scholarly journals Role of Amino-terminal Half of the S4-S5 Linker in Type 1 Ryanodine Receptor (RyR1) Channel Gating

2011 ◽  
Vol 286 (41) ◽  
pp. 35571-35577 ◽  
Author(s):  
Takashi Murayama ◽  
Nagomi Kurebayashi ◽  
Toshiharu Oba ◽  
Hideto Oyamada ◽  
Katsuji Oguchi ◽  
...  
Keyword(s):  
Ryanodine Receptor ◽  
Single Channel ◽  
Signal Transmission ◽  
Channel Gating ◽  
Helical Structure ◽  
Release Channel ◽  
Amino Terminal ◽  
Molecular Events

The type 1 ryanodine receptor (RyR1) is a Ca2+ release channel found in the sarcoplasmic reticulum of skeletal muscle and plays a pivotal role in excitation-contraction coupling. The RyR1 channel is activated by a conformational change of the dihydropyridine receptor upon depolarization of the transverse tubule, or by Ca2+ itself, i.e. Ca2+-induced Ca2+ release (CICR). The molecular events transmitting such signals to the ion gate of the channel are unknown. The S4-S5 linker, a cytosolic loop connecting the S4 and S5 transmembrane segments in six-transmembrane type channels, forms an α-helical structure and mediates signal transmission in a wide variety of channels. To address the role of the S4-S5 linker in RyR1 channel gating, we performed alanine substitution scan of N-terminal half of the putative S4-S5 linker (Thr4825–Ser4829) that exhibits high helix probability. The mutant RyR1 was expressed in HEK cells, and CICR activity was investigated by caffeine-induced Ca2+ release, single-channel current recordings, and [3H]ryanodine binding. Four mutants (T4825A, I4826A, S4828A, and S4829A) had reduced CICR activity without changing Ca2+ sensitivity, whereas the L4827A mutant formed a constitutive active channel. T4825I, a disease-associated mutation for malignant hyperthermia, exhibited enhanced CICR activity. An α-helical wheel representation of the N-terminal S4-S5 linker provides a rational explanation to the observed activities of the mutants. These results suggest that N-terminal half of the S4-S5 linker may form an α-helical structure and play an important role in RyR1 channel gating.

scholarly journals Nicotinic acid–adenine dinucleotide phosphate activates the skeletal muscle ryanodine receptor

2002 ◽  
Vol 367 (2) ◽  
pp. 423-431 ◽  
Author(s):  
Martin HOHENEGGER ◽  
Josef SUKO ◽  
Regina GSCHEIDLINGER ◽  
Helmut DROBNY ◽  
Andreas ZIDAR
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Nicotinic Acid ◽  
Ryanodine Receptor ◽  
Spatial Information ◽  
Single Channel ◽  
Reversal Potential ◽  
Open Probability ◽  
Release Channel

Calcium is a universal second messenger. The temporal and spatial information that is encoded in Ca2+-transients drives processes as diverse as neurotransmitter secretion, axonal outgrowth, immune responses and muscle contraction. Ca2+-release from intracellular Ca2+ stores can be triggered by diffusible second messengers like InsP3, cyclic ADP-ribose or nicotinic acid—adenine dinucleotide phosphate (NAADP). A target has not yet been identified for the latter messenger. In the present study we show that nanomolar concentrations of NAADP trigger Ca2+-release from skeletal muscle sarcoplasmic reticulum. This was due to a direct action on the Ca2+-release channel/ryanodine receptor type-1, since in single channel recordings, NAADP increased the open probability of the purified channel protein. The effects of NAADP on Ca2+-release and open probability of the ryanodine receptor occurred over a similar concentration range (EC5030nM) and were specific because (i) they were blocked by Ruthenium Red and ryanodine, (ii) the precursor of NAADP, NADP, was ineffective at equimolar concentrations, (iii) NAADP did not affect the conductance and reversal potential of the ryanodine receptor. Finally, we also detected an ADP-ribosyl cyclase activity in the sarcoplasmic reticulum fraction of skeletal muscle. This enzyme was not only capable of synthesizing cyclic GDP-ribose but also NAADP, with an activity of 0.25nmol/mg/min. Thus, we conclude that NAADP is generated in the vicinity of type 1 ryanodine receptor and leads to activation of this ion channel.

scholarly journals Regulation by Ca2+ and Inositol 1,4,5-Trisphosphate (Insp3) of Single Recombinant Type 3 Insp3 Receptor Channels

2001 ◽  
Vol 117 (5) ◽  
pp. 435-446 ◽  
Author(s):  
Don-On Daniel Mak ◽  
Sean McBride ◽  
J. Kevin Foskett
Keyword(s):  
Mammalian Cells ◽  
Single Channel ◽  
Channel Gating ◽  
Hill Coefficient ◽  
Release Channel ◽  
Recombinant Type ◽  
Inositol 1,4,5 Trisphosphate ◽  
Type 3 ◽  
Insp3 Receptor

The inositol 1,4,5-trisphosphate (InsP3) receptor (InsP3R) is an endoplasmic reticulum–localized Ca2+-release channel that controls complex cytoplasmic Ca2+ signaling in many cell types. At least three InsP3Rs encoded by different genes have been identified in mammalian cells, with different primary sequences, subcellular locations, variable ratios of expression, and heteromultimer formation. To examine regulation of channel gating of the type 3 isoform, recombinant rat type 3 InsP3R (r-InsP3R-3) was expressed in Xenopus oocytes, and single-channel recordings were obtained by patch-clamp electrophysiology of the outer nuclear membrane. Gating of the r-InsP3R-3 exhibited a biphasic dependence on cytoplasmic free Ca2+ concentration ([Ca2+]i). In the presence of 0.5 mM cytoplasmic free ATP, r-InsP3R-3 gating was inhibited by high [Ca2+]i with features similar to those of the endogenous Xenopus type 1 InsP3R (X-InsP3R-1). Ca2+ inhibition of channel gating had an inhibitory Hill coefficient of ∼3 and half-maximal inhibiting [Ca2+]i (Kinh) = 39 μM under saturating (10 μM) cytoplasmic InsP3 concentrations ([InsP3]). At [InsP3] < 100 nM, the r-InsP3R-3 became more sensitive to Ca2+ inhibition, with the InsP3 concentration dependence of Kinh described by a half-maximal [InsP3] of 55 nM and a Hill coefficient of ∼4. InsP3 activated the type 3 channel by tuning the efficacy of Ca2+ to inhibit it, by a mechanism similar to that observed for the type 1 isoform. In contrast, the r-InsP3R-3 channel was uniquely distinguished from the X-InsP3R-1 channel by its enhanced Ca2+ sensitivity of activation (half-maximal activating [Ca2+]i of 77 nM instead of 190 nM) and lack of cooperativity between Ca2+ activation sites (activating Hill coefficient of 1 instead of 2). These differences endow the InsP3R-3 with high gain InsP3–induced Ca2+ release and low gain Ca2+–induced Ca2+ release properties complementary to those of InsP3R-1. Thus, distinct Ca2+ signals may be conferred by complementary Ca2+ activation properties of different InsP3R isoforms.

scholarly journals Spontaneous Channel Activity of the Inositol 1,4,5-Trisphosphate (InsP3) Receptor (InsP3R). Application of Allosteric Modeling to Calcium and InsP3 Regulation of InsP3R Single-channel Gating

2003 ◽  
Vol 122 (5) ◽  
pp. 583-603 ◽  
Author(s):  
Don-On Daniel Mak ◽  
Sean M.J. McBride ◽  
J. Kevin Foskett
Keyword(s):  
Binding Sites ◽  
Single Channel ◽  
Channel Gating ◽  
Quantitative Prediction ◽  
Open Probability ◽  
Release Channel ◽  
Gating Mechanism ◽  
Inhibitory Site ◽  
Type 3

The InsP3R Ca2+ release channel has a biphasic dependence on cytoplasmic free Ca2+ concentration ([Ca2+]i). InsP3 activates gating primarily by reducing the sensitivity of the channel to inhibition by high [Ca2+]i. To determine if relieving Ca2+ inhibition is sufficient for channel activation, we examined single-channel activities in low [Ca2+]i in the absence of InsP3, by patch clamping isolated Xenopus oocyte nuclei. For both endogenous Xenopus type 1 and recombinant rat type 3 InsP3R channels, spontaneous InsP3-independent channel activities with low open probability Po (∼0.03) were observed in [Ca2+]i < 5 nM with the same frequency as in the presence of InsP3, whereas no activities were observed in 25 nM Ca2+. These results establish the half-maximal inhibitory [Ca2+]i of the channel to be 1.2–4.0 nM in the absence of InsP3, and demonstrate that the channel can be active when all of its ligand-binding sites (including InsP3) are unoccupied. In the simplest allosteric model that fits all observations in nuclear patch-clamp studies of [Ca2+]i and InsP3 regulation of steady-state channel gating behavior of types 1 and 3 InsP3R isoforms, including spontaneous InsP3-independent channel activities, the tetrameric channel can adopt six different conformations, the equilibria among which are controlled by two inhibitory and one activating Ca2+-binding and one InsP3-binding sites in a manner outlined in the Monod-Wyman-Changeux model. InsP3 binding activates gating by affecting the Ca2+ affinities of the high-affinity inhibitory sites in different conformations, transforming it into an activating site. Ca2+ inhibition of InsP3-liganded channels is mediated by an InsP3-independent low-affinity inhibitory site. The model also suggests that besides the ligand-regulated gating mechanism, the channel has a ligand-independent gating mechanism responsible for maximum channel Po being less than unity. The validity of this model was established by its successful quantitative prediction of channel behavior after it had been exposed to ultra-low bath [Ca2+].

Characterization of the ryanodine receptor/channel of invertebrate muscle

1998 ◽  
Vol 274 (2) ◽  
pp. R494-R502 ◽  
Author(s):  
Kerry E. Quinn ◽  
Loriana Castellani ◽  
Karol Ondrias ◽  
Barbara E. Ehrlich
Keyword(s):  
Ryanodine Receptor ◽  
Single Channel ◽  
Microscopic Analysis ◽  
Ruthenium Red ◽  
Electron Microscopic ◽  
Release Channel ◽  
Single Channel Currents ◽  
Channel Currents ◽  
Bell Shaped Curve

Electron-microscopic analysis was used to show that invertebrate muscle has feetlike structures on the sarcoplasmic reticulum (SR) displaying the typical four-subunit appearance of the calcium (Ca2+) release channel/ryanodine receptor (RyR) observed in vertebrate skeletal muscle (K. E. Loesser, L. Castellani, and C. Franzini-Armstrong. J. Muscle Res. Cell Motil. 13: 161–173, 1992). SR vesicles from invertebrate muscle exhibited specific ryanodine binding and single channel currents that were activated by Ca2+, caffeine, and ATP and inhibited by ruthenium red. The single channel conductance of this invertebrate RyR was lower than that of the vertebrate RyR (49 and 102 pS, respectively). Activation of lobster and scallop SR Ca2+ release channel, in response to cytoplasmic Ca2+ (1 nM–10 mM), reflected a bell-shaped curve, as is found with the mammalian RyR. In contrast to a previous report (J.-H. Seok, L. Xu, N. R. Kramarcy, R. Sealock, and G. Meissner. J. Biol. Chem. 267: 15893–15901, 1992), our results show that regulation of the invertebrate and vertebrate RyRs is quite similar and suggest remarkably similar paths in these diverse organisms.

scholarly journals Evidence for the transport of glutathione through ryanodine receptor channel type 1

2003 ◽  
Vol 376 (3) ◽  
pp. 807-812 ◽  
Author(s):  
Gábor BÁNHEGYI ◽  
Miklós CSALA ◽  
Gábor NAGY ◽  
Vincenzo SORRENTINO ◽  
Rosella FULCERI ◽  
...  
Keyword(s):  
Ryanodine Receptor ◽  
Skeletal Muscles ◽  
Ruthenium Red ◽  
Lanthanum Chloride ◽  
Human Embryonic Kidney ◽  
Hek 293 ◽  
Channel Type ◽  
Receptor Channel

In the present study, we have investigated the role of RyR1 (ryanodine receptor calcium channel type 1) in glutathione (GSH) transport through the sarcoplasmic reticulum (SR) membrane of skeletal muscles. Lanthanum chloride, a prototypic blocker of cation channels, inhibited the influx and efflux of GSH in SR vesicles. Using a rapid-filtration-based assay and lanthanum chloride as a transport blocker, an uptake of radiolabelled GSH into SR vesicles was observed. Pretreatment of SR vesicles with the RyR1 antagonists Ruthenium Red and ryanodine as well as with lanthanum chloride blocked the GSH uptake. An SR-like GSH uptake appeared in microsomes obtained from an HEK-293 (human embryonic kidney 293) cell line after transfection of RyR1. These observations strongly suggest that RyR1 mediates GSH transport through the SR membranes of skeletal muscles.

scholarly journals Modification of ryanodine receptor/Ca2+ release channel with dinitrofluorobenzene

1999 ◽  
Vol 342 (1) ◽  
pp. 239-248 ◽  
Author(s):  
Nurit HADAD ◽  
Wei FENG ◽  
Varda SHOSHAN-BARMATZ
Keyword(s):  
Ryanodine Receptor ◽  
Single Channel ◽  
Ruthenium Red ◽  
Amino Group ◽  
Channel Activity ◽  
Closed Channel ◽  
Release Channel ◽  
Atp Binding Site ◽  
Ph Values ◽  
Stimulation Of

Modification of the ryanodine receptor (RyR)/Ca2+ release channel with 2,4-dinitrofluorobenzene (DNFB) indicated that two classes of amino group interact with the reagent, as can be distinguished on the basis of their reactivity/accessibility and the effects on ryanodine binding and single channel activities. One group interacted very rapidly (t½ < 30 s) at 25 °C with low concentrations of DNFB [C50 (concentration of DNFB required for 50% inhibition or stimulation of ryanodine binding) = 5 μM], and at pH values of 6.2 and higher. This interaction resulted in the marked stimulation of ryanodine binding and the complete inhibition of a single Ca2+ release channel incorporated into planar lipid bilayer. The second group is accessible at higher temperatures (37 °C); at pH values higher than 7.4 it reacted slowly (t½ = 20 min) with high concentrations of DNFB (C50 = 70 μM). This interaction led to the inhibition of ryanodine binding and single channel activity. Modification of RyR with DNFB under the stimulatory conditions resulted in 3.6-fold and 6-fold increases in ryanodine-binding and Ca2+-binding affinities respectively. Modification with DNFB under the inhibitory conditions resulted in a decrease in the total ryanodine-binding sites. The exposure of the RyR single channel to DNFB under both inhibitory and stimulatory conditions led to the complete closure of the channel. However, when modified under the stimulatory conditions, but not under the inhibitory ones, the DNFB-modified closed channel could be re-activated by sub-micromolar concentrations of ryanodine, in the presence of nanomolar concentrations of Ca2+. The DNFB-modified ryanodine-activated RyR channel showed fast transitions between open, closed and several sub-conductance states, and was completely closed by Ruthenium Red. ATP re-activated the DNFB-modified closed channel or, if present during modification, prevented the inhibition of RyR channel activity by DNFB. Neither the stimulation nor the inhibition of ryanodine binding by modification with DNFB was affected by the presence of ATP. By using the photoreactive ATP analogue 3′-O-(4-benzoyl)benzoyl-[α-32P]ATP we found that DNFB modification had no effect on the ATP-binding site of RyR. The results are discussed with regard to the involvement of amino group residues in channel gating, ryanodine association/dissociation and occlusion, and the relationship between the open/closed state of the RyR and its capacity to bind ryanodine.

scholarly journals Divergent Activity Profiles of Type 1 Ryanodine Receptor Channels Carrying Malignant Hyperthermia and Central Core Disease Mutations in the Amino-Terminal Region

PLoS ONE ◽  
2015 ◽  
Vol 10 (6) ◽  
pp. e0130606 ◽  
Author(s):  
Takashi Murayama ◽  
Nagomi Kurebayashi ◽  
Toshiko Yamazawa ◽  
Hideto Oyamada ◽  
Junji Suzuki ◽  
...  
Keyword(s):  
Malignant Hyperthermia ◽  
Ryanodine Receptor ◽  
Terminal Region ◽  
Central Core ◽  
Central Core Disease ◽  
Amino Terminal ◽  
Disease Mutations

scholarly journals Atp-Dependent Adenophostin Activation of Inositol 1,4,5-Trisphosphate Receptor Channel Gating

2001 ◽  
Vol 117 (4) ◽  
pp. 299-314 ◽  
Author(s):  
Don-On Daniel Mak ◽  
Sean McBride ◽  
J. Kevin Foskett
Keyword(s):  
Single Channel ◽  
Free Acid ◽  
Channel Gating ◽  
Xenopus Laevis Oocyte ◽  
Open Probability ◽  
Release Channel ◽  
Gating Kinetics ◽  
Inositol 1,4,5 Trisphosphate ◽  
Kinetics Of ◽  
In Cells

The inositol 1,4,5-trisphosphate (InsP3) receptor (InsP3R) is a ligand-gated intracellular Ca2+ release channel that plays a central role in modulating cytoplasmic free Ca2+ concentration ([Ca2+]i). The fungal metabolite adenophostin A (AdA) is a potent agonist of the InsP3R that is structurally different from InsP3 and elicits distinct calcium signals in cells. We have investigated the effects of AdA and its analogues on single-channel activities of the InsP3R in the outer membrane of isolated Xenopus laevis oocyte nuclei. InsP3R activated by either AdA or InsP3 have identical channel conductance properties. Furthermore, AdA, like InsP3, activates the channel by tuning Ca2+ inhibition of gating. However, gating of the AdA-liganded InsP3R has a critical dependence on cytoplasmic ATP free acid concentration not observed for InsP3-liganded channels. Channel gating activated by AdA is indistinguishable from that elicited by InsP3 in the presence of 0.5 mM ATP, although the functional affinity of the channel is 60-fold higher for AdA. However, in the absence of ATP, gating kinetics of AdA-liganded InsP3R were very different. Channel open time was reduced by 50%, resulting in substantially lower maximum open probability than channels activated by AdA in the presence of ATP, or by InsP3 in the presence or absence of ATP. Also, the higher functional affinity of InsP3R for AdA than for InsP3 is nearly abolished in the absence of ATP. Low affinity AdA analogues furanophostin and ribophostin activated InsP3R channels with gating properties similar to those of AdA. These results provide novel insights for interpretations of observed effects of AdA on calcium signaling, including the mechanisms that determine the durations of elementary Ca2+ release events in cells. Comparisons of single-channel gating kinetics of the InsP3R activated by InsP3, AdA, and its analogues also identify molecular elements in InsP3R ligands that contribute to binding and activation of channel gating.

Ryanodine receptor function in newborn rat heart

2005 ◽  
Vol 288 (5) ◽  
pp. H2527-H2540 ◽  
Author(s):  
Claudia G. Pérez ◽  
Julio A. Copello ◽  
Yanxia Li ◽  
Kimberly L. Karko ◽  
Leticia Gómez ◽  
...  
Keyword(s):  
Lipid Bilayers ◽  
Ryanodine Receptor ◽  
Functional Properties ◽  
Rat Heart ◽  
Single Channel ◽  
Binding Assays ◽  
Adult Rats ◽  
Intracellular Ca ◽  
Cardiac Excitation

The role of ryanodine receptor (RyR) in cardiac excitation-contraction (E-C) coupling in newborns (NB) is not completely understood. To determine whether RyR functional properties change during development, we evaluated cellular distribution and functionality of sarcoplasmic reticulum (SR) in NB rats. Sarcomeric arrangement of immunostained SR Ca2+-ATPase (SERCA2a) and the presence of sizeable caffeine-induced Ca2+ transients demonstrated that functional SR exists in NB. E-C coupling properties were then defined in NB and compared with those in adult rats (AD). Ca2+ transients in NB reflected predominantly sarcolemmal Ca2+ entry, whereas the RyR-mediated component was ∼13%. Finally, the RyR density and functional properties at the single-channel level in NB were compared with those in AD. Ligand binding assays revealed that in NB, RyR density can be up to 36% of that found in AD, suggesting that some RyRs do not contribute to the Ca2+ transient. To test the hypothesis that RyR functional properties change during development, we incorporated single RyRs into lipid bilayers. Our results show that permeation and gating kinetics of NB RyRs are identical to those of AD. Also, endogenous ligands had similar effects on NB and AD RyRs: sigmoidal Ca2+ dependence, stronger Mg2+-induced inhibition at low cytoplasmic Ca2+ concentrations, comparable ATP-activating potency, and caffeine sensitivity. These observations indicate that NB rat heart contains fully functional RyRs and that the smaller contribution of RyR-mediated Ca2+ release to the intracellular Ca2+ transient in NB is not due to different single RyR channel properties or to the absence of functional intracellular Ca2+ stores.

Sign in / Sign up

Export Citation Format

Share Document