouse Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism
chemical_list: D000071063:Endoglin; C502004:Eng protein, mouse; D026561:Low Density Lipoprotein Receptor-Related Protein-2; C055127:Lrp2 protein, mouse; D015335:Molecular Probes