Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P21817 (RyR1)
 1,154 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Using whole cell patch clamp recordings on unfertilized eggs of the ascidian Ciona intestinalis, we are able to detect ryanodine receptors within the oocytes. Our approach is based on measurements of the voltage-activated inward calcium currents. Two types of Ca2+ currents have been described on the oocyte membrane of Ciona: a low threshold slowly activating current, and a high threshold faster one. We show here that caffeine induces a decrease in the intensity of the Ca2+ currents, when applied either externally or internally from the mouth of a patch pipette. Caffeine application mimics fertilization which transiently decreases the high threshold Ca2+ current density during density during the first meiotic cycle. Ryanodine (> 1 nM) has an effect similar to caffeine. This partial decrease in Ca2+ current density elicited by caffeine or ryanodine is prevented by intracellular application of the calcium chelator BAPTA, then imputable to calcium release. In summary, the depolarization-induced Ca2+ current intensity allows monitoring of an intracellular calcium store which is sensitive to low concentrations of ryanodine in Ciona oocytes. Further identification of a ryanodine receptor was obtained by immunological staining with antibodies against mammalian skeletal muscle ryanodine receptor. Ryanodine receptors were asymmetrically localized in the cortex of Ciona eggs. We discuss the methodological relevance of our patch-clamp approach, in connection with the possible biological role of such a ryanodine receptor in the early stages of development.
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PMID:A ryanodine-sensitive calcium store in ascidian eggs monitored by whole-cell patch-clamp recordings. 913

In the present study, we compare functional consequences of dissociation and reconstitution of binding proteins FKBP12 and FKBP12.6 with ryanodine receptors from cardiac (RyR2) and skeletal muscle (RyR1). The skeletal muscle RyR1 channel became activated on removal of endogenously bound FKBP12, consistent with previous reports. Both FKBP12 and FKBP12.6 rebind to FKBP-depleted RyR1 and restore its quiescent channel behavior by altering ligand sensitivity, as studied by single-channel recordings in planar lipid bilayers, and macroscopic behavior of the channels (ryanodine binding and net energized Ca2- uptake). By contrast, removal of FKBP12.6 from the cardiac RyR2 did not modulate the function of the channel using the same types of assays as for RyR1. FKBP12 or FKBP12.6 had no effect on channel activity of FKBP12.6-depleted cardiac RyR2, although FKBP12.6 rebinds. Our studies reveal important differences between the two ryanodine receptor isoforms with respect to their functional interaction with FKBP12 and FKBP12.6.
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PMID:Different interactions of cardiac and skeletal muscle ryanodine receptors with FK-506 binding protein isoforms. 917 65

Malignant hyperthermia susceptibility (MHS) is characterized by genetic heterogeneity. However, except for the MHS1 locus, which corresponds to the skeletal muscle ryanodine receptor (RYR1) and for which several mutations have been described, no direct molecular evidence for a mutation in another gene has been reported so far. In this study we show that the CACNL1A3 gene encoding the alpha 1-subunit of the human skeletal muscle dihydropyridine-sensitive L-type voltage-dependent calcium channel (VDCC) represents a new MHS locus and is responsible for the disease in a large French family. Linkage analysis performed with an intragenic polymorphic microsatellite marker of the CACLN1A3 gene generated a two-point LOD score of 4.38 at a recombinant fraction of 0. Sequence analysis of the coding region of the CACLN1A3 gene showed the presence of an Arg-His substitution at residue 1086, resulting from the transition of A for G3333, which segregates perfectly with the MHS phenotype in the family. The mutation is localized in a very different part of the alpha 1-subunit of the human skeletal muscle VDCC, compared with previously reported mutations found in patients with hypokalemic periodic paralysis, and these two diseases might be discussed in terms of allelic diseases. This report is the first direct evidence that the skeletal muscle VDCC is involved in MHS, and it suggests a direct interaction between the skeletal muscle VDCC and the ryanodine receptor in the skeletal muscle sarcoplasmic reticulum.
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PMID:Malignant-hyperthermia susceptibility is associated with a mutation of the alpha 1-subunit of the human dihydropyridine-sensitive L-type voltage-dependent calcium-channel receptor in skeletal muscle. 919 49

In vertebrate skeletal muscles, the type 1 isoform of ryanodine receptor (RyR1) is essential in triggering contraction by releasing Ca2+ from the sarcoplasmic reticulum in response to plasma membrane depolarisation. Recently, the presence of another RyR isoform, RyR3, has been detected in mammalian skeletal muscle cells, raising the question of the eventual relevance of RyR3 for muscle cell physiology. The expression of RyR3 was investigated during differentiation of skeletal muscle cells. Using antibodies able to distinguish the different RyR isoforms and Western blot analysis, the RyR3 protein was detected in the microsomal fractions of differentiated skeletal muscle cells but not of undifferentiated cells. Accordingly, blocking muscle differentiation by the addition of either transforming growth factor-beta or basic fibroblast growth factor prevented the expression of the RyR3 protein. In differentiated skeletal muscle cells, RyR3 was expressed independent of cell fusion and myotube formation. The expression of RyR3 was also investigated during development of the diaphragm muscle. The RyR3 content in the diaphragm muscle increased between the late stage of fetal development and the first postnatal days. However, at variance with RyR1, which reached maximum levels of expression 2-3 weeks after birth, the expression of RyR3 was found to be higher in the neonatal phase of the diaphragm muscle development (2-15 days after birth) than in the same muscle from adult mice. The differential content of RyR3 in adult skeletal muscles was found not to be mediated by neurotrophic factors or electrical activity. These findings indicate that RyR3 is preferentially expressed in differentiated skeletal muscle cells. In addition, during skeletal muscle development, its expression is regulated differently from that of RyR1.
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PMID:Expression of the ryanodine receptor type 3 calcium release channel during development and differentiation of mammalian skeletal muscle cells. 924 41

Previously, we reported the presence of a heterogeneous population of mRNAs in the squid giant axon. The construction of a cDNA library to this mRNA population has facilitated the identification of several of the constituent mRNAs which encode several cytoskeletal and motor proteins as well as enolase, a glycolytic enzyme. In this communication, we report the isolation of a novel mRNA species (pA6) from the axonal cDNA library. The pA6 mRNA is relatively small (550 nucleotides in length) and is expressed in both nervous tissue and skeletal muscle. The axonal localization of pA6 mRNA was unequivocally established by in situ hybridization histochemistry. The results of quantitative RT-PCR analysis indicate that there are 1.8 x 10(6) molecules of pA6 mRNA (approximately 0.45 pg) in the analyzed segment of the giant axon and suggest that the level of pA6 mRNA in the axonal domain of the giant fiber system might be equal to or greater than the level present in the parental cell soma. Sequence analysis of pA6 suggests that the mRNA encodes an integral membrane protein comprising 84 amino acids. The putative protein contains a single transmembrane domain located in the middle of the molecule and a phosphate-binding loop situated near the N terminus. The C-terminal region of the protein contains two potential phosphorylation sites. These four structural motifs manifest striking similarity to domains present in the ryanodine receptor, raising the possibility that pA6 represents a cephalopod intracellular calcium release channel protein.
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PMID:Molecular cloning and characterization of a novel mRNA present in the squid giant axon. 927 37

We investigated type 3 isoform (RyR3) of ryanodine receptor in rabbit skeletal muscles using an antibody specific for RyR3. By Western blot analysis and by immunoprecipitation, a single polypeptide for RyR3 was detected in sarcoplasmic reticulum vesicles from rabbit diaphragm but not in those from back muscle. The molecular mass was slightly smaller than that of RyR1, the major isoform in skeletal muscles. Each of RyR1 and RyR3 formed a homotetramer in rabbit diaphragm. RyR3 had a single class of [3H]ryanodine binding sites of high affinity (KD = 1.6 nM). From the Bmax of the binding, the content of RyR3 was estimated to be only 0.6% of RyR1 in rabbit diaphragm. -3H-Ryanodine binding to RyR3 was biphasically dependent on Ca2+, as is true of RyR1, and was stimulated further by adenine nucleotide, caffeine, or high salt concentration. Procaine and ruthenium red inhibited the binding. RyR3 was more resistant to Mg2+ inhibition than RyR1. Interestingly, RyR3 showed about a 7-fold lower Ca2+ sensitivity for activation than RyR1. Comparison with the counterparts in bullfrog skeletal muscles indicates that the Ca2+ sensitivities of RyR3 homologs are similar to each other, whereas those of RyR1 homologs are species-specific.
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PMID:Characterization of type 3 ryanodine receptor (RyR3) of sarcoplasmic reticulum from rabbit skeletal muscles. 929 56

In the present report we studied the interaction between the skeletal muscle ryanodine receptor and the ubiquitous S100A1 Ca2+ binding protein. S100A1 did not affect equilibrium [3H]ryanodine binding to purified rabbit skeletal muscle terminal cisternae at 100 microM free [Ca2+]. At nanomolar free [Ca2+], however, S100A1 activated by 40 +/- 6.7% (mean +/- SE, n = 5) the [3H]ryanodine binding activity; the half-maximal concentration for stimulation of [3H]ryanodine binding was approximately 70 nM, a value well below the estimated S100A1 concentration in skeletal muscle fibers. Scatchard analysis of [3H]ryanodine binding performed in the presence of 100 microM EGTA indicates that S100A1 increases the apparent affinity of the receptor for ryanodine (Kd = 191 vs 383 nM in the presence and in the absence of 100 nM S100A1, respectively). The effect of S100A1 was also tested on the single-channel gating properties of the purified ryanodine receptor after reconstitution into a lipid planar bilayer. Currents carried by purified ryanodine receptor channels were modulated by both cis Ca2+ and ruthenium red. In the presence of nanomolar [Ca2+], S100A1 activated the channel by increasing (6.0 +/- 2.8)-fold (mean +/- SE, n = 3) the normalized open probability. The interaction between S100A1 and the purified RYR was verified using the optical biosensor BIAcore: we show that the two proteins interact directly both at millimolar and at nanomolar calcium concentrations. We next mapped the regions of the skeletal muscle RYR involved in the interaction with S100A1 by performing ligand overlays on a panel RYR of fusion proteins in the presence of 100 nM S100A1. Our results indicate that the skeletal muscle RYR contains three potential S100A1 binding domains. Binding of S100A1 to the RYR fusion proteins occurred at both nanomolar and millimolar free [Ca2+]. S100A1 binding domain 1 binds the ligand in the presence of 1 mM free [Ca2+] or 1 mM EGTA. Maximal binding to S100A1#2 was achieved in the presence of 1 mM free [Ca2+]. The S100A1#3 domain, which overlaps with calcium-dependent calmodulin binding domain 3 (CaM 3), exhibits weak and strong S100A1 binding activity in the presence of either millimolar or nanomolar Ca2+, respectively. The interaction between S100A1 and the purified RYR complex was also investigated by affinity chromatography: in the presence of nanomolar Ca2+, we observed binding of native RYR complex to S100A1-conjugated Sepharose. This interaction could be inhibited by the presence of RYR polypeptides encompassing S100A1 binding sites S100A1#1, S100A1#2, and S100A1#3.
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PMID:Interaction of S100A1 with the Ca2+ release channel (ryanodine receptor) of skeletal muscle. 929 70

To investigate the channel properties of the mammalian type 3 ryanodine receptor (RyR3), we have cloned the RyR3 cDNA from rabbit uterus by reverse transcriptase-polymerase chain reaction and expressed the cDNA in HEK293 cells. Immunoblotting studies showed that the cloned RyR3 was indistinguishable from the native mammalian RyR3 in molecular size and immunoreactivity. Ca2+ release measurements using the fluorescence Ca2+ indicator fluo 3 revealed that the cloned RyR3 functioned as a caffeine- and ryanodine-sensitive Ca2+ release channel in HEK293 cells. Functional properties of the cloned RyR3 were further characterized by using single channel recordings in lipid bilayers. The cloned RyR3 channel exhibited a K+ conductance of 777 picosiemens in 250 mM KCl and a Ca2+ conductance of 137 picosiemens in 250 mM CaCl2 and displayed a pCa2+/pK+ ratio of 6.3 and an open time constant of about 1.16 ms. The response of the cloned RyR3 to cytoplasmic Ca2+ concentrations was biphasic. The channel was activated by Ca2+ at about 100 nM and inactivated at about 10 mM. Ca2+ alone was able to activate the cloned RyR3 fully. Calmodulin activated the cloned RyR3 at low Ca2+ concentrations but inhibited the channel at high Ca2+ concentrations. The cloned RyR3 was activated by ATP, caffeine, and perchlorate, inhibited by Mg2+ and ruthenium red, and modified by ryanodine. Cyclic ADP-ribose did not seem to affect single channel activity of the cloned RyR3. The most prominent differences of the cloned RyR3 from the rabbit skeletal muscle ryanodine receptor were in the gating kinetics, extent of maximal activation by Ca2+, and sensitivity to Ca2+ inactivation. Results of the present study provide initial insights into the single channel properties of the mammalian RyR3.
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PMID:Functional characterization of the recombinant type 3 Ca2+ release channel (ryanodine receptor) expressed in HEK293 cells. 930 76

The skeletal muscle relaxant dantrolene inhibits the release of Ca2+ from the sarcoplasmic reticulum during excitation-contraction coupling and suppresses the uncontrolled Ca2+ release that underlies the skeletal muscle pharmacogenetic disorder malignant hyperthermia; however, the molecular mechanism by which dantrolene selectively affects skeletal muscle Ca2+ regulation remains to be defined. Here we provide evidence of a high-affinity, monophasic inhibition by dantrolene of ryanodine receptor Ca2+ channel function in isolated sarcoplasmic reticulum vesicles prepared from malignant hyperthermia-susceptible and normal pig skeletal muscle. In media simulating resting myoplasm, dantrolene increased the half-time for 45Ca2+ release from both malignant hyperthermia and normal vesicles approximately 3.5-fold and inhibited sarcoplasmic reticulum vesicle [3H]ryanodine binding (Ki approximately 150 nM for both malignant hyperthermia and normal). Inhibition of vesicle [3H]ryanodine binding by dantrolene was associated with a decrease in the extent of activation by both calmodulin and Ca2+. Dantrolene also inhibited [3H]ryanodine binding to purified skeletal muscle ryanodine receptor protein reconstituted into liposomes. In contrast, cardiac sarcoplasmic reticulum vesicle 45Ca2+ release and [3H]ryanodine binding were unaffected by dantrolene. Together, these results demonstrate selective effects of dantrolene on skeletal muscle ryanodine receptors that are consistent with the actions of dantrolene in vivo and suggest a mechanism of action in which dantrolene may act directly at the skeletal muscle ryanodine receptor complex to limit its activation by calmodulin and Ca2+. The potential implications of these results for understanding how dantrolene and malignant hyperthermia mutations may affect the voltage-dependent activation of Ca2+ release in intact skeletal muscle are discussed.
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PMID:Dantrolene inhibition of sarcoplasmic reticulum Ca2+ release by direct and specific action at skeletal muscle ryanodine receptors. 934 Nov 33

A 94 kDa large subunit thiol-protease, as identified by anti-calpain antibodies, has been isolated from skeletal muscle junctional sarcoplasmic reticulum (SR). This protease cleaves specifically the skeletal muscle ryanodine receptor (RyR)/Ca2+ release channel at one site resulting in the 375 kDa and 150 kDa fragments. The 94 kDa thiol-protease degrades neither other SR proteins nor the ryanodine receptor of cardiac nor brain membranes. The partially purified 94 kDa protease, like the SR associated protease, had an optimal pH of about 7.0, was absolutely dependent on the presence of thiol reducing reagents, and was completely inhibited by HgCl2, leupeptin and the specific calpain I inhibitor. However, while the SR membrane-associated protease requires Ca2+ at a submicromolar concentration, the isolated thiol-protease has lost the Ca2+ requirement. The 94 kDa thiol-protease had no effect on ryanodine binding but modified the channel activity of RyR reconstituted into planar lipid bilayer: in a time-dependent manner, the channel activity decreases and within several minutes the channel is converted into a subconducting state. The protease-modified channel activity is still Ca2+-dependent and ryanodine sensitive. This 94 kDa thiol-protease cross react with anti-calpain antibodies thus, may represent the novel large subunit of the skeletal muscle specific calpain p94.
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PMID:Identification, characterization and partial purification of a thiol-protease which cleaves specifically the skeletal muscle ryanodine receptor/Ca2+ release channel. 943 Jun 19


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