Thapsigargin selectively rescues the trafficking defective LQT2 channels G601S and F805C.
Several mutations in the human ether-a-go-go-related K+ channel gene (HERG or KCNH2) cause long QT syndrome (LQT2) by reducing the intracellular transport (trafficking) of the channel protein to the cell surface. Drugs that bind to and block HERG channels (i.e. E4031) rescue the surface expression of some trafficking defective LQT2 mutations. Because these drugs potently block HERG current, their ability to correct congenital LQT is confounded by their risk of causing acquired LQT. We tested the hypothesis that pharmacological rescue can occur without HERG channel block. Thapsigargin (1 microM), a sarcoplasmic/endoplasmic reticulum Ca2+-ATPase inhibitor, rescued the surface expression of G601S, and it did so without blocking current. Thapsigargin-induced rescue and E4031-induced rescue caused complex glycosylation that was evident within 3 h of drug exposure. Disruption of the Golgi apparatus with brefeldin A prevented thapsigargin- and E4031-induced rescue of IG01S. Confocal imaging showed that G601S protein is predominantly "trapped" intracellularly and that both thapsigargin and E4031 promote its relocation to the surface membrane. We also studied two other trafficking defective LQT2 mutations. Thapsigargin rescued the C terminus mutation F805C but not N470D, whereas E4031 rescued N470D but not F805C. Other sarcoplasmic/endoplasmic reticulum Ca2+-ATPase inhibitors did not rescue G601S or F805C. This study 1) supports the hypothesis that the LQT2 trafficking defective phenotype can be reversed without blocking the channel; 2) demonstrates pharmacological rescue of a C terminus LQT2 mutation; and 3) shows that thapsigargin can correct trafficking defective phenotypes in more than one channel type and disease (i.e. LQT2 and cystic fibrosis).
Delisle
Brian P
BP
Department of Medicine (Cardiology), University of Wisconsin, Madison, Wisconsin 53706, USA.
Anderson
Corey L
CL
Balijepalli
Ravi C
RC
Anson
Blake D
BD
Kamp
Timothy J
TJ
January
Craig T
CT
D019943:Amino Acid Substitution; D001216:Asparagine; D001224:Aspartic Acid; D027682:Cation Transport Proteins; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D002467:Cell Nucleus / Q000187:drug effects / Q000502:physiology; D004268:DNA-Binding Proteins; D000072237:ERG1 Potassium Channel; D051638:Ether-A-Go-Go Potassium Channels; D005998:Glycine; D006801:Humans; D008133:Long QT Syndrome / Q000235:genetics / Q000503:physiopathology; D016297:Mutagenesis, Site-Directed; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers / Q000494:pharmacology; D015221:Potassium Channels / Q000187:drug effects / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D024642:Potassium Channels, Voltage-Gated; D021381:Protein Transport / Q000187:drug effects; D011994:Recombinant Proteins / Q000187:drug effects / Q000378:metabolism; D012694:Serine; D019284:Thapsigargin / Q000494:pharmacology*; D015534:Trans-Activators; D000071230:Transcriptional Regulator ERG
D027682:Cation Transport Proteins; D004268:DNA-Binding Proteins; C072312:ERG protein, human; D000072237:ERG1 Potassium Channel; D051638:Ether-A-Go-Go Potassium Channels; C000606913:KCNH2 protein, human; C092509:KCNH6 protein, human; D026902:Potassium Channel Blockers; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011994:Recombinant Proteins; D015534:Trans-Activators; D000071230:Transcriptional Regulator ERG; D001224:Aspartic Acid; D012694:Serine; D019284:Thapsigargin; D001216:Asparagine; D005998:Glycine