Jian Guo1, Yuemei Zhang1, Anton Mihic1, Shu-Hong Li1, Zhuo Sun1, Zhengbo Shao1, Jun Wu1, Richard D Weisel1, Ren-Ke Li2. 1. 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. 2. 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.
Abstract
AIMS: 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. METHODS AND 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. Published on behalf of the European Society of Cardiology. All rights reserved.
AIMS: 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. METHODS AND 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 humanCNPY2 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. Published on behalf of the European Society of Cardiology. All rights reserved.
Authors: Feng Hong; Bei Liu; Bill X Wu; Jordan Morreall; Brady Roth; Christopher Davies; Shaoli Sun; J Alan Diehl; Zihai Li Journal: Nat Struct Mol Biol Date: 2017-09-04 Impact factor: 15.369