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PMID: 12042350 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Novel functional properties of Ca(2+) channel beta subunits revealed by their expression in adult rat heart cells.

The Journal of physiology ·Vol. 541 ·No. Pt 2 ·2002-06-01 ·Pages 435-52

Colecraft HM, Alseikhan B, Takahashi SX, Chaudhuri D, Mittman S, Yegnasubramanian V, Alvania RS, Johns DC, Marbán E, Yue DT

Abstract

Recombinant adenoviruses were used to overexpress green fluorescent protein (GFP)-fused auxiliary Ca(2+) channel beta subunits (beta(1)-beta(4)) in cultured adult rat heart cells, to explore new dimensions of beta subunit functions in vivo. Distinct beta-GFP subunits distributed differentially between the surface sarcolemma, transverse elements, and nucleus in single heart cells. All beta-GFP subunits increased the native cardiac whole-cell L-type Ca(2+) channel current density, but produced distinctive effects on channel inactivation kinetics. The degree of enhancement of whole-cell current density was non-uniform between beta subunits, with a rank order of potency beta(2a) approximately equal to beta(4) > beta(1b) > beta(3). For each beta subunit, the increase in L-type current density was accompanied by a correlative increase in the maximal gating charge (Q(max)) moved with depolarization. However, beta subunits produced characteristic effects on single L-type channel gating, resulting in divergent effects on channel open probability (P(o)). Quantitative analysis and modelling of single-channel data provided a kinetic signature for each channel type. Spurred on by ambiguities regarding the molecular identity of the actual endogenous cardiac L-type channel beta subunit, we cloned a new rat beta(2) splice variant, beta(2b), from heart using 5' rapid amplification of cDNA ends (RACE) PCR. By contrast with beta(2a), expression of beta(2b) in heart cells yielded channels with a microscopic gating signature virtually identical to that of native unmodified channels. Our results provide novel insights into beta subunit functions that are unattainable in traditional heterologous expression studies, and also provide new perspectives on the molecular identity of the beta subunit component of cardiac L-type Ca(2+) channels. Overall, the work establishes a powerful experimental paradigm to explore novel functions of ion channel subunits in their native environments.

MeSH Terms
Adenoviridae/metabolism Animals Blotting, Western Calcium Channels/biosynthesis,metabolism Calcium Channels, L-Type/drug effects DNA Primers Electrophysiology Genetic Vectors Ion Channel Gating/drug effects Membrane Potentials/physiology Models, Neurological Myocardium/chemistry,metabolism Patch-Clamp Techniques Plasmids Rats Recombinant Proteins/biosynthesis Reverse Transcriptase Polymerase Chain Reaction Sarcolemma/drug effects,metabolism Transfection
Chemicals
Cacnb3 protein, rat Cacnb4 protein, rat Calcium Channels Calcium Channels, L-Type DNA Primers Recombinant Proteins
Authors & Affiliations
10 authors, click to expand affiliations / ORCID
Colecraft Henry M
Program in Molecular and Cellular Systems Physiology, Department of Biomedical Engineering, Johns Hopkins University School of Medicine, Traylor Building, Room 710A, 720 Rutland Avenue, Baltimore, MD 21205, USA. [email protected]
Alseikhan Badr
Takahashi Shoji X
Chaudhuri Dipayan
Mittman Scott
Yegnasubramanian Vasan
Alvania Rebecca S
Johns David C
Marbán Eduardo
Yue David T
References (82)
82 references, click to expand
  1. Voltage-dependent gating of Shaker A-type potassium channels in Drosophila muscle.
    J Gen Physiol. 1990 Jan;95(1):29-60 PMID: 2299331
  2. Beta-adrenergic stimulation of calcium channels occurs by potentiation of high-activity gating modes.
    Proc Natl Acad Sci U S A. 1990 Jan;87(2):753-7 PMID: 1689051
  3. Intramembrane charge movement in guinea-pig and rat ventricular myocytes.
    J Physiol. 1989 Aug;415:601-24 PMID: 2484211
  4. Two distinct functional effects of protein phosphatase inhibitors on guinea-pig cardiac L-type Ca2+ channels.
    J Physiol. 1995 May 1;484 ( Pt 3):583-92 PMID: 7623278
  5. Adenovirus-mediated expression of a voltage-gated potassium channel in vitro (rat cardiac myocytes) and in vivo (rat liver). A novel strategy for modifying excitability.
    J Clin Invest. 1995 Aug;96(2):1152-8 PMID: 7635952
  6. The nuclear pore complex.
    Annu Rev Biochem. 1995;64:865-96 PMID: 7574503
  7. Ca(2+) release mechanisms, Ca(2+) sparks, and local control of excitation-contraction coupling in normal heart muscle.
    Circ Res. 1999 Oct 29;85(9):770-6 PMID: 10532944
  8. Cloning of the beta(2a) subunit of the voltage-dependent calcium channel from human heart: cooperative effect of alpha(2)/delta and beta(2a) on the membrane expression of the alpha(1C) subunit.
    Biochem Biophys Res Commun. 2000 Jan 7;267(1):156-63 PMID: 10623591
  9. Expression of Ca(2+) channel subunits during cardiac ontogeny in mice and rats: identification of fetal alpha(1C) and beta subunit isoforms.
    J Cell Biochem. 2000 Jan;76(4):695-703 PMID: 10653988
  10. Differential occurrence of reluctant openings in G-protein-inhibited N- and P/Q-type calcium channels.
    J Gen Physiol. 2000 Feb;115(2):175-92 PMID: 10653895
  11. Complexes of the alpha1C and beta subunits generate the necessary signal for membrane targeting of class C L-type calcium channels.
    J Biol Chem. 1999 Jan 22;274(4):2137-44 PMID: 9890976
  12. Voltage sensors in domains III and IV, but not I and II, are immobilized by Na+ channel fast inactivation.
    Neuron. 1999 Jan;22(1):73-87 PMID: 10027291
  13. Inducible genetic suppression of neuronal excitability.
    J Neurosci. 1999 Mar 1;19(5):1691-7 PMID: 10024355
  14. Differential plasma membrane targeting of voltage-dependent calcium channel subunits expressed in a polarized epithelial cell line.
    J Physiol. 1999 Mar 15;515 ( Pt 3):685-94 PMID: 10066897
  15. Differential regulation of skeletal muscle L-type Ca2+ current and excitation-contraction coupling by the dihydropyridine receptor beta subunit.
    Biophys J. 1999 Apr;76(4):1744-56 PMID: 10096875
  16. Subunit expression of the cardiac L-type calcium channel is differentially regulated in diastolic heart failure of the cardiac allograft.
    Circulation. 1999 Jul 13;100(2):155-63 PMID: 10402445
  17. Cardiac-specific overexpression of the alpha(1) subunit of the L-type voltage-dependent Ca(2+) channel in transgenic mice. Loss of isoproterenol-induced contraction.
    J Biol Chem. 1999 Jul 30;274(31):21503-6 PMID: 10419451
  18. Atrial L-type Ca2+ currents and human atrial fibrillation.
    Circ Res. 1999 Sep 3;85(5):428-36 PMID: 10473672
  19. Subunit regulation of the neuronal alpha 1A Ca2+ channel expressed in Xenopus oocytes.
    J Physiol. 1995 Jun 15;485 ( Pt 3):619-34 PMID: 7562605
  20. Roles of a membrane-localized beta subunit in the formation and targeting of functional L-type Ca2+ channels.
    J Biol Chem. 1995 Dec 15;270(50):30036-44 PMID: 8530407
  21. Ventricular action potential and L-type calcium channel in infarct-induced hypertrophy in rats.
    J Cardiovasc Electrophysiol. 1995 Nov;6(11):1004-14 PMID: 8589869
  22. Normalization of current kinetics by interaction between the alpha 1 and beta subunits of the skeletal muscle dihydropyridine-sensitive Ca2+ channel.
    Nature. 1991 Aug 8;352(6335):527-30 PMID: 1650913
  23. The roles of the subunits in the function of the calcium channel.
    Science. 1991 Sep 27;253(5027):1553-7 PMID: 1716787
  24. Cloning and tissue-specific expression of the brain calcium channel beta-subunit.
    FEBS Lett. 1991 Oct 21;291(2):253-8 PMID: 1657644
  25. Heterologous regulation of the cardiac Ca2+ channel alpha 1 subunit by skeletal muscle beta and gamma subunits. Implications for the structure of cardiac L-type Ca2+ channels.
    J Biol Chem. 1991 Nov 15;266(32):21943-7 PMID: 1718988
  26. Cloning and expression of a cardiac/brain beta subunit of the L-type calcium channel.
    J Biol Chem. 1992 Jan 25;267(3):1792-7 PMID: 1370480
  27. Calcium channel beta subunit heterogeneity: functional expression of cloned cDNA from heart, aorta and brain.
    EMBO J. 1992 Mar;11(3):885-90 PMID: 1312465
  28. Modulation of N-type calcium channels in bullfrog sympathetic neurons by luteinizing hormone-releasing hormone: kinetics and voltage dependence.
    J Neurosci. 1993 Feb;13(2):516-33 PMID: 7678856
  29. Cloning and expression of a third calcium channel beta subunit.
    J Biol Chem. 1993 Feb 15;268(5):3450-5 PMID: 7679112
  30. Cloning and characterization of a Lambert-Eaton myasthenic syndrome antigen.
    Ann Neurol. 1993 Jan;33(1):113-20 PMID: 8494331
  31. Molecular cloning of three isoforms of the L-type voltage-dependent calcium channel beta subunit from normal human heart.
    Circ Res. 1993 Jun;72(6):1337-44 PMID: 7916667
  32. Cloning and expression of a neuronal calcium channel beta subunit.
    J Biol Chem. 1993 Jun 15;268(17):12359-66 PMID: 7685340
  33. Potentiation by the beta subunit of the ratio of the ionic current to the charge movement in the cardiac calcium channel.
    Science. 1993 Oct 22;262(5133):575-8 PMID: 8211185
  34. Functional properties of a neuronal class C L-type calcium channel.
    Neuropharmacology. 1993 Nov;32(11):1117-26 PMID: 8107966
  35. Regulation and modulation of calcium channels in cardiac, skeletal, and smooth muscle cells.
    Physiol Rev. 1994 Apr;74(2):365-507 PMID: 8171118
  36. Mechanism of Ca(2+)-sensitive inactivation of L-type Ca2+ channels.
    Neuron. 1994 Jun;12(6):1301-18 PMID: 8011340
  37. Ca2+ channel regulation by a conserved beta subunit domain.
    Neuron. 1994 Aug;13(2):495-503 PMID: 8060623
  38. The amino terminus of a calcium channel beta subunit sets rates of channel inactivation independently of the subunit's effect on activation.
    Neuron. 1994 Dec;13(6):1433-8 PMID: 7993634
  39. Reduced calcium current density in single myocytes isolated from hypertrophied failing guinea pig hearts.
    J Mol Cell Cardiol. 1994 Sep;26(9):1133-43 PMID: 7815457
  40. Dual activation of the cardiac Ca2+ channel alpha 1C-subunit and its modulation by the beta-subunit.
    Am J Physiol. 1995 Mar;268(3 Pt 1):C732-40 PMID: 7900777
  41. Comparison of fura-2 imaging and electrophysiological analysis of murine calcium channel alpha 1 subunits coexpressed with novel beta 2 subunit isoforms.
    Mol Pharmacol. 1995 Apr;47(4):707-16 PMID: 7723731
  42. Molecular architecture of membranes involved in excitation-contraction coupling of cardiac muscle.
    J Cell Biol. 1995 May;129(3):659-71 PMID: 7730402
  43. Immunolocalization of sarcolemmal dihydropyridine receptor and sarcoplasmic reticular triadin and ryanodine receptor in rabbit ventricle and atrium.
    J Cell Biol. 1995 May;129(3):673-82 PMID: 7730403
  44. Calcium currents in diseased human cardiac cells.
    J Cardiovasc Pharmacol. 1995 Feb;25(2):282-91 PMID: 7752654
  45. The control of calcium release in heart muscle.
    Science. 1995 May 19;268(5213):1045-9 PMID: 7754384
  46. The beta subunit increases Ca2+ currents and gating charge movements of human cardiac L-type Ca2+ channels.
    Biophys J. 1996 Mar;70(3):1285-93 PMID: 8785284
  47. Enhancement of ionic current and charge movement by coexpression of calcium channel beta 1A subunit with alpha 1C subunit in a human embryonic kidney cell line.
    J Physiol. 1996 Apr 1;492 ( Pt 1):89-96 PMID: 8730585
  48. Identification of palmitoylation sites within the L-type calcium channel beta2a subunit and effects on channel function.
    J Biol Chem. 1996 Oct 25;271(43):26465-8 PMID: 8900112
  49. Coupling between charge movement and pore opening in vertebrate neuronal alpha 1E calcium channels.
    J Physiol. 1996 Dec 15;497 ( Pt 3):675-86 PMID: 9003553
  50. Regional expression and cellular localization of the alpha1 and beta subunit of high voltage-activated calcium channels in rat brain.
    J Neurosci. 1997 Feb 15;17(4):1339-49 PMID: 9006977
  51. Construction of adenovirus vectors through Cre-lox recombination.
    J Virol. 1997 Mar;71(3):1842-9 PMID: 9032314
  52. Bursts of action potential waveforms relieve G-protein inhibition of recombinant P/Q-type Ca2+ channels in HEK 293 cells.
    J Physiol. 1997 Mar 15;499 ( Pt 3):637-44 PMID: 9130160
  53. Importance of the different beta subunits in the membrane expression of the alpha1A and alpha2 calcium channel subunits: studies using a depolarization-sensitive alpha1A antibody.
    Eur J Neurosci. 1997 Apr;9(4):749-59 PMID: 9153581
  54. Beta subunit heterogeneity in neuronal L-type Ca2+ channels.
    J Biol Chem. 1997 May 23;272(21):13877-82 PMID: 9153247
  55. Suppression of neuronal and cardiac transient outward currents by viral gene transfer of dominant-negative Kv4.2 constructs.
    J Biol Chem. 1997 Dec 12;272(50):31598-603 PMID: 9395498
  56. Unique regulatory properties of the type 2a Ca2+ channel beta subunit caused by palmitoylation.
    Proc Natl Acad Sci U S A. 1998 Apr 14;95(8):4690-5 PMID: 9539800
  57. Subunit interaction sites in voltage-dependent Ca2+ channels: role in channel function.
    Trends Neurosci. 1998 Apr;21(4):148-54 PMID: 9554724
  58. Effects of temperature on human L-type cardiac Ca2+ channels expressed in Xenopus oocytes.
    Pflugers Arch. 1998 Jul;436(2):238-47 PMID: 9594024
  59. Changes in L-type calcium channel abundance and function during the transition to pacing-induced congestive heart failure.
    Cardiovasc Res. 1998 Feb;37(2):432-44 PMID: 9614498
  60. Preferential closed-state inactivation of neuronal calcium channels.
    Neuron. 1998 May;20(5):1027-38 PMID: 9620706
  61. Mechanism of auxiliary subunit modulation of neuronal alpha1E calcium channels.
    J Gen Physiol. 1998 Aug;112(2):125-43 PMID: 9689023
  62. Membrane targeting of L-type calcium channels. Role of palmitoylation in the subcellular localization of the beta2a subunit.
    J Biol Chem. 1998 Sep 4;273(36):23590-7 PMID: 9722599
  63. Increased availability and open probability of single L-type calcium channels from failing compared with nonfailing human ventricle.
    Circulation. 1998 Sep 8;98(10):969-76 PMID: 9737516
  64. Structures and functions of calcium channel beta subunits.
    J Bioenerg Biomembr. 1998 Aug;30(4):357-75 PMID: 9758332
  65. beta subunit heterogeneity of L-type Ca(2+) channels in smooth muscle tissues.
    FEBS Lett. 2000 Feb 4;467(1):65-9 PMID: 10664458
  66. Ca(2+) channel modulation by recombinant auxiliary beta subunits expressed in young adult heart cells.
    Circ Res. 2000 Feb 4;86(2):175-84 PMID: 10666413
  67. Structure and regulation of voltage-gated Ca2+ channels.
    Annu Rev Cell Dev Biol. 2000;16:521-55 PMID: 11031246
  68. Immunogold-labeled L-type calcium channels are clustered in the surface plasma membrane overlying junctional sarcoplasmic reticulum in guinea-pig myocytes-implications for excitation-contraction coupling in cardiac muscle.
    J Mol Cell Cardiol. 2000 Nov;32(11):1981-94 PMID: 11040103
  69. Distribution of proteins implicated in excitation-contraction coupling in rat ventricular myocytes.
    Biophys J. 2000 Nov;79(5):2682-91 PMID: 11053140
  70. The [beta]2a subunit is a molecular groom for the Ca2+ channel inactivation gate.
    J Neurosci. 2000 Dec 15;20(24):9046-52 PMID: 11124981
  71. Atrial L-type Ca2+-channel, beta-adrenorecptor, and 5-hydroxytryptamine type 4 receptor mRNAs in human atrial fibrillation.
    Basic Res Cardiol. 2001 Feb;96(1):82-90 PMID: 11215536
  72. Downregulation of the voltage-dependent calcium channel (VDCC) beta-subunit mRNAs in pancreatic islets of type 2 diabetic rats.
    Biochem Biophys Res Commun. 2001 Jan 26;280(3):923-32 PMID: 11162613
  73. Reduction in density of transverse tubules and L-type Ca(2+) channels in canine tachycardia-induced heart failure.
    Cardiovasc Res. 2001 Feb 1;49(2):298-307 PMID: 11164840
  74. G-protein inhibition of N- and P/Q-type calcium channels: distinctive elementary mechanisms and their functional impact.
    J Neurosci. 2001 Feb 15;21(4):1137-47 PMID: 11160384
  75. Mechanisms of L-type Ca(2+) current downregulation in rat atrial myocytes during heart failure.
    Circ Res. 2001 Sep 28;89(7):607-13 PMID: 11577026
  76. Cloning of a functional splice variant of L-type calcium channel beta 2 subunit from rat heart.
    J Biol Chem. 2001 Dec 14;276(50):47163-70 PMID: 11604404
  77. Cleavage of structural proteins during the assembly of the head of bacteriophage T4.
    Nature. 1970 Aug 15;227(5259):680-5 PMID: 5432063
  78. Inactivation of the sodium channel. II. Gating current experiments.
    J Gen Physiol. 1977 Nov;70(5):567-90 PMID: 591912
  79. Calcium-induced release of calcium from the cardiac sarcoplasmic reticulum.
    Am J Physiol. 1983 Jul;245(1):C1-14 PMID: 6346892
  80. Different modes of Ca channel gating behaviour favoured by dihydropyridine Ca agonists and antagonists.
    Nature. 1984 Oct 11-17;311(5986):538-44 PMID: 6207437
  81. Primary structure of the beta subunit of the DHP-sensitive calcium channel from skeletal muscle.
    Science. 1989 Sep 8;245(4922):1115-8 PMID: 2549640
  82. Nonlinear charge movement in mammalian cardiac ventricular cells. Components from Na and Ca channel gating.
    J Gen Physiol. 1989 Jul;94(1):65-93 PMID: 2553859
Article Info
Journal
The Journal of physiology
Abbr.
J Physiol
ISSN
0022-3751
DOI
10.1113/jphysiol.2002.018515
Email
[email protected]
Published
2002-06-01
Pages
435-52
Language
English
Region
England
NLM ID
0266262
PMCID
PMC2290333
Subset
IM
External Links
PubMed source DOI: 10.1113/jphysiol.2002.018515 PMC Full Text View journal details
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