title: A secreted protein (Canopy 2, CNPY2) enhances angiogenesis and promotes smooth muscle cell migration and proliferation.
abstract: OBJECTIVE
  Ischaemic heart disease is a leading cause of mortality. After ischaemic injury, tissue hypoxia induces the activity of angiogenic factors that promote revascularization. Increased understanding of hypoxia-responsive genes and their role in angiogenesis will lead to new therapies for ischaemic injury. We delineated the function of Canopy 2 (CNPY2), a newly discovered, hypoxia-regulated gene.
  
  
  RESULTS
  We found CNPY2 in a screen for genes induced by low oxygen in human smooth muscle cells (SMCs). CNPY2 protein co-localized with the endoplasmic reticulum and the Golgi. Treatment with Brefeldin A, which destroys Golgi stacks, resulted in CNPY2 accumulation in the endoplasmic reticulum. Secreted CNPY2 was detected in the blood of healthy mice and humans, and the medium of cultured SMCs. SMCs under hypoxia or treated with a prolyl-4-hydroxylase inhibitor stabilized HIF-1α protein and up-regulated CNPY2, while CNPY2 induction was lost after HIF-1α silencing. Chromatin immunoprecipitation demonstrated that HIF-1α binds to a hypoxia response element (HRE-1157) upstream of the human CNPY2 promoter, which was verified by a luciferase reporter driven by HRE-1157-containing constructs. CNPY2 stimulation activated Cdc42, PAK1, and FAK in SMCs, resulting in enhanced proliferation and migration in vitro, and dramatic aortic ring sprouting ex vivo. CNPY2 significantly increased revascularization of the mouse retina after reperfusion injury.
  
  
  CONCLUSIONS
  CNPY2 is a HIF-1α-regulated, secreted angiogenic growth factor that promotes SMC migration, proliferation, and tissue revascularization. This new target may have a broader profile than currently available proteins.
authors.affiliation: Division of Cardiovascular Surgery, Toronto General Research Institute, University Health Network, MaRS Centre, Toronto Medical Discovery Tower, Room 3-702, 101 College Street, Toronto, Ontario, Canada M5G 1L7 Department of Surgery, Division of Cardiac Surgery, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Jian
authors.initials: J
authors.lastname: Guo
authors.affiliation: Division of Cardiovascular Surgery, Toronto General Research Institute, University Health Network, MaRS Centre, Toronto Medical Discovery Tower, Room 3-702, 101 College Street, Toronto, Ontario, Canada M5G 1L7 Department of Surgery, Division of Cardiac Surgery, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Yuemei
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Division of Cardiovascular Surgery, Toronto General Research Institute, University Health Network, MaRS Centre, Toronto Medical Discovery Tower, Room 3-702, 101 College Street, Toronto, Ontario, Canada M5G 1L7 Department of Surgery, Division of Cardiac Surgery, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Anton
authors.initials: A
authors.lastname: Mihic
authors.affiliation: Division of Cardiovascular Surgery, Toronto General Research Institute, University Health Network, MaRS Centre, Toronto Medical Discovery Tower, Room 3-702, 101 College Street, Toronto, Ontario, Canada M5G 1L7 Department of Surgery, Division of Cardiac Surgery, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Shu-Hong
authors.initials: SH
authors.lastname: Li
authors.affiliation: Division of Cardiovascular Surgery, Toronto General Research Institute, University Health Network, MaRS Centre, Toronto Medical Discovery Tower, Room 3-702, 101 College Street, Toronto, Ontario, Canada M5G 1L7 Department of Surgery, Division of Cardiac Surgery, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Zhuo
authors.initials: Z
authors.lastname: Sun
authors.affiliation: Division of Cardiovascular Surgery, Toronto General Research Institute, University Health Network, MaRS Centre, Toronto Medical Discovery Tower, Room 3-702, 101 College Street, Toronto, Ontario, Canada M5G 1L7 Department of Surgery, Division of Cardiac Surgery, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Zhengbo
authors.initials: Z
authors.lastname: Shao
authors.affiliation: Division of Cardiovascular Surgery, Toronto General Research Institute, University Health Network, MaRS Centre, Toronto Medical Discovery Tower, Room 3-702, 101 College Street, Toronto, Ontario, Canada M5G 1L7 Department of Surgery, Division of Cardiac Surgery, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Jun
authors.initials: J
authors.lastname: Wu
authors.affiliation: Division of Cardiovascular Surgery, Toronto General Research Institute, University Health Network, MaRS Centre, Toronto Medical Discovery Tower, Room 3-702, 101 College Street, Toronto, Ontario, Canada M5G 1L7 Department of Surgery, Division of Cardiac Surgery, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Richard D
authors.initials: RD
authors.lastname: Weisel
authors.affiliation: Division of Cardiovascular Surgery, Toronto General Research Institute, University Health Network, MaRS Centre, Toronto Medical Discovery Tower, Room 3-702, 101 College Street, Toronto, Ontario, Canada M5G 1L7 Department of Surgery, Division of Cardiac Surgery, University of Toronto, Toronto, Ontario, Canada renkeli@uhnres.utoronto.ca.
authors.forename: Ren-Ke
authors.initials: RK
authors.lastname: Li
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000097:blood / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D015687:Cell Hypoxia; D002465:Cell Movement; D049109:Cell Proliferation; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism; D047908:Intracellular Signaling Peptides and Proteins / Q000097:blood / Q000235:genetics / Q000378:metabolism*; D008565:Membrane Proteins / Q000097:blood / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D009131:Muscle, Smooth, Vascular / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D018919:Neovascularization, Physiologic; D011401:Promoter Regions, Genetic; D051381:Rats; D015427:Reperfusion Injury / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology; D015861:Retinal Neovascularization / Q000235:genetics / Q000378:metabolism / Q000503:physiopathology; D015398:Signal Transduction; D013997:Time Factors; D014158:Transcription, Genetic
chemical_list: D048868:Adaptor Proteins, Signal Transducing; C502333:CNPY2 protein, human; C000603106:Canopy 2 protein, mouse; C497442:HIF1A protein, human; C497443:Hif1a protein, mouse; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins
keywords: Angiogenesis; CNPY2; HIF-1α; Hypoxia; Hypoxia response element; Revascularization