scholarly journals Rem uncouples excitation–contraction coupling in adult skeletal muscle fibers

2015 ◽  
Vol 146 (1) ◽  
pp. 97-108 ◽  
Author(s):  
Donald Beqollari ◽  
Christin F. Romberg ◽  
Dilyana Filipova ◽  
Ulises Meza ◽  
Symeon Papadopoulos ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Direct Interaction ◽  
Charge Movement ◽  
Triple Mutant ◽  
Membrane Expression ◽  
Direct Role ◽  
Release Channel ◽  
Adult Skeletal Muscle ◽  
Ec Coupling ◽  
Conformational Coupling

In skeletal muscle, excitation–contraction (EC) coupling requires depolarization-induced conformational rearrangements in L-type Ca2+ channel (CaV1.1) to be communicated to the type 1 ryanodine-sensitive Ca2+ release channel (RYR1) of the sarcoplasmic reticulum (SR) via transient protein–protein interactions. Although the molecular mechanism that underlies conformational coupling between CaV1.1 and RYR1 has been investigated intensely for more than 25 years, the question of whether such signaling occurs via a direct interaction between the principal, voltage-sensing α1S subunit of CaV1.1 and RYR1 or through an intermediary protein persists. A substantial body of evidence supports the idea that the auxiliary β1a subunit of CaV1.1 is a conduit for this intermolecular communication. However, a direct role for β1a has been difficult to test because β1a serves two other functions that are prerequisite for conformational coupling between CaV1.1 and RYR1. Specifically, β1a promotes efficient membrane expression of CaV1.1 and facilitates the tetradic ultrastructural arrangement of CaV1.1 channels within plasma membrane–SR junctions. In this paper, we demonstrate that overexpression of the RGK protein Rem, an established β subunit–interacting protein, in adult mouse flexor digitorum brevis fibers markedly reduces voltage-induced myoplasmic Ca2+ transients without greatly affecting CaV1.1 targeting, intramembrane gating charge movement, or releasable SR Ca2+ store content. In contrast, a β1a-binding–deficient Rem triple mutant (R200A/L227A/H229A) has little effect on myoplasmic Ca2+ release in response to membrane depolarization. Thus, Rem effectively uncouples the voltage sensors of CaV1.1 from RYR1-mediated SR Ca2+ release via its ability to interact with β1a. Our findings reveal Rem-expressing adult muscle as an experimental system that may prove useful in the definition of the precise role of the β1a subunit in skeletal-type EC coupling.

scholarly journals Stac3 has a direct role in skeletal muscle-type excitation–contraction coupling that is disrupted by a myopathy-causing mutation

2016 ◽  
Vol 113 (39) ◽  
pp. 10986-10991 ◽  
Author(s):  
Alexander Polster ◽  
Benjamin R. Nelson ◽  
Eric N. Olson ◽  
Kurt G. Beam
Keyword(s):  
Skeletal Muscle ◽  
Native American ◽  
Plasma Membrane ◽  
Membrane Trafficking ◽  
Support Surface ◽  
Membrane Expression ◽  
Direct Role ◽  
Supported Membrane ◽  
Ec Coupling ◽  
Conformational Coupling

In skeletal muscle, conformational coupling between CaV1.1 in the plasma membrane and type 1 ryanodine receptor (RyR1) in the sarcoplasmic reticulum (SR) is thought to underlie both excitation–contraction (EC) coupling Ca2+ release from the SR and retrograde coupling by which RyR1 increases the magnitude of the Ca2+ current via CaV1.1. Recent work has shown that EC coupling fails in muscle from mice and fish null for the protein Stac3 (SH3 and cysteine-rich domain 3) but did not establish the functional role of Stac3 in the CaV1.1–RyR1 interaction. We investigated this using both tsA201 cells and Stac3 KO myotubes. While confirming in tsA201 cells that Stac3 could support surface expression of CaV1.1 (coexpressed with its auxiliary β1a and α2-δ1 subunits) and the generation of large Ca2+ currents, we found that without Stac3 the auxiliary γ1 subunit also supported membrane expression of CaV1.1/β1a/α2-δ1, but that this combination generated only tiny Ca2+ currents. In Stac3 KO myotubes, there was reduced, but still substantial CaV1.1 in the plasma membrane. However, the CaV1.1 remaining in Stac3 KO myotubes did not generate appreciable Ca2+ currents or EC coupling Ca2+ release. Expression of WT Stac3 in Stac3 KO myotubes fully restored Ca2+ currents and EC coupling Ca2+ release, whereas expression of Stac3W280S (containing the Native American myopathy mutation) partially restored Ca2+ currents but only marginally restored EC coupling. We conclude that membrane trafficking of CaV1.1 is facilitated by, but does not require, Stac3, and that Stac3 is directly involved in conformational coupling between CaV1.1 and RyR1.

STIM1 negatively regulates Ca2+ release from the sarcoplasmic reticulum in skeletal myotubes

2013 ◽  
Vol 453 (2) ◽  
pp. 187-200 ◽  
Author(s):  
Keon Jin Lee ◽  
Jin Seok Woo ◽  
Ji-Hye Hwang ◽  
Changdo Hyun ◽  
Chung-Hyun Cho ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Direct Interaction ◽  
Dominant Negative ◽  
Wild Type ◽  
Triple Mutant ◽  
Direct Role ◽  
Independent Manner ◽  
C Terminus ◽  
The Times

STIM1 (stromal interaction molecule 1) mediates SOCE (store-operated Ca2+ entry) in skeletal muscle. However, the direct role(s) of STIM1 in skeletal muscle, such as Ca2+ release from the SR (sarcoplasmic reticulum) for muscle contraction, have not been identified. The times required for the maximal expression of endogenous STIM1 or Orai1, or for the appearance of puncta during the differentiation of mouse primary skeletal myoblasts to myotubes, were all different, and the formation of puncta was detected with no stimulus during differentiation, suggesting that, in skeletal muscle, the formation of puncta is a part of the differentiation. Wild-type STIM1 and two STIM1 mutants (Triple mutant, missing Ca2+-sensing residues but possessing the intact C-terminus; and E136X, missing the C-terminus) were overexpressed in the myotubes. The wild-type STIM1 increased SOCE, whereas neither mutant had an effect on SOCE. It was interesting that increases in the formation of puncta were observed in the Triple mutant as well as in wild-type STIM1, suggesting that SOCE-irrelevant puncta could exist in skeletal muscle. On the other hand, overexpression of wild-type or Triple mutant, but not E136X, attenuated Ca2+ releases from the SR in response to KCl [evoking ECC (excitation–contraction coupling) via activating DHPR (dihydropyridine receptor)] in a dominant-negative manner. The attenuation was removed by STIM1 knockdown, and STIM1 was co-immunoprecipitated with DHRP in a Ca2+-independent manner. These results suggest that STIM1 negatively regulates Ca2+ release from the SR through the direct interaction of the STIM1 C-terminus with DHPR, and that STIM1 is involved in both ECC and SOCE in skeletal muscle.

Functional analysis of the R1086H malignant hyperthermia mutation in the DHPR reveals an unexpected influence of the III-IV loop on skeletal muscle EC coupling

2004 ◽  
Vol 287 (4) ◽  
pp. C1094-C1102 ◽  
Author(s):  
Regina G. Weiss ◽  
Kristen M. S. O’Connell ◽  
Bernhard E. Flucher ◽  
Paul D. Allen ◽  
Manfred Grabner ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Malignant Hyperthermia ◽  
Voltage Dependence ◽  
Charge Movement ◽  
Caffeine Concentration ◽  
Maximal Conductance ◽  
Release Channel ◽  
Ec Coupling ◽  
Negative Allosteric Modulator ◽  
Similar Decrease

Malignant hyperthermia (MH) is an inherited pharmacogenetic disorder caused by mutations in the skeletal muscle ryanodine receptor (RyR1) and the dihydropyridine receptor (DHPR) α1S-subunit. We characterized the effects of an MH mutation in the DHPR cytoplasmic III-IV loop of α1S (R1086H) on DHPR-RyR1 coupling after reconstitution in dysgenic (α1S null) myotubes. Compared with wild-type α1S, caffeine-activated Ca2+ release occurred at approximately fivefold lower concentrations in nonexpressing and R1086H-expressing myotubes. Although maximal voltage-gated Ca2+ release was similar in α1S- and R1086H-expressing myotubes, the voltage dependence of Ca2+ release was shifted ∼5 mV to more negative potentials in R1086H-expressing myotubes. Our results demonstrate that α1S functions as a negative allosteric modulator of release channel activation by caffeine/voltage and that the R1086H MH mutation in the intracellular III-IV linker disrupts this negative regulatory influence. Moreover, a low caffeine concentration (2 mM) caused a similar shift in voltage dependence of Ca2+ release in α1S- and R1086H-expressing myotubes. Compared with α1S-expressing myotubes, maximal L channel conductance ( Gmax) was reduced in R1086H-expressing myotubes (α1S 130 ± 10.2, R1086H 88 ± 6.8 nS/nF; P < 0.05). The decrease in Gmax did not result from a change in retrograde coupling with RyR1 as maximal conductance-charge movement ratio ( Gmax/Qmax) was similar in α1S- and R1086H-expressing myotubes and a similar decrease in Gmax was observed for an analogous mutation engineered into the cardiac L channel (R1217H). In addition, both R1086H and R1217H DHPRs targeted normally and colocalized with RyR1 in sarcoplasmic reticulum (SR)-sarcolemmal junctions. These results indicate that the R1086H MH mutation in α1S enhances RyR1 sensitivity to activation by both endogenous (voltage sensor) and exogenous (caffeine) activators.

scholarly journals Interfering with calcium release suppresses I gamma, the "hump" component of intramembranous charge movement in skeletal muscle.

1991 ◽  
Vol 97 (5) ◽  
pp. 845-884 ◽  
Author(s):  
L Csernoch ◽  
G Pizarro ◽  
I Uribe ◽  
M Rodríguez ◽  
E Ríos
Keyword(s):  
Skeletal Muscle ◽  
Time Course ◽  
Calcium Release ◽  
Time Derivative ◽  
Ruthenium Red ◽  
Total Charge ◽  
Charge Movement ◽  
Release Channel ◽  
Release Flux ◽  
Highly Correlated

Four manifestations of excitation-contraction (E-C) coupling were derived from measurements in cut skeletal muscle fibers of the frog, voltage clamped in a Vaseline-gap chamber: intramembranous charge movement currents, myoplasmic [Ca2+] transients, flux of calcium release from the sarcoplasmic reticulum (SR), and the intrinsic optical transparency change that accompanies calcium release. In attempts to suppress Ca release by direct effects on the SR, three interventions were applied: (a) a conditioning pulse that causes calcium release and inhibits release in subsequent pulses by Ca-dependent inactivation; (b) a series of brief, large pulses, separated by long intervals (greater than 700 ms), which deplete Ca2+ in the SR; and (c) intracellular application of the release channel blocker ruthenium red. All these reduced calcium release flux. None was expected to affect directly the voltage sensor of the T-tubule; however, all of them reduced or eliminated a component of charge movement current with the following characteristics: (a) delayed onset, peaking 10-20 ms into the pulse; (b) current reversal during the pulse, with an inward phase after the outward peak; and (c) OFF transient of smaller magnitude than the ON, of variable polarity, and sometimes biphasic. When the total charge movement current had a visible hump, the positive phase of the current eliminated by the interventions agreed with the hump in timing and size. The component of charge movement current blocked by the interventions was greater and had a greater inward phase in slack fibers with high [EGTA] inside than in stretched fibers with no EGTA. Its amplitude at -40 mV was on average 0.26 A/F (SEM 0.03) in slack fibers. The waveform of release flux determined from the Ca transients measured simultaneously with the membrane currents had, as described previously (Melzer, W., E. Ríos, and M. F. Schneider. 1984. Biophysical Journal. 45:637-641), an early peak followed by a descent to a steady level during the pulse. The time at which this peak occurred was highly correlated with the time to peak of the current suppressed, occurring on average 6.9 ms later (SEM 0.73 ms). The current suppressed by the above interventions in all cases had a time course similar to the time derivative of the release flux; specifically, the peak of the time derivative of release flux preceded the peak of the current suppressed by 0.7 ms (SEM 0.6 ms). The magnitude of the current blocked was highly correlated with the inhibitory effect of the interventions on Ca2+ release flux.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals The voltage sensor of excitation-contraction coupling in skeletal muscle. Ion dependence and selectivity.

1989 ◽  
Vol 94 (3) ◽  
pp. 405-428 ◽  
Author(s):  
G Pizarro ◽  
R Fitts ◽  
I Uribe ◽  
E Ríos
Keyword(s):  
Skeletal Muscle ◽  
Voltage Sensor ◽  
Charge Movement ◽  
Ca Channel ◽  
Extracellular Medium ◽  
Metal Free ◽  
Metal Ligands ◽  
Ec Coupling ◽  
Intramembrane Charge Movement ◽  
Ion Species

Manifestations of excitation-contraction (EC) coupling of skeletal muscle were studied in the presence of metal ions of the alkaline and alkaline-earth groups in the extracellular medium. Single cut fibers of frog skeletal muscle were voltage clamped in a double Vaseline gap apparatus, and intramembrane charge movement and myoplasmic Ca2+ transients were simultaneously measured. In metal-free extracellular media both charge movement of the charge 1 type and Ca transients were suppressed. Under metal-free conditions the nonlinear charge distribution was the same in depolarized (holding potential of 0 mV) and normally polarized fibers (holding potentials between -80 and -90 mV). The manifestations of EC coupling recovered when ions of groups Ia and IIa of the periodic table were included in the extracellular solution; the extent of recovery depended on the ion species. These results are consistent with the idea that the voltage sensor of EC coupling has a binding site for metal cations--the "priming" site--that is essential for function. A state model of the voltage sensor in which metal ligands bind preferentially to the priming site when the sensor is in noninactivated states accounts for the results. This theory was used to derive the relative affinities of the various ions for the priming site from the magnitude of the EC coupling response. The selectivity sequence thus constructed is: Ca greater than Sr greater than Mg greater than Ba for group IIa cations and Li greater than Na greater than K greater than Rb greater than Cs for group Ia. Ca2+, the most effective of all ions tested, was 1,500-fold more effective than Na+. This selectivity sequence is qualitatively and quantitatively similar to that of the intrapore binding sites of the L-type cardiac Ca channel. This provides further evidence of molecular similarity between the voltage sensor and Ca channels.

scholarly journals Cross-linking analysis of the ryanodine receptor and α1-dihydropyridine receptor in rabbit skeletal muscle triads

1997 ◽  
Vol 324 (2) ◽  
pp. 689-696 ◽  
Author(s):  
Brendan E. MURRAY ◽  
Kay OHLENDIECK
Keyword(s):  
Skeletal Muscle ◽  
Ryanodine Receptor ◽  
Dihydropyridine Receptor ◽  
Cross Linking ◽  
Binding Studies ◽  
Electron Microscopic ◽  
Release Channel ◽  
Adult Skeletal Muscle ◽  
Junctional Protein ◽  
Physical Interactions

In mature skeletal muscle, excitation–contraction (EC) coupling is thought to be mediated by direct physical interactions between the transverse tubular, voltage-sensing dihydropyridine receptor (DHPR) and the ryanodine receptor (RyR) Ca2+ release channel of the sarcoplasmic reticulum (SR). Although previous attempts at demonstrating interactions between purified RyR and α1-DHPR have failed, the cross-linking analysis shown here indicates low-level complex formation between the SR RyR and the junctional α1-DHPR. After cross-linking of membranes highly enriched in triads with dithiobis-succinimidyl propionate, distinct complexes of more than 3000 kDa were detected. This agrees with numerous physiological and electron-microscopic findings, as well as co-immunoprecipitation experiments with triad receptors and receptor domain-binding studies. However, a distinct overlap of immunoreactivity between receptors was not observed in crude microsomal preparations, indicating that the triad complex is probably of low abundance. Disulphide-bonded, high-molecular-mass clusters of triadin, the junctional protein proposed to mediate interactions in triads, were confirmed to be linked to the RyR. Calsequestrin and the SR Ca2+-ATPase were not found in cross-linked complexes of the RyR and α1-DHPR. Thus, whereas recent studies indicate that the two receptors exhibit temporal differences in their developmental inductions and that receptor interactions are not essential for the formation and maintenance of triads, this study supports the signal transduction hypothesis of direct physical interactions between triad receptors in adult skeletal muscle.

scholarly journals Molecular architecture of membranes involved in excitation-contraction coupling of cardiac muscle.

1995 ◽  
Vol 129 (3) ◽  
pp. 659-671 ◽  
Author(s):  
X H Sun ◽  
F Protasi ◽  
M Takahashi ◽  
H Takeshima ◽  
D G Ferguson ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Cardiac Muscle ◽  
Calcium Release ◽  
Surface Membrane ◽  
Freeze Fracture ◽  
Muscle Membrane ◽  
Charge Movement ◽  
Molecular Architecture ◽  
Direct Role ◽  
Large Particles

Peripheral couplings are junctions between the sarcoplasmic reticulum (SR) and the surface membrane (SM). Feet occupy the SR/SM junctional gap and are identified as the SR calcium release channels, or ryanodine receptors (RyRs). In cardiac muscle, the activation of RyRs during excitation-contraction (e-c) coupling is initiated by surface membrane depolarization, followed by the opening of surface membrane calcium channels, the dihydropyridine receptors (DHPRs). We have studied the disposition of DHPRs and RyRs, and the structure of peripheral couplings in chick myocardium, a muscle that has no transverse tubules. Immunolabeling shows colocalization of RyRs and DHPRs in clusters at the fiber's periphery. The positions of DHPR and RyR clusters change coincidentally during development. Freeze-fracture of the surface membrane reveals the presence of domains (junctional domains) occupied by clusters of large particles. Junctional domains in the surface membrane and arrays of feet in the junctional gap have similar sizes and corresponding positions during development, suggesting that both are components of peripheral couplings. As opposed to skeletal muscle, membrane particles in junctional domains of cardiac muscle do not form tetrads. Thus, despite their proximity to the feet, they do not appear to be specifically associated with them. Two observations establish the identify of the structurally identified feet arrays/junctional domain complexes with the immunocytochemically defined RyRs/DHPRs coclusters: the concomitant changes during development and the identification of feet as the cytoplasmic domains of RyRs. We suggest that the large particles in junctional domains of the surface membrane represent DHPRs. These observations have two important functional consequences. First, the apposition of DHPRs and RyRs indicates that most of the inward calcium current flows into the restricted space where feet are located. Secondly, contrary to skeletal muscle, presumptive DHPRs do not show a specific association with the feet, which is consistent with a less direct role of charge movement in cardiac than in skeletal e-c coupling.

scholarly journals PKA phosphorylation activates the calcium release channel (ryanodine receptor) in skeletal muscle

2003 ◽  
Vol 160 (6) ◽  
pp. 919-928 ◽  
Author(s):  
Steven Reiken ◽  
Alain Lacampagne ◽  
Hua Zhou ◽  
Aftab Kherani ◽  
Stephan E. Lehnart ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Ryanodine Receptor ◽  
Calcium Release ◽  
Calcium Release Channel ◽  
Skeletal Muscle Function ◽  
Release Channel ◽  
Fk506 Binding Protein ◽  
Ec Coupling ◽  
Increased Activity

The type 1 ryanodine receptor (RyR1) on the sarcoplasmic reticulum (SR) is the major calcium (Ca2+) release channel required for skeletal muscle excitation–contraction (EC) coupling. RyR1 function is modulated by proteins that bind to its large cytoplasmic scaffold domain, including the FK506 binding protein (FKBP12) and PKA. PKA is activated during sympathetic nervous system (SNS) stimulation. We show that PKA phosphorylation of RyR1 at Ser2843 activates the channel by releasing FKBP12. When FKB12 is bound to RyR1, it inhibits the channel by stabilizing its closed state. RyR1 in skeletal muscle from animals with heart failure (HF), a chronic hyperadrenergic state, were PKA hyperphosphorylated, depleted of FKBP12, and exhibited increased activity, suggesting that the channels are “leaky.” RyR1 PKA hyperphosphorylation correlated with impaired SR Ca2+ release and early fatigue in HF skeletal muscle. These findings identify a novel mechanism that regulates RyR1 function via PKA phosphorylation in response to SNS stimulation. PKA hyperphosphorylation of RyR1 may contribute to impaired skeletal muscle function in HF, suggesting that a generalized EC coupling myopathy may play a role in HF.

Malignant Hyperthermia: An Inherited Disorder of Skeletal Muscle Ca2+ Regulation

2001 ◽  
Vol 21 (2) ◽  
pp. 155-168 ◽  
Author(s):  
Charles F. Louis ◽  
Edward M. Balog ◽  
Bradley R. Fruen
Keyword(s):  
Skeletal Muscle ◽  
Malignant Hyperthermia ◽  
Muscle Relaxants ◽  
Release Channel ◽  
Major Locus ◽  
Channel Proteins ◽  
Ec Coupling ◽  
Physiologic Mechanism ◽  
Intact Muscle ◽  
Life Threatening

Malignant hyperthermia (MH) is a pharmacogenetic disorder of skeletal muscle characterized by muscle contracture and life-threatening hypermetabolic crisis following exposure to halogenated anesthetics and depolarizing muscle relaxants during surgery. Susceptibility to MH results from mutations in Ca2+ channel proteins that mediate excitation–contraction (EC) coupling, with the ryanodine receptor Ca2+ release channel (RyR1) representing the major locus. Here we review recent studies characterizing the effects of MH mutations on the sensitivity of the RyR1 to drugs and endogenous channel effectors including Ca2+ and calmodulin. In addition, we present a working model that incorporates these effects of MH mutations on the isolated RyR1 with their effects on the physiologic mechanism that activates Ca2+ release during EC coupling in intact muscle.

scholarly journals An allosteric model of the molecular interactions of excitation-contraction coupling in skeletal muscle.

1993 ◽  
Vol 102 (3) ◽  
pp. 449-481 ◽  
Author(s):  
E Ríos ◽  
M Karhanek ◽  
J Ma ◽  
A González
Keyword(s):  
Skeletal Muscle ◽  
Calcium Release ◽  
Kinetic Equations ◽  
Voltage Sensor ◽  
Charge Movement ◽  
Calcium Release Channel ◽  
Open Probability ◽  
Release Channel ◽  
Voltage Sensors ◽  
Function Relationship

A contact interaction is proposed to exist between the voltage sensor of the transverse tubular membrane of skeletal muscle and the calcium release channel of the sarcoplasmic reticulum. This interaction is given a quantitative formulation inspired in the Monod, Wyman, and Changeux model of allosteric transitions in hemoglobin (Monod, J., J. Wyman, and J.-P. Changeux. 1965. Journal of Molecular Biology. 12:88-118), and analogous to one proposed by Marks and Jones for voltage-dependent Ca channels (Marks, T. N., and S. W. Jones. 1992. Journal of General Physiology. 99:367-390). The allosteric protein is the calcium release channel, a homotetramer, with two accessible states, closed and open. The kinetics and equilibrium of this transition are modulated by voltage sensors (dihydropyridine receptors) pictured as four units per release channel, each undergoing independent voltage-driven transitions between two states (resting and activating). For each voltage sensor that moves to the activating state, the tendency of the channel to open increases by an equal (large) factor. The equilibrium and kinetic equations of the model are solved and shown to reproduce well a number of experimentally measured relationships including: charge movement (Q) vs. voltage, open probability of the release channel (Po) vs. voltage, the transfer function relationship Po vs. Q, and the kinetics of charge movement, release activation, and deactivation. The main consequence of the assumption of allosteric coupling is that primary effects on the release channel are transmitted backward to the voltage sensor and give secondary effects. Thus, the model reproduces well the effects of perchlorate, described in the two previous articles, under the assumption that the primary effect is to increase the intrinsic tendency of the release channel to open, with no direct effects on the voltage sensor. This modification of the open-closed equilibrium of the release channel causes a shift in the equilibrium dependency of charge movement with voltage. The paradoxical slowing of charge movement by perchlorate also results from reciprocal effects of the channel on the allosterically coupled voltage sensors. The observations of the previous articles plus the simulations in this article constitute functional evidence of allosteric transmission.

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