title: A knock-in mouse model of human PHD2 gene-associated erythrocytosis establishes a haploinsufficiency mechanism.
abstract: The central pathway for controlling red cell mass is the PHD (prolyl hydroxylase domain protein):hypoxia-inducible factor (HIF) pathway. HIF, which is negatively regulated by PHD, activates numerous genes, including ones involved in erythropoiesis, such as the ERYTHROPOIETIN (EPO) gene. Recent studies have implicated PHD2 as the key PHD isoform regulating red cell mass. Studies of humans have identified erythrocytosis-associated, heterozygous point mutations in the PHD2 gene. A key question concerns the mechanism by which human mutations lead to phenotypes. In the present report, we generated and characterized a mouse line in which a P294R knock-in mutation has been introduced into the mouse Phd2 locus to model the first reported human PHD2 mutation (P317R). Phd2(P294R/+) mice display a degree of erythrocytosis equivalent to that seen in Phd2(+/-) mice. The Phd2(P294R/+)-associated erythrocytosis is reversed in a Hif2a(+/-), but not a Hif1a(+/-) background. Additional studies using various conditional knock-outs of Phd2 reveal that erythrocytosis can be induced by homozygous and heterozygous knock-out of Phd2 in renal cortical interstitial cells using a Pax3-Cre transgene or by homozygous knock-out of Phd2 in hematopoietic progenitors driven by a Vav1-Cre transgene. These studies formally prove that a missense mutation in PHD2 is the cause of the erythrocytosis, show that this occurs through haploinsufficiency, and point to multifactorial control of red cell mass by PHD2.
authors.affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Patrick R
authors.initials: PR
authors.lastname: Arsenault
authors.affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Fei
authors.initials: F
authors.lastname: Pei
authors.affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Rebecca
authors.initials: R
authors.lastname: Lee
authors.affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Heddy
authors.initials: H
authors.lastname: Kerestes
authors.affiliation: Department of Haematology, Belfast City Hospital, Belfast, Northern Ireland BT9 7AB, United Kingdom.
authors.forename: Melanie J
authors.initials: MJ
authors.lastname: Percy
authors.affiliation: Department of Cancer Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104; Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Brian
authors.initials: B
authors.lastname: Keith
authors.affiliation: Abramson Family Cancer Research Institute, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104; Department of Cell and Developmental Biology, University of Pennsylvania, Philadelphia, Pennsylvania 19104; Howard Hughes Medical Institute, Philadelphia, Pennsylvania 19104.
authors.forename: M Celeste
authors.initials: MC
authors.lastname: Simon
authors.affiliation: Centre for Cancer Research and Cell Biology, Queen's University, Belfast, Northern Ireland BT9 7BL, United Kingdom.
authors.forename: Terence R J
authors.initials: TRJ
authors.lastname: Lappin
authors.affiliation: Department of Physiology and Pennsylvania Muscle Institute, University of Pennsylvania, Philadelphia, Pennsylvania 19104.
authors.forename: Tejvir S
authors.initials: TS
authors.lastname: Khurana
authors.affiliation: Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104. Electronic address: franklee@mail.med.upenn.edu.
authors.forename: Frank S
authors.initials: FS
authors.lastname: Lee
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D004195:Disease Models, Animal; D055879:Gene Knock-In Techniques; D057895:Haploinsufficiency; D006801:Humans; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D020125:Mutation, Missense; D011086:Polycythemia / Q000235:genetics / Q000378:metabolism* / Q000473:pathology
chemical_list: C490190:EGLN1 protein, human; C490189:Egln1 protein, mouse; D064799:Hypoxia-Inducible Factor-Proline Dioxygenases
keywords: Erythropoeisis; Erythropoietin; Hypoxia; Hypoxia-inducible Factor; Hypoxia-inducible Factor (HIF); Prolyl Hydroxylase Domain Protein