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EHD2 Stabilizes the Surface Expression of Cardiac K ATP Channels and Protects Against Ischemic Injury
Author(s) -
Foster Monique,
Jana Kundan,
Bergman Lisen,
Pennegård Elin,
Ördberg Malin,
Neubert Thomas,
Coetzee William
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.1025.16
Subject(s) - microbiology and biotechnology , myocyte , chemistry , potassium channel , hek 293 cells , transfection , ischemic preconditioning , ischemia , biophysics , biology , biochemistry , medicine , gene
Opening of ATP‐sensitive potassium (K ATP ) channels mediate ischemic preconditioning (IPC), which is an endogenous protection mechanism against ischemic injury. We found that internalization of K ATP channels during ischemia is prevented by IPC. We investigated the role of EHD2 (identified in our proteomic assays as a K ATP channel interacting protein) in K ATP channel internalization. A dominant‐negative EHD2‐G65R caused Kir6.2/SUR2A K ATP channels to redistribute in transfected COS‐7 cells. Both the dominant negative Rab11‐S25N and EHD2‐G65R led to accumulation of Kir6.2 in the endocytic recycling complex (ERC), suggesting that EHD2 controls exit from the ERC to the plasma membrane. Using biotinylation assays, we found that EHD2 decreased (whereas EHD2‐G65R accelerated) the rate of K ATP channel endocytosis. Surface biotinylation experiments showed that EHD2 stabilized and EHD2‐G65R decreased the surface density of Kir6.2/SUR2A channels in HEK‐293 cells. Consistently, patch clamping demonstrated that EHD2 increases, whereas EHD2‐G65R decreases, K ATP channel current density without affecting K ATP channel properties. Similar data were obtained with isolated adult mouse cardiac myocytes infected with Ad‐mCherry‐EHD2 or Ad‐mCherry‐EHD2‐G65R. We employed a cellular model of pharmacological preconditioning, in which myocytes are pretreated with phenylephrine prior to ischemic pelleting. Adult mouse cardiomyocytes infected with Ad‐mCherry‐EHD2 resisted ischemic injury, whereas cell death was more prominent in Ad‐mCherry‐EHD2‐G65R. Our data demonstrate that EHD2 positively regulates K ATP channel surface expression and participates in the protection afforded by preconditioning.