(Journal Article): Sequence and expression of mRNAs encoding the alpha 1 and alpha 2 subunits of a DHP-sensitive calcium channel.
Ellis SB, Williams ME, Ways NR, Brenner R, Sharp AH, Leung AT, Campbell KP, McKenna E, Koch WJ, Hui A
IN:
Science
1988; 241(4873):1661-1664
Impact Factor(s) of Science: 30.927 (2005), 31.853 (2004), 29.162 (2003), 26.682 (2002), 23.329 (2001)
Fulltext: HTML
ABSTRACT: Complementary DNAs were isolated and used to deduce the primary structures of the alpha 1 and alpha 2 subunits of the dihydropyridine-sensitive, voltage-dependent calcium channel from rabbit skeletal muscle. The alpha 1 subunit, which contains putative binding sites for calcium antagonists, is a hydrophobic protein with a sequence that is consistent with multiple transmembrane domains and shows structural and sequence homology with other voltage-dependent ion channels. In contrast, the alpha 2 subunit is a hydrophilic protein without homology to other known protein sequences. Nucleic acid hybridization studies suggest that the alpha 1 and alpha 2 subunit mRNAs are expressed differentially in a tissue-specific manner and that there is a family of genes encoding additional calcium channel subtypes.
TYPE OF PUBLICATION: Original article
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(Journal Article): Molecular pharmacology of voltage-dependent calcium channels.
Mori Y
IN:
Jpn J Pharmacol
1996; 72(2):83-109
Impact Factor(s) of Jpn J Pharmacol: 1.724 (2004), 1.23 (2002), 1.347 (2001)
Fulltext: HTML
TYPE OF PUBLICATION: Original article
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(Journal Article): Structure and regulation of voltage-gated Ca2+ channels.
Catterall WA
IN:
Annu Rev Cell Dev Biol
2000; 16(1):521-555
Impact Factor(s) of Annu Rev Cell Dev Biol: 23.69 (2005), 17.804 (2004), 22.638 (2003), 22.87 (2002), 27.106 (2001)
ABSTRACT: Voltage-gated Ca(2+) channels mediate Ca(2+) entry into cells in response to membrane depolarization. Electrophysiological studies reveal different Ca(2+) currents designated L-, N-, P-, Q-, R-, and T-type. The high-voltage-activated Ca(2+) channels that have been characterized biochemically are complexes of a pore-forming alpha1 subunit of approximately 190-250 kDa; a transmembrane, disulfide-linked complex of alpha2 and delta subunits; an intracellular beta subunit; and in some cases a transmembrane gamma subunit. Ten alpha1 subunits, four alpha2delta complexes, four beta subunits, and two gamma subunits are known. The Cav1 family of alpha1 subunits conduct L-type Ca(2+) currents, which initiate muscle contraction, endocrine secretion, and gene transcription, and are regulated primarily by second messenger-activated protein phosphorylation pathways. The Cav2 family of alpha1 subunits conduct N-type, P/Q-type, and R-type Ca(2+) currents, which initiate rapid synaptic transmission and are regulated primarily by direct interaction with G proteins and SNARE proteins and secondarily by protein phosphorylation. The Cav3 family of alpha1 subunits conduct T-type Ca(2+) currents, which are activated and inactivated more rapidly and at more negative membrane potentials than other Ca(2+) current types. The distinct structures and patterns of regulation of these three families of Ca(2+) channels provide a flexible array of Ca(2+) entry pathways in response to changes in membrane potential and a range of possibilities for regulation of Ca(2+) entry by second messenger pathways and interacting proteins.
TYPE OF PUBLICATION: Review
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(Journal Article): Growth and function of the embryonic heart depend upon the cardiac-specific L-type calcium channel alpha1 subunit.
Rottbauer W, Baker K, Wo ZG, Mohideen MA, Cantiello HF, Fishman MC
IN:
Dev Cell
2001; 1(2):265-275
Impact Factor(s) of Dev Cell: 15.434 (2004), 14.807 (2003), 11.531 (2002)
ABSTRACT: The heart must function from the moment of its embryonic assembly, but the molecular underpinnings of the first heart beat are not known, nor whether function determines form at this early stage. Here, we find by positional cloning that the embryonic lethal island beat (isl) mutation in zebrafish disrupts the alpha1 C L-type calcium channel subunit (C-LTCC). The isl atrium is relatively normal in size, and individual cells contract chaotically, in a pattern resembling atrial fibrillation. The ventricle is completely silent. Unlike another mutation with a silent ventricle, isl fails to acquire the normal number of myocytes. Thus, calcium signaling via C-LTCC can regulate heart growth independently of contraction, and plays distinctive roles in fashioning both form and function of the two developing chambers.
TYPE OF PUBLICATION: Original article
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(Journal Article): Direct interaction of gbetagamma with a C-terminal gbetagamma-binding domain of the Ca2+ channel alpha1 subunit is responsible for channel inhibition by G protein-coupled receptors.
Qin N, Platano D, Olcese R, Stefani E, Birnbaumer L
IN:
Proc Natl Acad Sci U S A
1997; 94(16):8866-8871
Impact Factor(s) of Proc Natl Acad Sci U S A: 10.452 (2004), 10.272 (2003), 10.7 (2002), 10.896 (2001)
ABSTRACT: Several classes of voltage-gated Ca2+ channels (VGCCs) are inhibited by G proteins activated by receptors for neurotransmitters and neuromodulatory peptides. Evidence has accumulated to indicate that for non-L-type Ca2+ channels the executing arm of the activated G protein is its betagamma dimer (Gbetagamma). We report below the existence of two Gbetagamma-binding sites on the A-, B-, and E-type alpha1 subunits that form non-L-type Ca2+ channels. One, reported previously, is in loop 1 connecting transmembrane domains I and II. The second is located approximately in the middle of the ca. 600-aa-long C-terminal tails. Both Gbetagamma-binding regions also bind the Ca2+ channel beta subunit (CCbeta), which, when overexpressed, interferes with inhibition by activated G proteins. Replacement in alpha1E of loop 1 with that of the G protein-insensitive and Gbetagamma-binding-negative loop 1 of alpha1C did not abolish inhibition by G proteins, but the exchange of the alpha1E C terminus with that of alpha1C did. This and properties of alpha1E C-terminal truncations indicated that the Gbetagamma-binding site mediating the inhibition of Ca2+ channel activity is the one in the C terminus. Binding of Gbetagamma to this site was inhibited by an alpha1-binding domain of CCbeta, thus providing an explanation for the functional antagonism existing between CCbeta and G protein inhibition. The data do not support proposals that Gbetagamma inhibits alpha1 function by interacting with the site located in the loop I-II linker. These results define the molecular mechanism by which presynaptic G protein-coupled receptors inhibit neurotransmission.
TYPE OF PUBLICATION: Original article
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(Journal Article): Mechanism of SNARE protein binding and regulation of Cav2 channels by phosphorylation of the synaptic protein interaction site.
Yokoyama CT, Myers SJ, Fu J, Mockus SM, Scheuer T, Catterall WA (Department of Pharmacology, University of Washington, Seattle, WA 98195, USA)
IN:
Mol Cell Neurosci
2005; 28(1):1-17
Impact Factor(s) of Mol Cell Neurosci: 3.789 (2004), 4.231 (2003), 5.446 (2001)
ABSTRACT: Ca(v)2.1 and Ca(v)2.2 channels conduct P/Q-type and N-type Ca(2+) currents that initiate neurotransmission and bind SNARE proteins through a synaptic protein interaction (synprint) site. PKC and CaMKII phosphorylate the synprint site and inhibit SNARE protein binding in vitro. Here we identify two separate microdomains that each bind syntaxin 1A and SNAP-25 in vitro and are regulated by PKC phosphorylation at serines 774 and 898 and CaMKII phosphorylation at serines 784 and 896. Activation of PKC resulted in its recruitment to and phosphorylation of Ca(V)2.2 channels, but PKC phosphorylation did not dissociate Ca(V)2.2 channel/syntaxin 1A complexes. Chimeric Ca(V)2.1a channels containing the synprint site of Ca(v)2.2 gain modulation by syntaxin 1A, which is blocked by PKC phosphorylation at the sites identified above. Our results support a bipartite model for the synprint site in which each SNARE-binding microdomain is controlled by a separate PKC and CaMKII phosphorylation site that regulates channel modulation by SNARE proteins.
TYPE OF PUBLICATION: Original article
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(Journal Article): Molecular physiology of low-voltage-activated t-type calcium channels.
Perez-Reyes E (Department of Pharmacology, University of Virginia, Charlottesville, Virginia 22908-0735, USA,
eperez@virginia.edu
)
IN:
Physiol Rev
2003; 83:117-161
Impact Factor(s) of Physiol Rev: 28.721 (2005), 33.918 (2004), 36.831 (2003), 26.532 (2002), 30.061 (2001)
ABSTRACT: T-type Ca2+ channels were originally called low-voltage-activated (LVA) channels because they can be activated by small depolarizations of the plasma membrane. In many neurons Ca2+ influx through LVA channels triggers low-threshold spikes, which in turn triggers a burst of action potentials mediated by Na+ channels. Burst firing is thought to play an important role in the synchronized activity of the thalamus observed in absence epilepsy, but may also underlie a wider range of thalamocortical dysrhythmias. In addition to a pacemaker role, Ca2+ entry via T-type channels can directly regulate intracellular Ca2+ concentrations, which is an important second messenger for a variety of cellular processes. Molecular cloning revealed the existence of three T-type channel genes. The deduced amino acid sequence shows a similar four-repeat structure to that found in high-voltage-activated (HVA) Ca2+ channels, and Na+ channels, indicating that they are evolutionarily related. Hence, the alpha1-subunits of T-type channels are now designated Cav3. Although mRNAs for all three Cav3 subtypes are expressed in brain, they vary in terms of their peripheral expression, with Cav3.2 showing the widest expression. The electrophysiological activities of recombinant Cav3 channels are very similar to native T-type currents and can be differentiated from HVA channels by their activation at lower voltages, faster inactivation, slower deactivation, and smaller conductance of Ba2+. The Cav3 subtypes can be differentiated by their kinetics and sensitivity to block by Ni2+. The goal of this review is to provide a comprehensive description of T-type currents, their distribution, regulation, pharmacology, and cloning.
TYPE OF PUBLICATION: Review
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(Journal Article): Cloning of a novel four repeat protein related to voltage-gated sodium and calcium channels.
Lee JH, Cribbs LL, Perez-Reyes E (Department of Physiology, Loyola University Medical Center, Maywood, IL 60153, USA)
IN:
FEBS Lett
1999; 445(2-3):231-236
Impact Factor(s) of FEBS Lett: 3.843 (2004), 3.609 (2003), 3.912 (2002), 3.644 (2001)
ABSTRACT: Cloning has led to the discovery of more ion channels than predicted by functional studies, yet there remain channels that have not been cloned. We report the cloning of a novel protein that contains the four domain structure found in voltage-gated Ca2+ and Na+ channels. Phylogenetic relationships suggested that the protein might have diverged from an ancestral four repeat channel before the divergence of Ca2+ and Na+ channels. Northern blot analysis showed that mRNA transcripts encoding the protein are expressed predominantly in the brain, moderately in the heart, and weakly in the pancreas. Despite extensive expression attempts, currents from the putative channel were not detected. Based on its sequence, we propose that the novel protein might be a voltage-activated cation channel with unique gating properties.
TYPE OF PUBLICATION: Original article
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(Journal Article): Dihydropyridine-sensitive calcium channels in cardiac and skeletal muscle membranes: studies with antibodies against the alpha subunits.
Takahashi M, Catterall WA (Department of Pharmacology, University of Washington, Seattle 98195, USA)
IN:
Biochemistry
1987; 26(17):5518-5526
Impact Factor(s) of Biochemistry: 4.008 (2004), 3.922 (2003), 4.114 (2001)
ABSTRACT: Polyclonal antibodies (PAC-2) against the purified skeletal muscle calcium channel were prepared and shown to be directed against alpha subunits of this protein by immunoblotting and immunoprecipitation. These polypeptides have an apparent molecular weight of 162,000 without reduction of disulfide bonds. Under conditions where the functional properties of the purified skeletal muscle calcium channel are retained, beta subunits (Mr 50,000) and gamma subunits (Mr 33,000) are coprecipitated, demonstrating specific noncovalent association of these three polypeptides in the purified skeletal muscle channel. PAC-2 immunoprecipitated cardiac calcium channels labeled with [3H]isopropyl 4-(2,1,3-benzoxadiazol-4-yl)-1,4-dihydro-2,6-dimethyl-5- (methoxycarbonyl)pyridine-3-carboxylate ([3H]PN200-110) at a 3-fold higher concentration than skeletal muscle channels. Preincubation with cardiac calcium channels blocked only 49% of the immunoreactivity of PAC-2 toward skeletal muscle channels, indicating that these two proteins have both homologous and distinct epitopes. The immunoreactive component of the cardiac calcium channel was identified by immunoprecipitation and polyacrylamide gel electrophoresis as a polypeptide with an apparent molecular weight of 170,000 before reduction of disulfide bonds and 141,000 after reduction, in close analogy with the properties of the alpha 2 subunits of the skeletal muscle channel. It is concluded that these two calcium channels have a homologous, but distinct, alpha subunit as a major polypeptide component.
TYPE OF PUBLICATION: Original article
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(Journal Article): Differences in apparent pore sizes of low and high voltage-activated Ca2+ channels.
Cataldi M, Perez-Reyes E, Tsien RW (Department of Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California 94305-5345, USA)
IN:
J Biol Chem
2002; 277(48):45969-45976
Impact Factor(s) of J Biol Chem: 6.355 (2004), 6.482 (2003), 7.258 (2001)
ABSTRACT: Pore size is of considerable interest in voltage-gated Ca(2+) channels because they exemplify a fundamental ability of certain ion channels: to display large pore diameter, but also great selectivity for their ion of choice. We determined the pore size of several voltage-dependent Ca(2+) channels of known molecular composition with large organic cations as probes. T-type channels supported by the Ca(V)3.1, Ca(V)3.2, and Ca(V)3.3 subunits; L-type channels encoded by the Ca(V)1.2, beta(1), and alpha(2)delta(1) subunits; and R-type channels encoded by the Ca(V)2.3 and beta(3) subunits were each studied using a Xenopus oocyte expression system. The weak permeabilities to organic cations were resolved by looking at inward tails generated upon repolarization after a large depolarizing pulse. Large inward NH(4)(+) currents and sizable methylammonium and dimethylammonium currents were observed in all of the channels tested, whereas trimethylammonium permeated only through L- and R-type channels, and tetramethylammonium currents were observed only in L-type channels. Thus, our experiments revealed an unexpected heterogeneity in pore size among different Ca(2+) channels, with L-type channels having the largest pore (effective diameter = 6.2 A), T-type channels having the tiniest pore (effective diameter = 5.1 A), and R-type channels having a pore size intermediate between these extremes. These findings ran counter to first-order expectations for these channels based simply on their degree of selectivity among inorganic cations or on the bulkiness of their acidic side chains at the locus of selectivity.
TYPE OF PUBLICATION: Original article
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(Journal Article): Molecular studies of the asymmetric pore structure of the human cardiac voltage-dependent Ca2+ channel.
Klockner U, Mikala G, Schwartz A, Varadi G (Department of Physiology, University of Cologne, Robert-Koch-Strasse 39, 50931 Cologne, Germany)
IN:
J Biol Chem
1996; 271(37):22293-22296
Impact Factor(s) of J Biol Chem: 6.355 (2004), 6.482 (2003), 7.258 (2001)
ABSTRACT: Proton transfer to calcium channels results in rapid fluctuations between two non-zero conductance levels when the current is carried by monovalent cations. A combination of site-directed mutagenesis and single-channel recording techniques were used to identify the unique proton acceptor site as Glu-1086, a conserved glutamate residue located in the S5-S6 linker of motif III of calcium channels. Glu-1086 is part of an array of four glutamate residues in the pore-lining region of the channel conferring the high selectivity of calcium channels. Titration of Glu-1086 yielded a pKa value of 7.91 which is different from that expected for a free glutamic acid side-chain carboxyl. Proposed electrostatic interactions between charged nearby residues can account only in part for this phenomenon since individual elimination of the other three glutamate residues only slightly decreased the pKa of Glu-1086. These data, in addition to identifying the proton acceptor site, provide evidence for the influence of the microenvironment in forming the asymmetry of the conducting pathway of calcium channels.
TYPE OF PUBLICATION: Original article
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(Journal Article): Differential contribution by conserved glutamate residues to an ion-selectivity site in the L-type Ca2+ channel pore.
Mikala G, Bahinski A, Yatani A, Tang S, Schwartz A (Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH 45267-0575, USA)
IN:
FEBS Lett
1993; 335(2):265-269
Impact Factor(s) of FEBS Lett: 3.843 (2004), 3.609 (2003), 3.912 (2002), 3.644 (2001)
ABSTRACT: In voltage-gated cation channels, it is thought that residues responsible for ion-selectivity are located within the pore-lining SS1-SS2 segments. In this study, we compared the ion permeation properties of mutant calcium channels in which highly conserved glutamate residues, located at analogous positions in the SS2 regions of all four motifs, were individually replaced. All of the mutants exhibited a loss of selectivity for divalent over monovalent cations. However, the permeation properties of the individual mutants varied in a position dependent manner. The results provide strong evidence that these glutamate residues, positioned at equivalent locations in the aligned sequences, play significantly different roles in forming the selectivity barrier of the calcium channel, and are probably arranged in an asymmetrical manner inside the ion-conducting pore.
TYPE OF PUBLICATION: Original article
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(Journal Article): Architecture of Ca(2+) channel pore-lining segments revealed by covalent modification of substituted cysteines.
Koch SE, Bodi I, Schwartz A, Varadi G (Institute of Molecular Pharmacology and Biophysics, University of Cincinnati, College of Medicine, Cincinnati, Ohio 45267-0828, USA)
IN:
J Biol Chem
2000; 275(44):34493-34500
Impact Factor(s) of J Biol Chem: 6.355 (2004), 6.482 (2003), 7.258 (2001)
ABSTRACT: The cysteine accessibility method was used to explore calcium channel pore topology. Cysteine mutations were introduced into the SS1-SS2 segments of Motifs I-IV of the human cardiac L-type calcium channel, expressed in Xenopus oocytes and the current block by methanethiosulfonate compounds was measured. Our studies revealed that several consecutive mutants of motifs II and III are accessible to methanethiosulfonates, suggesting that these segments exist as random coils. Motif I cysteine mutants exhibited an intermittent sensitivity to these compounds, providing evidence for a beta-sheet secondary structure. Motif IV showed a periodic sensitivity, suggesting the presence of an alpha-helix. These studies reveal that the SS1-SS2 segment repeat in each motif have non-uniform secondary structures. Thus, the channel architecture evolves as a highly distorted 4-fold pore symmetry.
TYPE OF PUBLICATION: Original article
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(Journal Article): Voltage sensor movements.
Bezanilla F (Department of Physiology, David Geffen School of Medicine at University of California at Los Angeles, Los Angeles, CA 90095, USA)
IN:
J Gen Physiol
2002; 120(4):465-473
Impact Factor(s) of J Gen Physiol: 5.105 (2004), 5.12 (2003), 5.477 (2001)
ABSTRACT: The voltage dependence of K+, Na+, and Ca2+ channels is brought about by a voltage sensor that moves 12–13 e0 across the entire electric field (Schoppa et al., 1992; Hirschberg et al., 1996; Noceti et al., 1996). In the case of Shaker K+ channel it is known which residues are responsible for this large amount of gating charge. This was found by measuring the total gating charge movement per channel after each of the putative charged residues (basic or acidic) were neutralized one by one (Aggarwal and MacKinnon, 1996; Seoh et al., 1996). These studies revealed that the four most extracellularly located basic residues of the S4 segment (R362, R365, R368, and R371) and the most intracellular acidic residue in the S2 segment (E293) are the major contributors to the gating charge movement. The simple assumption that all these residues move across the entire electric field accounts for more than the measured total gating charge. This means that the movement of at least some of the charged residues is only partial within the electric field.
TYPE OF PUBLICATION: Review
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(Journal Article): The principle of gating charge movement in a voltage-dependent K+ channel.
Jiang Y, Ruta V, Chen J, Lee A, MacKinnon R (Howard Hughes Medical Institute, Laboratory of Molecular Neurobiology and Biophysics, Rockefeller University, 1230 York Avenue, New York, New York 10021, USA)
IN:
Nature
2003; 423(6935):42-48
Impact Factor(s) of Nature: 29.273 (2005), 32.182 (2004), 30.979 (2003), 30.432 (2002), 27.955 (2001)
ABSTRACT: The steep dependence of channel opening on membrane voltage allows voltage-dependent K+ channels to turn on almost like a switch. Opening is driven by the movement of gating charges that originate from arginine residues on helical S4 segments of the protein. Each S4 segment forms half of a 'voltage-sensor paddle' on the channel's outer perimeter. Here we show that the voltage-sensor paddles are positioned inside the membrane, near the intracellular surface, when the channel is closed, and that the paddles move a large distance across the membrane from inside to outside when the channel opens. KvAP channels were reconstituted into planar lipid membranes and studied using monoclonal Fab fragments, a voltage-sensor toxin, and avidin binding to tethered biotin. Our findings lead us to conclude that the voltage-sensor paddles operate somewhat like hydrophobic cations attached to levers, enabling the membrane electric field to open and close the pore.
TYPE OF PUBLICATION: Original article
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(Journal Article): Subunits of purified calcium channels.
De Jongh KS, Warner C, Catterall WA (Department of Pharmacology, University of Washington, Seattle 98195, USA)
IN:
J Biol Chem
1990; 265(25):14738-14741
Impact Factor(s) of J Biol Chem: 6.355 (2004), 6.482 (2003), 7.258 (2001)
ABSTRACT: Polyacrylamide gel electrophoresis of purified rabbit skeletal muscle L-type calcium channel before and after reduction of disulfide bonds confirmed that 27- and 24-kDa forms of the delta subunit are disulfide-linked to the 143-kDa alpha 2 subunit. The amino acid sequences of three peptides obtained by tryptic digestion of the delta subunits corresponded to amino acid sequences predicted from the 3' region of the mRNA encoding alpha 2. One of these peptides had the same sequence as the N terminus of the 24- and 27-kDa forms of the delta subunit and corresponded to residues 935-946 of the predicted alpha 2 primary sequence. Anti-peptide antibodies directed to regions on the N-terminal side of this site recognized the 143-kDa alpha 2 subunit in immunoblots of purified calcium channels under reducing conditions, whereas an antipeptide antibody directed toward a sequence on the C-terminal side of this site recognized 24- and 27-kDa forms of the delta subunit. A similar result was obtained after immunoblotting using purified transverse tubules or crude microsomal membrane preparations indicating that alpha 2 and delta occur as distinct disulfide-linked polypeptides in skeletal muscle membranes. Thus, the delta subunits are encoded by the same gene as the alpha 2 subunit and are integral components of the skeletal muscle calcium channel.
TYPE OF PUBLICATION: Original article
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(Journal Article): Molecular cloning and characterization of the human voltage-gated calcium channel alpha(2)delta-4 subunit.
Qin N, Yagel S, Momplaisir ML, Codd EE, D'Andrea MR (Johnson & Johnson Pharmaceutical Research and Development, Spring House, Pennsylvania 19477-0776, USA,
nqin@prius.jnj.com
)
IN:
Mol Pharmacol
2002; 62(3):485-496
Impact Factor(s) of Mol Pharmacol: 5.8 (2004), 5.65 (2003), 5.48 (2002), 5.297 (2001)
ABSTRACT: The voltage-gated calcium channel is composed of a pore-forming alpha(1) subunit and several regulatory subunits: alpha(2)delta, beta, and gamma. We report here the identification of a novel alpha(2)delta subunit, alpha(2)delta-4, from the expressed sequence tag database followed by its cloning and characterization. The novel alpha(2)delta-4 subunit gene contains 39 exons spanning about 130 kilobases and is co-localized with the CHCNA1C gene (alpha(1C) subunit) on human chromosome 12p13.3. Alternative splicing of the alpha(2)delta-4 gene gives rise to four potential variants, a through d. The open reading frame of human alpha(2)delta-4a is composed of 3363 base pairs encoding a protein with 1120 residues and a calculated molecular mass of 126 kDa. The alpha(2)delta-4a subunit shares 30, 32, and 61% identity with the human calcium channel alpha(2)delta-1, alpha(2)delta-2, and alpha(2)delta-3 subunits, respectively. Primary sequence comparison suggests that alpha(2)delta-4 lacks the gabapentin binding motifs characterized for alpha(2)delta-1 and alpha(2)delta-2; this was confirmed by a [(3)H]gabapentin-binding assay. In human embryonic kidney 293 cells, the alpha(2)delta-4 subunit associated with Ca(V)1.2 and beta(3) subunits and significantly increased Ca(V)1.2/beta(3)-mediated Ca(2+) influx. Immunohistochemical study revealed that the alpha(2)delta-4 subunit has limited distribution in special cell types of the pituitary, adrenal gland, colon, and fetal liver. Whether the alpha(2)delta-4 subunit plays a distinct physiological role in select endocrine tissues remains to be demonstrated.
TYPE OF PUBLICATION: Original article
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(Journal Article): Use of transgenic mice to study voltage-dependent Ca2+ channels.
Muth JN, Varadi G, Schwartz A (Institute of Molecular Pharmacology and Biophysics and the Dept of Cell Biology, Neurobiology and Anatomy, PO Box 670828, 231 Albert Sabin Way, Cincinnati, OH 45267-0828, USA,
schwara@email.uc.edu
)
IN:
Trends Pharmacol Sci
2001; 22(10):526-532
Impact Factor(s) of Trends Pharmacol Sci: 13.054 (2004), 13.965 (2003), 13.276 (2002), 11.394 (2001)
ABSTRACT: During the past decade a great number of genes encoding high- and low-voltage-dependent Ca(2+) channels and their accessory subunits have been cloned. Studies of Ca(2+) channel structure-function relationships and channel regulation using cDNA expression in heterologous expression systems have revealed intricate details of subunit interaction, regulation of channels by protein kinase A (PKA) and protein kinase C (PKC), drug binding sites, mechanisms of drug action, the ion conduction pathway and other aspects of channel function. In recent years, however, we have arrived at the brink of an entirely new strategy to study Ca(2+) channels by overexpressing or knocking out genes encoding these channels in transgenic mice. In this article, various models of gene knockout or gene overexpression will be discussed. This new approach will reveal many secrets regarding Ca(2+) channel regulation and the control of Ca(2+)-dependent cellular processes.
TYPE OF PUBLICATION: Review
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(Journal Article): Molecular basis for Ca2+ channel diversity.
Hofmann F, Biel M, Flockerzi V (Institut fur Pharmakologie und Toxikologie, TU Munchen, Germany)
IN:
Annu Rev Neurosci
1994; 17(1):399-418
Impact Factor(s) of Annu Rev Neurosci: 24.184 (2005), 23.143 (2004), 30.167 (2003), 24.091 (2002), 27.152 (2001)
ABSTRACT: n/a
TYPE OF PUBLICATION: Review
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(Journal Article): Acceleration of activation and inactivation by the beta subunit of the skeletal muscle calcium channel.
Varadi G, Lory P, Schultz D, Varadi M, Schwartz A (Department of Pharmacology and Cell Biophysics, University of Cincinnati, Ohio 45267-0575, USA)
IN:
Nature
1991; 352(6331):159-162
Impact Factor(s) of Nature: 29.273 (2005), 32.182 (2004), 30.979 (2003), 30.432 (2002), 27.955 (2001)
ABSTRACT: The L-type voltage-dependent calcium channel is an important link in excitation-contraction coupling of muscle cells (reviewed in refs 2 and 3). The channel has two functional characteristics: calcium permeation and receptor sites for calcium antagonists. In skeletal muscle the channel is a complex of five subunits, alpha 1, alpha 2, beta, gamma and delta. Complementary DNAs to these subunits have been cloned and their amino-acid sequences deduced. The skeletal muscle alpha 1 subunit cDNA expressed in L cells manifests as specific calcium-ion permeation, as well as sensitivity to the three classes of organic calcium-channel blockers. We report here that coexpression of the alpha 1 subunit with other subunits results in significant changes in dihydropyridine binding and gating properties. The available number of drug receptor sites increases 10-fold with an alpha 1 beta combination, whereas the affinity of the dihydropyridine binding site remains unchanged. Also, the presence of the beta subunit accelerates activation and inactivation kinetics of the calcium-channel current.
TYPE OF PUBLICATION: Original article
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(Journal Article): Gabapentin inhibits high-threshold calcium channel currents in cultured rat dorsal root ganglion neurones.
Sutton KG, Martin DJ, Pinnock RD, Lee K, Scott RH (Department of Biology, Pfizer Global R&D, Cambridge Laboratories, Cambridge, CB2 2QB, UK,
Kathy_sutton@merck.com
)
IN:
Br J Pharmacol
2002; 135(1):257-265
Impact Factor(s) of Br J Pharmacol: 3.325 (2004), 3.611 (2003), 3.45 (2002), 3.502 (2001)
ABSTRACT: 1. This study examined the action of gabapentin (gabapentin,1-(aminomethyl) cyclohexane acetic acid (Neurontin) on voltage-gated calcium (Ca(2+)) channel influx recorded in cultured rat dorsal root ganglion (DRG) neurones. 2. Voltage-gated Ca(2+) influx was monitored using both fura-2 based fluorescence Ca(2+) imaging and the whole-cell patch clamp technique. 3. Imaging of intracellular Ca(2+) transients revealed that gabapentin inhibited KCl (30 mM)-evoked voltage-dependent Ca(2+) influx. Both the duration for 50% of the maximum response (W50) and total Ca(2+) influx were significantly reduced by approximately 25-30% in the presence of gabapentin (25 microM). 4. Gabapentin potently inhibited the peak whole-cell Ca(2+) channel current (I(Ba)) in a dose-dependent manner with an estimated IC(50) value of 167 nM. Block was incomplete and saturated at a maximal concentration of 25 microM. 5. Inhibition was significantly decreased in the presence of the neutral amino acid L-isoleucine (25 microM) but unaffected by application of the GABA(B) antagonist, saclofen (200 microM), suggesting a direct action on the alpha(2)delta subunit of the Ca(2+) channel. 6. Gabapentin inhibition was voltage-dependent, producing an approximately 7 mV hyperpolarizing shift in current voltage properties and reducing a non-inactivating component of whole-cell current activated at relatively depolarized potentials. 7. The use of specific Ca(2+) channel antagonists revealed a mixed pharmacology of the gabapentin-sensitive current (N-, L- and P/Q-type), which is dominated by N-type current. 8. The present study is the first to demonstrate that gabapentin directly mediates inhibition of voltage-gated Ca(2+) influx in DRG neurones, providing a potential means for gabapentin to effectively mediate spinal anti-nociception.
TYPE OF PUBLICATION: Original article
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(Journal Article): Calcium channel alpha(2)delta subunits-structure and Gabapentin binding.
Marais E, Klugbauer N, Hofmann F (Institut fur Pharmakologie und Toxikologie der Technischen Universitat Munchen, Munchen, Germany)
IN:
Mol Pharmacol
2001; 59(5):1243-1248
Impact Factor(s) of Mol Pharmacol: 5.8 (2004), 5.65 (2003), 5.48 (2002), 5.297 (2001)
ABSTRACT: High-voltage activated calcium channels are modulated by a series of auxiliary proteins, including those of the alpha(2)delta family. Until recently, only a single alpha(2)delta subunit was known, but two further members, alpha(2)delta-2 and -3, have since been identified. In this study, the structure of these two novel subunits has been characterized and binding of the antiepileptic drug gabapentin investigated. Using antibodies directed against the amino terminal portion of the proteins, the gross structure of the subunits could be analyzed by Western blotting. Similar to alpha(2)delta-1, both alpha(2)delta-2 and -3 subunits consist of two proteins-a larger alpha(2) and a smaller delta that can be separated by reduction. The subunits are also highly N-glycosylated with approximately 30 kDa of their mass consisting of oligosaccharides. alpha(2)delta-1 was detected in all mouse tissues studied, whereas alpha(2)delta-2 was found at high levels in brain and heart. The alpha(2)delta-3 subunit was observed only in brain. alpha(2)delta-1 and alpha(2)delta-2, but not alpha(2)delta-3, were found to bind gabapentin. The K(d) value of gabapentin binding to alpha(2)delta-2 was 153 nM compared with the higher affinity binding to alpha(2)delta-1 (K(d) = 59 nM).
TYPE OF PUBLICATION: Original article
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(Journal Article): Injury type-specific calcium channel alpha 2 delta-1 subunit up-regulation in rat neuropathic pain models correlates with antiallodynic effects of gabapentin.
Luo ZD, Calcutt NA, Higuera ES, Valder CR, Song YH, Svensson CI, Myers RR (Department of Anesthesiology, University of California San Diego, La Jolla, California 92093-0818, USA,
zluo@ucsd.edu
)
IN:
J Pharmacol Exp Ther
2002; 303(3):1199-1205
Impact Factor(s) of J Pharmacol Exp Ther: 4.335 (2004), 4.337 (2003), 3.555 (2001)
ABSTRACT: The calcium channel alpha2delta-1 subunit is a structural subunit important for functional calcium channel assembly. In vitro studies have shown that this subunit is the binding site for gabapentin, an anticonvulsant that exerts antihyperalgesic effects by unknown mechanisms. Increased expression of this subunit in the spinal cord and dorsal root ganglia (DRG) has been suggested to play a role in enhanced nociceptive responses of spinal nerve-injured rats to innocuous mechanical stimulation (allodynia). To investigate whether a common mechanism underlies allodynic states derived from different etiologies, and if so, whether similar alpha2delta-1 subunit up-regulation correlates with these allodynic states, we compared DRG and spinal cord alpha2delta-1 subunit levels and gabapentin sensitivity in allodynic rats with mechanical nerve injuries (sciatic nerve chronic constriction injury, spinal nerve transection, or ligation), a metabolic disorder (diabetes), or chemical neuropathy (vincristine neurotoxicity). Our data indicated that even though allodynia occurred in all types of nerve injury investigated, DRG and/or spinal cord alpha2delta-1 subunit up-regulation and gabapentin sensitivity only coexisted in the mechanical and diabetic neuropathies. Thus, induction of the alpha2delta-1 subunit in the DRG and spinal cord is likely regulated by factors that are specific for individual neuropathies and may contribute to gabapentin-sensitive allodynia. However, the calcium channel alpha2delta-1 subunit is not the sole molecular change that uniformly characterizes the neuropathic pain states.
TYPE OF PUBLICATION: Original article
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(Journal Article): Cerebellar ataxia, seizures, premature death, and cardiac abnormalities in mice with targeted disruption of the Cacna2d2 gene.
Ivanov SV, Ward JM, Tessarollo L, McAreavey D, Sachdev V, Fananapazir L, Banks MK, Morris N, Djurickovic D, Devor-Henneman DE, Wei MH, Alvord GW, Gao B, Richardson JA, Minna JD, Rogawski MA, Lerman MI (Basic Research Program, Science Applications International Corporation-Frederick, Inc., National Cancer Institute at Frederick, Frederick, Maryland, USA,
ivanov@mail.ncifcrf.gov
)
IN:
Am J Pathol
2004; 165(3):1007-1018
Impact Factor(s) of Am J Pathol: 6.441 (2004), 6.946 (2003), 6.75 (2002), 7.103 (2001)
ABSTRACT: CACNA2D2 is a putative tumor suppressor gene located in the human chromosome 3p21.3 region that shows frequent allelic imbalances in lung, breast, and other cancers. The alpha2delta-2 protein encoded by the gene is a regulatory subunit of voltage-dependent calcium channels and is expressed in brain, heart, and other tissues. Here we report that mice homozygous for targeted disruption of the Cacna2d2 gene exhibit growth retardation, reduced life span, ataxic gait with apoptosis of cerebellar granule cells followed by Purkinje cell depletion, enhanced susceptibility to seizures, and cardiac abnormalities. The Cacna2d2(tm1NCIF) null phenotype has much in common with that of Cacna1a mutants, such as cerebellar neuro-degeneration associated with ataxia, seizures, and premature death. A tendency to bradycardia and limited response of null mutants to isoflurane implicate alpha2delta-2 in sympathetic regulation of cardiac function. In summary, our findings provide genetic evidence that the alpha2delta-2 subunit serves in vivo as a component of P/Q-type calcium channels, is indispensable for the central nervous system function, and may be involved in hereditary cerebellar ataxias and epileptic disorders in humans.
TYPE OF PUBLICATION: Original article
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(Journal Article): Structure-functional diversity of human L-type Ca2+ channel: perspectives for new pharmacological targets.
Abernethy DR, Soldatov NM (Section on Molecular and Clinical Pharmacology, Laboratory of Clinical Investigation, National Institute on Aging, Intramural Research Program, National Institutes of Health, Baltimore, Maryland, USA,
soldatovn@grc.nia.nih.gov
)
IN:
J Pharmacol Exp Ther
2002; 300(3):724-728
Impact Factor(s) of J Pharmacol Exp Ther: 4.335 (2004), 4.337 (2003), 3.555 (2001)
ABSTRACT: The L-type Ca2+ channels mediate depolarization-induced influx of Ca2+ into a wide variety of cells and thus play a central role in triggering cardiac and smooth muscle contraction. Because of this role, clinically important classes of 1,4-dihydropyridine, phenylalkylamine, and benzothiazepine Ca2+ channel blockers were developed as powerful medicines to treat hypertension and angina pectoris. Molecular cloning studies revealed that the channel is subject to extensive structure-functional variability due to alternative splicing. In this review, we will focus on a potentially important role of genetically driven variability of Ca2+ channels in expression regulation and mutations, Ca2+-induced inactivation, and modulation of sensitivity to Ca2+ channel blockers with the perspective for new pharmacological targets.
TYPE OF PUBLICATION: Review
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(Journal Article): Auxiliary subunits: essential components of the voltage-gated calcium channel complex
Arikkath J, Campbell KP (Howard Hughes Medical Institute and Department of Physiology and Biophysics, University of Iowa, College of Medicine, 400 Eckstein Medical Research Building, Iowa City, IA 52242, USA,
kevin-campbell@uiowa.edu
)
IN:
Curr Opin Neurobiol
2003; 13(3):298-307
Impact Factor(s) of Curr Opin Neurobiol: 7.937 (2004), 9.727 (2003), 10.718 (2002), 9.995 (2001)
ABSTRACT: Voltage-gated calcium channels are important mediators of several physiological processes, including neuronal excitability and muscle contraction. At the molecular level, the channels are composed of four subunits--the pore forming alpha(1) subunit and the auxiliary alpha(2)delta, beta and gamma subunits. The auxiliary subunits modulate the trafficking and the biophysical properties of the alpha(1) subunit. In the past several years there has been an acceleration of our understanding of the auxiliary subunits, primarily because of their molecular characterization and the availability of spontaneous and targeted mouse mutants. These studies have revealed the crucial role of the subunits in the functional effects that are mediated by voltage-gated calcium channels.
TYPE OF PUBLICATION: Review
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(Journal Article): Gamma subunit of voltage-activated calcium channels.
Kang MG, Campbell KP (Howard Hughes Medical Institute, Department of Physiology and Biophysics, University of Iowa College of Medicine, Iowa City, IA 52242, USA,
kevin-campbell@uiowa.edu
)
IN:
J Biol Chem
2003; 278(24):21315-21318
Impact Factor(s) of J Biol Chem: 6.355 (2004), 6.482 (2003), 7.258 (2001)
ABSTRACT: Voltage-activated Ca2+ channels play a major role in many fundamental physiological processes including neurotransmission, muscle contraction, intracellular signaling, hormone secretion, and development. Therefore, an understanding of the structure and regulation of Ca2+ channels is critical for the comprehension of these physiological phenomena. Five types of high voltage-activated Ca2+ channels (named L-, N-, P-, Q-, and R-type) and one type of low voltage-activated Ca2+ channel (known as T-type) have been identified based on the pharmacological and biophysical characteristics of native currents.
TYPE OF PUBLICATION: Review
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