title: The Human Ether-a-go-go-related Gene (hERG) Potassium Channel Represents an Unusual Target for Protease-mediated Damage.
abstract: The human ether-a-go-go-related gene (hERG) encodes the pore-forming subunit of the rapidly activating delayed rectifier potassium channel (IKr), which is important for cardiac repolarization. Dysfunction of hERG causes long QT syndrome and sudden death, which occur in patients with cardiac ischemia. Cardiac ischemia is also associated with activation, up-regulation, and secretion of various proteolytic enzymes. Here, using whole-cell patch clamp and Western blotting analysis, we demonstrate that the hERG/IKr channel was selectively cleaved by the serine protease, proteinase K (PK). Using molecular biology techniques including making a chimeric channel between protease-sensitive hERG and insensitive human ether-a-go-go (hEAG), as well as application of the scorpion toxin BeKm-1, we identified that the S5-pore linker of hERG is the target domain for proteinase K cleavage. To investigate the physiological relevance of the unique susceptibility of hERG to proteases, we show that cardiac ischemia in a rabbit model was associated with a reduction in mature ERG expression and an increase in the expression of several proteases, including calpain. Using cell biology approaches, we found that calpain-1 was actively released into the extracellular milieu and cleaved hERG at the S5-pore linker. Using protease cleavage-predicting software and site-directed mutagenesis, we identified that calpain-1 cleaves hERG at position Gly-603 in the S5-pore linker of hERG. Clarification of protease-mediated damage of hERG extends our understanding of hERG regulation. Damage of hERG mediated by proteases such as calpain may contribute to ischemia-associated QT prolongation and sudden cardiac death.
authors: From the Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors: Shawn M
authors: SM
authors: Lamothe
authors: From the Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors: Jun
authors: J
authors: Guo
authors: From the Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors: Wentao
authors: W
authors: Li
authors: From the Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors: Tonghua
authors: T
authors: Yang
authors: From the Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada shetuan.zhang@queensu.ca.
authors: Shetuan
authors: S
authors: Zhang
mesh_terms: D000818:Animals; D002154:Calpain / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D000072237:ERG1 Potassium Channel / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019286:Endopeptidase K / Q000737:chemistry; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D017202:Myocardial Ischemia / Q000235:genetics / Q000378:metabolism*; D059748:Proteolysis; D011817:Rabbits; D012604:Scorpion Venoms / Q000494:pharmacology
chemical_list: C425305:BeKm-1 toxin; D000072237:ERG1 Potassium Channel; C000606913:KCNH2 protein, human; D012604:Scorpion Venoms; D019286:Endopeptidase K; D002154:Calpain; C484806:CAPN1 protein, human
keywords: cell surface protein; electrophysiology; hERG; ion channel; patch clamp; potassium channel; protease