The Kv2.1 C terminus can autonomously transfer Kv2.1-like phosphorylation-dependent localization, voltage-dependent gating, and muscarinic modulation to diverse Kv channels.
Modulation of K+ channels is widely used to dynamically regulate neuronal membrane excitability. The voltage-gated K+ channel Kv2.1 is an abundant delayed rectifier K+ (IK) channel expressed at high levels in many types of mammalian central neurons where it regulates diverse aspects of membrane excitability. Neuronal Kv2.1 is constitutively phosphorylated, localized in high-density somatodendritic clusters, and has a relatively depolarized voltage dependence of activation. Here, we show that the clustering and voltage-dependent gating of endogenous Kv2.1 in cultured rat hippocampal neurons are modulated by cholinergic stimulation, a common form of neuromodulation. The properties of neuronal Kv2.1 are recapitulated in recombinant Kv2.1 expressed in human embryonic kidney 293 (HEK293) cells, but not COS-1 cells, because of cell background-specific differences in Kv2.1 phosphorylation. As in neurons, Kv2.1 in HEK293 cells is dynamically regulated by cholinergic stimulation, which leads to Ca2+/calcineurin-dependent dephosphorylation of Kv2.1, dispersion of channel clusters, and hyperpolarizing shifts in the voltage-dependent gating properties of the channel. Immunocytochemical, biochemical, and biophysical analyses of chimeric Kv channels show that the Kv2.1 cytoplasmic C-terminal domain can act as an autonomous domain sufficient to transfer Kv2.1-like clustering, voltage-dependent activation, and cholinergic modulation to diverse Kv channels. These findings provide novel mechanistic insights into cholinergic modulation of ion channels and regulation of the localization and voltage-dependent gating properties of the abundant neuronal Kv2.1 channel by cholinergic and other neuromodulatory stimuli.
Mohapatra
Durga P
DP
Department of Pharmacology, School of Medicine, University of California, Davis, California 95616, USA.
Trimmer
James S
JS
D000818:Animals; D019556:COS Cells / Q000378:metabolism; D019703:Calcineurin / Q000502:physiology; D002118:Calcium / Q000502:physiology; D002217:Carbachol / Q000494:pharmacology; D002460:Cell Line / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D002478:Cells, Cultured / Q000378:metabolism; D002522:Chlorocebus aethiops; D004591:Electrophoresis, Polyacrylamide Gel; D006624:Hippocampus / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D006801:Humans; D015640:Ion Channel Gating; D017136:Ion Transport; D015759:Ionomycin / Q000494:pharmacology; D007668:Kidney; D051666:Kv1.5 Potassium Channel / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D008564:Membrane Potentials; D018721:Muscarinic Agonists / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D009474:Neurons / Q000378:metabolism; D009928:Organ Specificity; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D011188:Potassium / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D051381:Rats; D011976:Receptors, Muscarinic / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000502:physiology; D051669:Shab Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D016037:Single-Blind Method; D013045:Species Specificity; D013329:Structure-Activity Relationship; D014162:Transfection
D051666:Kv1.5 Potassium Channel; D018721:Muscarinic Agonists; D009419:Nerve Tissue Proteins; D011976:Receptors, Muscarinic; D011993:Recombinant Fusion Proteins; D051669:Shab Potassium Channels; D015759:Ionomycin; D002217:Carbachol; D019703:Calcineurin; D011188:Potassium; D002118:Calcium