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Physiology (Bethesda, Md.) Feb 2008Phosphorylation of potassium channels affects their function and plays a major role in regulating cell physiology. Here, we review previous studies of potassium channel... (Review)
Review
Phosphorylation of potassium channels affects their function and plays a major role in regulating cell physiology. Here, we review previous studies of potassium channel phosphorylation, focusing first on studies employing site-directed mutagenesis of recombinant channels expressed in heterologous cells. We then discuss recent mass spectrometric-based approaches to identify and quantify phosphorylation at specific sites on native and recombinant potassium channels, and newly developed mass spectrometric-based techniques that may prove beneficial to future studies of potassium channel phosphorylation, its regulation, and its mechanism of channel modulation.
Topics: Animals; Humans; Ion Channel Gating; Mass Spectrometry; Membrane Potentials; Models, Molecular; Mutation; Phosphorylation; Potassium Channels; Protein Conformation; Protein Transport; Recombinant Proteins; Time Factors
PubMed: 18268365
DOI: 10.1152/physiol.00031.2007 -
Migration of PIP2 lipids on voltage-gated potassium channel surface influences channel deactivation.Scientific Reports Oct 2015Published studies of lipid-protein interactions have mainly focused on lipid binding to an individual site of the protein. Here, we show that a lipid can migrate between...
Published studies of lipid-protein interactions have mainly focused on lipid binding to an individual site of the protein. Here, we show that a lipid can migrate between different binding sites in a protein and this migration modulates protein function. Voltage-gated potassium (Kv) channels have several potential binding sites for phosphatidylinositol-4,5-bisphosphate (PIP2). Our molecular dynamics (MD) simulations on the KCNQ2 channel reveal that PIP2 preferentially binds to the S4-S5 linker when the channel is in the open state while maintains a certain probability of migrating to the S2-S3 linker. Guided by the MD results, electrophysiological experiments using KCNQ2, KCNQ1, and hERG channels show that the migration of PIP2 toward the S2-S3 linker controls the deactivation rate of the channel. The data suggest that PIP2 can migrate between different binding sites in Kv channels with significant impacts on channel deactivation, casting new insights into the dynamics and physiological functions of lipid-protein interactions.
Topics: Animals; CHO Cells; Cricetulus; Ether-A-Go-Go Potassium Channels; Humans; KCNQ2 Potassium Channel; Models, Molecular; Molecular Conformation; Molecular Dynamics Simulation; Mutation; Phosphatidylinositol 4,5-Diphosphate; Potassium Channels, Voltage-Gated; Protein Binding; Structure-Activity Relationship
PubMed: 26469389
DOI: 10.1038/srep15079 -
Marine Drugs Aug 2020Recently, Conorfamide-Sr3 (CNF-Sr3) was isolated from the venom of and was demonstrated to have an inhibitory concentration-dependent effect on the K channel. The...
Recently, Conorfamide-Sr3 (CNF-Sr3) was isolated from the venom of and was demonstrated to have an inhibitory concentration-dependent effect on the K channel. The voltage-gated potassium channels play critical functions on cellular signaling, from the regeneration of action potentials in neurons to the regulation of insulin secretion in pancreatic cells, among others. In mammals, there are at least 40 genes encoding voltage-gated K channels and the process of expression of some of them may include alternative splicing. Given the enormous variety of these channels and the proven use of conotoxins as tools to distinguish different ligand- and voltage-gated ion channels, in this work, we explored the possible effect of CNF-Sr3 on four human voltage-gated K channel subtypes homologous to the channel. CNF-Sr3 showed a 10 times higher affinity for the Kv1.6 subtype with respect to Kv1.3 (IC = 2.7 and 24 μM, respectively) and no significant effect on Kv1.4 and Kv1.5 at 10 µM. Thus, CNF-Sr3 might become a novel molecular probe to study diverse aspects of human Kv1.3 and Kv1.6 channels.
Topics: Animals; Conus Snail; Ion Channel Gating; Kv1.3 Potassium Channel; Kv1.4 Potassium Channel; Kv1.5 Potassium Channel; Kv1.6 Potassium Channel; Membrane Potentials; Mollusk Venoms; Oocytes; Potassium Channel Blockers; Shaker Superfamily of Potassium Channels; Xenopus laevis
PubMed: 32823677
DOI: 10.3390/md18080425 -
Current Biology : CB Mar 2001The number, type and distribution of ion channels on a neuron's surface determine its electrical response to stimulation. One way that a cell determines how many... (Review)
Review
The number, type and distribution of ion channels on a neuron's surface determine its electrical response to stimulation. One way that a cell determines how many molecules of each channel type are sent to the surface has been eludicated in a recent study of intrinsic protein transport signals within potassium channels.
Topics: Animals; Biological Transport; Endoplasmic Reticulum; Golgi Apparatus; Kv1.2 Potassium Channel; Kv1.4 Potassium Channel; Kv1.5 Potassium Channel; Potassium Channels; Potassium Channels, Inwardly Rectifying; Potassium Channels, Voltage-Gated; Protein Sorting Signals
PubMed: 11301268
DOI: 10.1016/s0960-9822(01)00111-7 -
Archives of Pathology & Laboratory... Jan 2001To describe the state of the art of our understanding of the long QT syndromes and to provide the genetic correlation of clinical severity of patients with this disorder. (Review)
Review
OBJECTIVES
To describe the state of the art of our understanding of the long QT syndromes and to provide the genetic correlation of clinical severity of patients with this disorder.
DATE SOURCES
In this review, we outline data that were obtained from work in our laboratory, as well as information reported in the literature.
STUDY SELECTION
The information in this review spans the last decade; data were obtained from the studies that had the most impact, as well as from recent work at our laboratory.
DATA EXTRACTION
The data reported herein were extracted from the world literature on sudden death and the clinical aspects of long QT syndrome. The genes identified to date, mutations in these genes, and the biophysical perturbations in the mutated ion channels, as well as the severity of disease, are detailed.
DATA SYNTHESIS
The extracted data are described as a state-of-the-art review.
CONCLUSIONS
The long QT syndromes, genetically heterogeneous disorders due to mutations in genes encoding ion channels, are relatively common causes of syncope and sudden death. The affected genes, along with the genetic background of individuals, determine the clinical severity of disease. An understanding of the mechanisms responsible for long QT syndrome is expected to enable development of specific therapies.
Topics: Cation Transport Proteins; Chromosome Mapping; DNA-Binding Proteins; ERG1 Potassium Channel; Electrocardiography; Ether-A-Go-Go Potassium Channels; Genes, Dominant; Genes, Recessive; Genotype; Humans; KCNQ Potassium Channels; KCNQ1 Potassium Channel; Long QT Syndrome; Mutation; NAV1.5 Voltage-Gated Sodium Channel; Phenotype; Potassium Channels; Potassium Channels, Voltage-Gated; Sodium Channels; Trans-Activators; Transcriptional Regulator ERG
PubMed: 11151064
DOI: 10.5858/2001-125-0116-GASOLQ -
Plant Physiology Jul 2013Phylogenetic analyses of small viral K channels suggests that they did not originate from their hosts, but instead could be the source of the postulated pore precursor... (Review)
Review
Phylogenetic analyses of small viral K channels suggests that they did not originate from their hosts, but instead could be the source of the postulated pore precursor in the evolution of K channels.
Topics: Evolution, Molecular; Potassium Channels; Selection, Genetic; Viral Proteins; Viruses
PubMed: 23719891
DOI: 10.1104/pp.113.219360 -
The Journal of Biological Chemistry Oct 2001Kv1.1 and Kv1.4 potassium channels are expressed as mature glycosylated proteins in brain, whereas they exhibited striking differences in degree of trans-Golgi...
Kv1.1 and Kv1.4 potassium channels are expressed as mature glycosylated proteins in brain, whereas they exhibited striking differences in degree of trans-Golgi glycosylation conversion and high cell surface expression when they were transiently expressed as homomers in cell lines. Kv1.4 exhibited a 70% trans-Golgi glycosylation conversion, whereas Kv1.1 showed none, and Kv1.4 exhibited a approximately 20-fold higher cell surface expression level as compared with Kv1.1. Chimeras between Kv1.4 and Kv1.1 and site-directed mutants were constructed to identify amino acid determinants that affected these processes. Truncating the cytoplasmic C terminus of Kv1.4 inhibited its trans-Golgi glycosylation and high cell surface expression (as shown by Li, D., Takimoto, K., and Levitan, E. S. (2000) J. Biol. Chem. 275, 11597-11602), whereas truncating this region on Kv1.1 did not affect either of these events, indicating that its C terminus is not a negative determinant for these processes. Exchanging the C terminus between these channels showed that there are other regions of the protein that exert a positive or negative effect on these processes. Chimeric constructs between Kv1.4 and Kv1.1 identified their outer pore regions as major positive and negative determinants, respectively, for both trans-Golgi glycosylation and cell surface expression. Site-directed mutagenesis identified a number of amino acids in the pore region that are involved in these processes. These data suggest that there are multiple positive and negative determinants on both Kv1.4 and Kv1.1 that affect channel folding, trans-Golgi glycosylation conversion, and cell surface expression.
Topics: Amino Acid Sequence; Amino Acids; Animals; Brain; CHO Cells; Cell Membrane; Cricetinae; Cytoplasm; DNA, Complementary; Endoplasmic Reticulum; Glycoside Hydrolases; Glycosylation; Golgi Apparatus; Immunoblotting; Kv1.1 Potassium Channel; Kv1.4 Potassium Channel; Molecular Sequence Data; Mutagenesis, Site-Directed; Potassium Channels; Potassium Channels, Voltage-Gated; Protein Binding; Protein Folding; Protein Structure, Tertiary; Rats; Recombinant Fusion Proteins; Sequence Homology, Amino Acid; Transfection
PubMed: 11487588
DOI: 10.1074/jbc.M107399200 -
Biochimica Et Biophysica Acta Feb 1999Large conductance, calcium-activated potassium (maxiK) channels are expressed in nerve, muscle, and other cell types and are important determinants of smooth muscle...
Large conductance, calcium-activated potassium (maxiK) channels are expressed in nerve, muscle, and other cell types and are important determinants of smooth muscle tone. To determine the mechanisms involved in the transcriptional regulation of maxiK channels, we characterized the promoter regions of the pore forming (alpha) and regulatory (beta) subunits of the human channel complex. Maximum promoter activity (up to 12.3-fold over control) occurred between nucleotides -567 and -220 for the alpha subunit (hSlo) gene. The minimal promoter is GC-rich with 5 Sp-1 binding sites and several TCC repeats. Other transcription factor-binding motifs, including c/EBP, NF-kB, PU.1, PEA-3, Myo-D, and E2A, were observed in the 5'-flanking sequence. Additionally, a CCTCCC sequence, which increases the transcriptional activity of the SM1/2 gene in smooth muscle, is located 27 bp upstream of the TATA-like sequence, a location identical to that found in the SM1/2 5'-flanking region. However, the promoter directed equivalent expression when transfected into smooth muscle and other cell types. Analysis of the hSlo beta subunit 5'-flanking region revealed a TATA box at position -77 and maximum promoter activity (up to 11.0-fold) in a 200 bp region upstream from the cap site. Binding sites for GATA-1, Myo-D, c-myb, Ets-1/Elk-1, Ap-1, and Ik-2 were identified within this sequence. Two CCTCCC elements are present in the hSlo beta subunit promoter, but tissue-specific transcriptional activity was not observed. The lack of tissue-specific promoter activity, particularly the finding of promoter activity in cells from tissues in which the maxiK gene is not expressed, suggests a complex channel regulatory mechanism for hSlo genes. Moreover, the lack of similarity of the promoters of the two genes suggests that regulation of coordinate expression of the subunits does not occur through equivalent cis-acting sequences.
Topics: Base Sequence; Cell Line; Cloning, Molecular; Genomic Library; Humans; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; Large-Conductance Calcium-Activated Potassium Channel beta Subunits; Large-Conductance Calcium-Activated Potassium Channels; Molecular Sequence Data; Muscle, Skeletal; Muscle, Smooth; Potassium Channels; Potassium Channels, Calcium-Activated; Promoter Regions, Genetic
PubMed: 10023076
DOI: 10.1016/s0167-4781(98)00276-0 -
Neuron Dec 2014Mechanical and thermal activation of ion channels is central to touch, thermosensation, and pain. The TRAAK/TREK K(2P) potassium channel subfamily produces background...
Mechanical and thermal activation of ion channels is central to touch, thermosensation, and pain. The TRAAK/TREK K(2P) potassium channel subfamily produces background currents that alter neuronal excitability in response to pressure, temperature, signaling lipids, and anesthetics. How such diverse stimuli control channel function is unclear. Here we report structures of K(2P)4.1 (TRAAK) bearing C-type gate-activating mutations that reveal a tilting and straightening of the M4 inner transmembrane helix and a buckling of the M2 transmembrane helix. These conformational changes move M4 in a direction opposite to that in classical potassium channel activation mechanisms and open a passage lateral to the pore that faces the lipid bilayer inner leaflet. Together, our findings uncover a unique aspect of K(2P) modulation, indicate a means for how the K(2P) C-terminal cytoplasmic domain affects the C-type gate which lies ∼40Å away, and suggest how lipids and bilayer inner leaflet deformations may gate the channel.
Topics: Animals; Cell Membrane; Cells, Cultured; Ion Channel Gating; Lipid Bilayers; Mutation; Oocytes; Physical Stimulation; Potassium Channels; Protein Structure, Secondary; Temperature; Xenopus laevis
PubMed: 25500157
DOI: 10.1016/j.neuron.2014.11.017 -
The Journal of Cell Biology Jan 2000Mechanisms of ion channel clustering by cytoplasmic membrane-associated guanylate kinases such as postsynaptic density 95 (PSD-95) and synapse-associated protein 97...
Mechanisms of ion channel clustering by cytoplasmic membrane-associated guanylate kinases such as postsynaptic density 95 (PSD-95) and synapse-associated protein 97 (SAP97) are poorly understood. Here, we investigated the interaction of PSD-95 and SAP97 with voltage-gated or Kv K(+) channels. Using Kv channels with different surface expression properties, we found that clustering by PSD-95 depended on channel cell surface expression. Moreover, PSD-95-induced clusters of Kv1 K(+) channels were present on the cell surface. This was most dramatically demonstrated for Kv1.2 K(+) channels, where surface expression and clustering by PSD-95 were coincidentally promoted by coexpression with cytoplasmic Kvbeta subunits. Consistent with a mechanism of plasma membrane channel-PSD-95 binding, coexpression with PSD-95 did not affect the intrinsic surface expression characteristics of the different Kv channels. In contrast, the interaction of Kv1 channels with SAP97 was independent of Kv1 surface expression, occurred intracellularly, and prevented further biosynthetic trafficking of Kv1 channels. As such, SAP97 binding caused an intracellular accumulation of each Kv1 channel tested, through the accretion of SAP97 channel clusters in large (3-5 microm) ER-derived intracellular membrane vesicles. Together, these data show that ion channel clustering by PSD-95 and SAP97 occurs by distinct mechanisms, and suggests that these channel-clustering proteins may play diverse roles in regulating the abundance and distribution of channels at synapses and other neuronal membrane specializations.
Topics: Adaptor Proteins, Signal Transducing; Amino Acid Sequence; Animals; COS Cells; Cell Membrane; Discs Large Homolog 1 Protein; Disks Large Homolog 4 Protein; Guanylate Kinases; Humans; Intracellular Signaling Peptides and Proteins; Kv1.1 Potassium Channel; Kv1.2 Potassium Channel; Kv1.4 Potassium Channel; Membrane Proteins; Mice; Mice, Inbred BALB C; Molecular Sequence Data; Nerve Tissue Proteins; Potassium Channels; Potassium Channels, Voltage-Gated; Subcellular Fractions
PubMed: 10629225
DOI: 10.1083/jcb.148.1.147