Small GTPase determinants for the Golgi processing and plasmalemmal expression of human ether-a-go-go related (hERG) K+ channels.
The pro-arrhythmic Long QT syndrome (LQT) is linked to 10 different genes (LQT1-10). Approximately 40% of genotype-positive LQT patients have LQT2, which is characterized by mutations in the human ether-a-go-go related gene (hERG). hERG encodes the voltage-gated K(+) channel alpha-subunits that form the pore of the rapidly activating delayed rectifier K(+) current in the heart. The purpose of this study was to elucidate the mechanisms that regulate the intracellular transport or trafficking of hERG, because trafficking is impaired for about 90% of LQT2 missense mutations. Protein trafficking is regulated by small GTPases. To identify the small GTPases that are critical for hERG trafficking, we coexpressed hERG and dominant negative (DN) GTPase mutations in HEK293 cells. The GTPases Sar1 and ARF1 regulate the endoplasmic reticulum (ER) export of proteins in COPII and COPI vesicles, respectively. Expression of DN Sar1 inhibited the Golgi processing of hERG, decreased hERG current (I(hERG)) by 85% (n > or = 8 cells per group, *, p < 0.01), and reduced the plasmalemmal staining of hERG. The coexpression of DN ARF1 had relatively small effects on hERG trafficking. Surprisingly, the coexpression of DN Rab11B, which regulates the endosomal recycling, inhibited the Golgi processing of hERG, decreased I(hERG) by 79% (n > or = 8 cells per group; *, p < 0.01), and reduced the plasmalemmal staining of hERG. These data suggest that hERG undergoes ER export in COPII vesicles and endosomal recycling prior to being processed in the Golgi. We conclude that hERG trafficking involves a pathway between the ER and endosomal compartments that influences expression in the plasmalemma.
Delisle
Brian P
BP
Department of Physiology, University of Kentucky, Lexington, Kentucky 40536. Electronic address: brian.delisle@uky.edu.
Underkofler
Heather A S
HAS
Cellular and Molecular Arrhythmia Research Program, Departments of Medicine and Physiology, University of Wisconsin, Madison, Wisconsin 53706.
Moungey
Brooke M
BM
Cellular and Molecular Arrhythmia Research Program, Departments of Medicine and Physiology, University of Wisconsin, Madison, Wisconsin 53706.
Slind
Jessica K
JK
Cellular and Molecular Arrhythmia Research Program, Departments of Medicine and Physiology, University of Wisconsin, Madison, Wisconsin 53706.
Kilby
Jennifer A
JA
Cellular and Molecular Arrhythmia Research Program, Departments of Medicine and Physiology, University of Wisconsin, Madison, Wisconsin 53706.
Best
Jabe M
JM
Cellular and Molecular Arrhythmia Research Program, Departments of Medicine and Physiology, University of Wisconsin, Madison, Wisconsin 53706.
Foell
Jason D
JD
Cellular and Molecular Arrhythmia Research Program, Departments of Medicine and Physiology, University of Wisconsin, Madison, Wisconsin 53706.
Balijepalli
Ravi C
RC
Cellular and Molecular Arrhythmia Research Program, Departments of Medicine and Physiology, University of Wisconsin, Madison, Wisconsin 53706.
Kamp
Timothy J
TJ
Cellular and Molecular Arrhythmia Research Program, Departments of Medicine and Physiology, University of Wisconsin, Madison, Wisconsin 53706.
January
Craig T
CT
Cellular and Molecular Arrhythmia Research Program, Departments of Medicine and Physiology, University of Wisconsin, Madison, Wisconsin 53706. Electronic address: ctj@medicine.wisc.edu.
D002462:Cell Membrane / Q000378:metabolism; D051638:Ether-A-Go-Go Potassium Channels / Q000235:genetics / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D018613:Microscopy, Confocal; D016297:Mutagenesis, Site-Directed; D020125:Mutation, Missense; D018408:Patch-Clamp Techniques
D051638:Ether-A-Go-Go Potassium Channels; D020558:GTP Phosphohydrolases