Calmodulin Regulates Human Ether à Go-Go 1 (hEAG1) Potassium Channels through Interactions of the Eag Domain with the Cyclic Nucleotide Binding Homology Domain.
The ether à go-go family of voltage-gated potassium channels is structurally distinct. The N terminus contains an eag domain (eagD) that contains a Per-Arnt-Sim (PAS) domain that is preceded by a conserved sequence of 25-27 amino acids known as the PAS-cap. The C terminus contains a region with homology to cyclic nucleotide binding domains (cNBHD), which is directly linked to the channel pore. The human EAG1 (hEAG1) channel is remarkably sensitive to inhibition by intracellular calcium (Ca(2+) i) through binding of Ca(2+)-calmodulin to three sites adjacent to the eagD and cNBHD. Here, we show that the eagD and cNBHD interact to modulate Ca(2+)-calmodulin as well as voltage-dependent gating. Sustained elevation of Ca(2+) i resulted in an initial profound inhibition of hEAG1 currents, which was followed by a phase when current amplitudes partially recovered, but activation gating was slowed and shifted to depolarized potentials. Deletion of either the eagD or cNBHD abolished the inhibition by Ca(2+) i However, deletion of just the PAS-cap resulted in a >15-fold potentiation in response to elevated Ca(2+) i Mutations of residues at the interface between the eagD and cNBHD have been linked to human cancer. Glu-600 on the cNBHD, when substituted with residues with a larger volume, resulted in hEAG1 currents that were profoundly potentiated by Ca(2+) i in a manner similar to the ΔPAS-cap mutant. These findings provide the first evidence that eagD and cNBHD interactions are regulating Ca(2+)-dependent gating and indicate that the binding of the PAS-cap with the cNBHD is required for the closure of the channels upon CaM binding.
From the Department of Molecular and Cell Biology, University of Leicester, Leicester LE1 9HN.
Eva
E
Lörinczi
From the Department of Molecular and Cell Biology, University of Leicester, Leicester LE1 9HN, the School of Physiology and Pharmacology, University of Bristol, Bristol BS5 1TD, and.
Matthew
M
Helliwell
From the Department of Molecular and Cell Biology, University of Leicester, Leicester LE1 9HN.
Alina
A
Finch
the Department of Biochemistry, University of Oxford, Oxford OX1 3QU, United Kingdom.
Phillip J
PJ
Stansfeld
From the Department of Molecular and Cell Biology, University of Leicester, Leicester LE1 9HN.
Noel W
NW
Davies
From the Department of Molecular and Cell Biology, University of Leicester, Leicester LE1 9HN.
Martyn
M
Mahaut-Smith
From the Department of Molecular and Cell Biology, University of Leicester, Leicester LE1 9HN.
Frederick W
FW
Muskett
From the Department of Molecular and Cell Biology, University of Leicester, Leicester LE1 9HN, jm109@le.ac.uk.
John S
JS
Mitcheson
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000235:genetics / Q000378:metabolism*; D051638:Ether-A-Go-Go Potassium Channels / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D000072417:Protein Domains; D017384:Sequence Deletion; D014982:Xenopus laevis
D002147:Calmodulin; D051638:Ether-A-Go-Go Potassium Channels; C496884:KCNH1 protein, human; D002118:Calcium
calcium imaging; calmodulin (CaM); electrophysiology; gating; potassium channel