Carbon monoxide promotes VEGF expression by increasing HIF-1alpha protein level via two distinct mechanisms, translational activation and stabilization of HIF-1alpha protein.
Carbon monoxide (CO) plays a significant role in vascular functions. We here examined the molecular mechanism by which CO regulates HIF-1 (hypoxia-inducible transcription factor-1)-dependent expression of vascular endothelial growth factor (VEGF), which is an important angiogenic factor. We found that astrocytes stimulated with CORM-2 (CO-releasing molecule) promoted angiogenesis by increasing VEGF expression and secretion. CORM-2 also induced HO-1 (hemeoxygenase-1) expression and increased nuclear HIF-1α protein level, without altering its promoter activity and mRNA level. VEGF expression was inhibited by treatment with HIF-1α siRNA and a hemeoxygenase inhibitor, indicating that CO stimulates VEGF expression via up-regulation of HIF-1α protein level, which is partially associated with HO-1 induction. CORM-2 activated the translational regulatory proteins p70(S6k) and eIF-4E as well as phosphorylating their upstream signal mediators Akt and ERK. These translational signal events and HIF-1α protein level were suppressed by inhibitors of phosphatidylinositol 3-kinase (PI3K), MEK, and mTOR, suggesting that the PI3K/Akt/mTOR and MEK/ERK pathways are involved in a translational increase in HIF-1α. In addition, CORM-2 also increased stability of the HIF-1α protein by suppressing its ubiquitination, without altering the proline hydroxylase-dependent HIF-1α degradation pathway. CORM-2 increased HIF-1α/HSP90α interaction, which is responsible for HIF-1α stabilization, and HSP90-specific inhibitors decreased this interaction, HIF-1α protein level, and VEGF expression. Furthermore, HSP90α knockdown suppressed CORM-2-induced increases in HIF-1α and VEGF protein levels. These results suggest that CO stimulates VEGF production by increasing HIF-1α protein level via two distinct mechanisms, translational stimulation and protein stabilization of HIF-1α.
Choi
Yoon Kyung
YK
Vascular System Research Center and Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon, Kangwon-do 200-701, Republic of Korea.
Kim
Chun-Ki
CK
Lee
Hansoo
H
Jeoung
Dooil
D
Ha
Kwon-Soo
KS
Kwon
Young-Guen
YG
Kim
Kyu-Won
KW
Kim
Young-Myeong
YM
D000818:Animals; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects / Q000502:physiology; D002248:Carbon Monoxide / Q000494:pharmacology*; D002465:Cell Movement; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology / Q000502:physiology; D005786:Gene Expression Regulation / Q000187:drug effects*; D018841:HSP90 Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism; D051547:Heme Oxygenase-1 / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000235:genetics / Q000378:metabolism*; D018919:Neovascularization, Physiologic / Q000187:drug effects; D009942:Organometallic Compounds / Q000494:pharmacology; D014176:Protein Biosynthesis / Q000187:drug effects*; D055550:Protein Stability; D034622:RNA Interference; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000378:metabolism*
D018841:HSP90 Heat-Shock Proteins; C533585:HSP90AA2P protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009942:Organometallic Compounds; D042461:Vascular Endothelial Growth Factor A; C447082:tricarbonyldichlororuthenium (II) dimer; D002248:Carbon Monoxide; D051547:Heme Oxygenase-1