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

Contraction of skeletal muscle is triggered by the release of Ca2+ from the sarcoplasmic reticulum (SR) after depolarization of transverse tubules. The ryanodine receptor exists as a 'foot' protein in the junctional gap between the sarcoplasmic reticulum and the transverse tubule in skeletal muscle, and is proposed to function as a calcium-release channel during excitation-contraction (E-C) coupling. Previous complementary DNA-cloning studies have defined three distinct subtypes of the ryanodine receptor in mammalian tissues, namely skeletal muscle, cardiac and brain types. We report here mice with a targeted mutation in the skeletal muscle ryanodine receptor gene. Mice homozygous for the mutation die perinatally with gross abnormalities of the skeletal muscle. The contractile response to electrical stimulation under physiological conditions is totally abolished in the mutant muscle, although ryanodine receptors other than the skeletal-muscle type seem to exist because the response to caffeine is retained. Our results show that the skeletal muscle ryanodine receptor is essential for both muscular maturation and E-C coupling, and also imply that the function of the skeletal muscle ryanodine receptor during E-C coupling cannot be substituted by other subtypes of the receptor.
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PMID:Excitation-contraction uncoupling and muscular degeneration in mice lacking functional skeletal muscle ryanodine-receptor gene. 751 81

Microsomal sarcoplasmic reticulum (SR) fractions from lobster skeletal muscle were found to bind [3H]-ryanodine. [3H]-ryanodine binding was enhanced by AMP, Ca2+ and caffeine, and significantly diminished by ATP, Ba2+ and Sr2+. Furthermore, dantrolene and ruthenium red, two classical inhibitors of Ca2+ release from the SR, blocked [3H]-ryanodine binding. Similarly, tetracaine, known to block the charge movement associated with excitation-contraction coupling in vertebrate muscle, inhibited the binding of the alkaloid. Our lobster SR preparation exhibited a single high-affinity ryanodine binding site (Kd = 6.6 nM, Bmax = 10 pmol/mg protein). Since SDS-PAGE of the SR proteins revealed a major band c. 565 kDa which comigrated with the putative ryanodine receptor from both rat and chicken skeletal muscle, we concluded that lobster skeletal muscle is equipped with the 565 kDa ryanodine receptor. Finally, incorporation of the SR microsomal fraction from lobster into planar bilayer membranes revealed the presence of a ryanodine-sensitive Ca2+ channel activity (160 pS in symmetrical 200 mM CsCl solutions). We concluded that both the crustacean and vertebrate skeletal muscle ryanodine receptor share the relevant properties such as molecular weight and affinity for ryanodine and inositol 1,4,5 triphosphate. However, there are important differences between the two receptors including differential effects of the alkaloid on the Ca2+ release channel and modulation of the receptor by nucleotides.
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PMID:Properties of the ryanodine receptor present in the sarcoplasmic reticulum from lobster skeletal muscle. 751 63

cDNAs encoding trpE fusion proteins containing fragments of the skeletal muscle Ca2+ release channel (ryanodine receptor) were expressed in bacteria. The fusion proteins, which covered about 90% of the linear sequence of the ryanodine receptor, were used to identify calmodulin- (CaM), Ca(2+)-, and ruthenium red-binding regions in the ryanodine receptor through the use of 125I-CaM, 45Ca2+, and ruthenium red overlay procedures. Six Ca(2+)-dependent CaM-binding domains were detected in the skeletal muscle ryanodine receptor. Strong CaM-binding domains were localized in regions 6, 11, 12, and 13, in subregions 6b, 11b, and 13b, and in short sequences 6b3, 11b1, and 13b2, lying between amino acid residues 2063 and 2091, 3611 and 3642, and 4303 and 4328. Weaker CaM-binding domains were localized in regions 4, 9, and 10 and in subregions 4b, 9b, and 10a, lying between residues 921 and 1173, 2804 and 2930, and 2961 and 3084. Most of these CaM-binding domains encompassed all or part of previously predicted CaM-binding sites. Strong 45Ca(2+)- and ruthenium red-binding sites domains were localized in the NH2- and COOH-terminal regions of the ryanodine receptor and in regions 6, 12, and 13. The 45Ca(2+- and ruthenium red-binding sites in regions 6 and 12 were localized in subregions 6b and 12b, lying between residues 1861-2094 and 3657-3776. These data together with earlier studies (Chen, S. R. W., Zhang, L., and MacLennan, D. H. (1992) J. Biol. Chem. 267, 23318-23326), show that strong CaM-, Ca(2+)-, and ruthenium red-binding domains are colocalized in the skeletal muscle ryanodine receptor.
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PMID:Identification of calmodulin-, Ca(2+)-, and ruthenium red-binding domains in the Ca2+ release channel (ryanodine receptor) of rabbit skeletal muscle sarcoplasmic reticulum. 752 30

Recent advances in determining the three-dimensional architecture of the skeletal muscle ryanodine receptor/calcium release channel (RyR) by cryo-electron microscopy and three-dimensional reconstruction are discussed. The tetrameric receptor is characterized by a large 4-fold symmetric cytoplasmic assembly that consists of many domains separated by solvent-containing crevices and holes. Experimental evidence suggests that at least one regulatory ligand, calmodulin, binds to sites on the cytoplasmic assembly that are at least 10 nanometers from the transmembrane channel.
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PMID:Three-dimensional architecture of the skeletal muscle ryanodine receptor. 764 82

In the present paper we have defined putative functional domains of the ryanodine receptor Ca2+ channel. cDNA fragments of the skeletal muscle ryanodine receptor were fused in-frame with the Escherichia coli trpe protein and the resulting fusion proteins were evaluated for their ability to react with anti-(ryanodine receptor) antibodies, which are known to block Ca(2+)-dependent activation of the Ca(2+)-release channel. Anti-(ryanodine receptor) antibodies react with epitopes lying within a 245-amino-acid-long polypeptide which is located in a region (residues 4380-4625) encompassing most of myoplasmic loop 2, the predicted transmembrane segment M5 and part of the next lumenal loop (45 residues). Purification of the anti-(ryanodine receptor) antibodies by affinity chromatography led to the isolation of a population of antibodies which was capable of decreasing (by > 30%) the doxorubicin-induced Ca2+ release from isolated terminal cisternae. Polyclonal antibodies raised against a ryanodine receptor fusion encompassing part (198 out of 245 residues) of the immunopositive polypeptide decreased by 2-fold the first-order rate constant of Ca(2+)-induced 45Ca2+ efflux from isolated terminal cisternae. These results suggest strongly that the Ca(2+)-activating domain of the skeletal muscle Ca(2+)-release channel is close to, or associated with, myoplasmic loop 2.
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PMID:Identification of the domain recognized by anti-(ryanodine receptor) antibodies which affect Ca(2+)-induced Ca2+ release. 768 74

In earlier studies (Chen, S. R. W., Zhang, L., and MacLennan, D. H. (1992) J. Biol. Chem. 267, 23318-23326), an amino acid sequence, designated 13c2, lying between amino acid residues 4478 and 4512 in the skeletal muscle ryanodine receptor was shown, through the use of a polyclonal antibody, to be involved in Ca(2+)-induced Ca2+ release. In the present study, an immobilized synthetic peptide, PEPEPEPEPE, corresponding to part of the predicted high affinity Ca2+ binding site between residues 4489 and 4499, was used to purify specific antibodies from an anti-13c2 rabbit antiserum. The effect of this affinity-purified, anti-peptide (anti-13cp1) antibody on Ca2+ release channel function was then characterized using single channel recordings across planar lipid bilayers. The anti-peptide antibody inhibited Ca(2+)- or caffeine-activated channel activities without closing the channel but did not diminish ATP-activated channel activity. The addition of ATP reversed the inhibition of the Ca(2+)- or caffeine-activated channel by the antibody, and the antibody-bound, ATP-activated channel was further modulated by Mg2+, ryanodine, and ruthenium red. The major epitopes in the anti-13c2 antibody, previously shown to activate the Ca2+ release channel by increasing the Ca2+ sensitivity of the channel, did not lie in the PE repeat. These results suggest that the PE repeat sequence forms a site involved in the Ca2+ activation pathway.
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PMID:Antibodies as probes for Ca2+ activation sites in the Ca2+ release channel (ryanodine receptor) of rabbit skeletal muscle sarcoplasmic reticulum. 768 61

Calmodulin (CaM) is a regulator of the calcium release channel (ryanodine receptor) of the sarcoplasmic reticulum of skeletal and cardiac muscle. The locations where CaM binds on the surface of the skeletal muscle ryanodine receptor were determined by electron microscopy. Wheat germ CaM was labeled specifically at Cys-27 with a maleimide derivative of a 1.4-nm-diameter gold cluster, and the gold-cluster-labeled CaM was bound to the purified ryanodine receptor. The complexes were imaged in the frozen-hydrated state by cryoelectron microscopy with no stains or fixatives present. In the micrographs, gold clusters were frequently observed near the corners of the square-shaped images of the ryanodine receptors. In some images, all four corners of the receptor were occupied by gold clusters. Image averaging allowed the site of CaM binding to be determined in two dimensions with an estimated precision of 4 nm. No changes were apparent in the quaternary structure of the ryanodine receptor upon binding CaM to the resolution attained, about 3 nm. Side views of the ryanodine receptor, in which the receptor is oriented approximately perpendicular to the much more frequent fourfold symmetric views, were occasionally observed, and showed that the CaM binding site is most likely on the surface of the receptor that faces the cytoplasm. We conclude that the CaM binding site is at least 10 nm from the transmembrane channel of the receptor and, consequently, that long-range conformational changes are involved in the modulation of the calcium channel activity of the receptor by CaM.
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PMID:Localization of calmodulin binding sites on the ryanodine receptor from skeletal muscle by electron microscopy. 769 69

We have tested the periodate-oxidized ATP analogue 2',3'-dialdehyde adenosine triphosphate (oATP) as a ligand for the skeletal muscle ryanodine receptor/Ca(2+)-release channel. Ca2+ efflux from passively loaded heavy sarcoplasmic reticulum vesicles of skeletal muscle is biphasic. oATP stimulates the initial phase of Ca2+ release in a concentration-dependent manner (EC50 160 microM), and the efflux proceeds with a half-time in the range 100-200 ms. This oATP-modulated initial rapid Ca2+ release was specifically inhibited by millimolar concentrations of Mg2+ and micromolar concentrations of Ruthenium Red, indicating that the effect of oATP was mediated via the ryanodine receptor. The purified Ca(2+)-release channel was incorporated into planar lipid bilayers, and single-channel recordings were carried out to verify a direct interaction of oATP with the ryanodine receptor. Addition of oATP to the cytoplasmic side activated the channel with an EC50 of 76 microM, which is roughly 30-fold higher than the apparent affinity of ATP. The oATP-induced increase in the open probability of the ryanodine receptor displays a steep concentration-response curve with a Hill coefficient of approximately 2, which suggests a co-operativity of the ATP binding sites in the tetrameric protein. oATP binds to the ryanodine receptor in a quasi-irreversible manner via Schiff base formation between the aldehyde groups of oATP and amino groups in the nucleotide binding pocket. This allows for the covalent specific incorporation of [alpha-32P]oATP by borhydride reduction. A typical adenine nucleotide binding site cannot be identified in the primary sequence of the ryanodine receptor. Our results demonstrate that oATP can be used to probe the structure and function of the nucleotide binding pocket of the ryanodine receptor and presumably of other ATP-regulated ion channels.
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PMID:Activation and labelling of the purified skeletal muscle ryanodine receptor by an oxidized ATP analogue. 775 53

The fluorogenic maleimide 7-diethylamino-3-(4'-maleimidylphenyl)-4-methylcoumarin (CPM) has been shown to selectively form Michael adducts with hyperreactive sulfhydryls on the skeletal sarcoplasmic reticulum (SR) ryanodine receptor (RyR1) and triadin which are essential for normal Ca2+ channel function (Liu, G., Abramson, J.J., Zable, A.C., and Pessah, I.N. (1994) Mol. Pharmacol. 45, 189-200). The present report demonstrates a functionally important interaction between RyR1 and triadin which involves, in part, redox cycling of hyperreactive sulfhydryls in response to channel activation and inactivation. Nanomolar CPM is shown to selectively label RyR1 and triadin only in the presence of Ca2+ channel inhibitors (Mg2+, neomycin, ruthenium red, or anti-triadin antibody). Treatment of SR with channel activators (micromolar Ca2+, nanomolar ryanodine, or millimolar caffeine), 1) slows CPM labeling kinetics > 10-fold, 2) negates CPM labeling of channel-associated sulfhydryls, and 3) stabilizes a high molecular weight complex (HMWC) which appears on nonreducing SDS-polyacrylamide gel electrophoresis gels. The HMWC is positively identified as RyR1 and triadin by Western blot and immunoprecipitation analyses. High-affinity [3H]ryanodine-binding sites are immunoprecipitated by either anti-RyR1 or anti-triadin antibody dose dependently. 1,4-Naphthoquinone (< or = 40 pmol/micrograms protein) selectively oxidizes hyperreactive sulfhydryls on RyR1 and triadin, induces Ca2+ efflux from SR, and stabilizes the HMWC. The HMWC is reduced by beta-mercaptoethanol or dithiothreitol into its component RyR1 and triadin promoters. The results provide direct evidence for the existence of a functionally important complex between RyR1 and triadin whose stability is determined by the redox state of hyperreactive sulfhydryl moieties which are allosterically regulated by physiological and pharmacological channel ligands. The present results suggest a possible molecular mechanism by which localized transient changes in the redox state within the RyR1-triadin complex can signal information across the SR membrane.
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PMID:Molecular interaction between ryanodine receptor and glycoprotein triadin involves redox cycling of functionally important hyperreactive sulfhydryls. 780 31

The ryanodine receptor is a channel for Ca2+ release from intracellular stores. By PCR analysis, we identified two alternatively spliced regions in mRNA of the mouse skeletal muscle ryanodine receptor (sRyR). The splice variants were characterized by the presence or absence of 15 bp (ASI) and 18 bp (ASII) exons. The exclusion of these exons results in the absence of the regions corresponding to Ala3481-Gln3485 and Val3865-Asn3870, respectively, of rabbit sRyR; these amino acid sequences exist in the modulatory region, where sites for phosphorylation and binding of Ca2+, calmodulin and ATP are postulated to be. We also detected sRyR in brain and heart as well as in skeletal muscle, and the splicing patterns were found to be tissue-specific. Only the ASII-lacking isoform was detected in heart, whereas in other tissues the ASII-containing isoform was predominant. The splicing patterns were also found to change during development. In skeletal muscle, the ASI-containing isoform increased gradually from embryo to adult. The ASII-lacking isoform abruptly increased upon birth, but the ASII-containing isoform increased steadily afterwards. In cerebrum, the ratio of the ASII-containing isoform to the ASII-lacking one increased abruptly during embryonic days 14 and 18. These findings suggest that the alternative splicing of ASI and ASII, by affecting the modulatory region, generates functionally different sRyR isoforms in a tissue-specific and developmentally regulated manner.
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PMID:Tissue-specific and developmentally regulated alternative splicing in mouse skeletal muscle ryanodine receptor mRNA. 783 48


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