Complex Formation between Junctin, Triadin, Calsequestrin, and the Ryanodine Receptor

Abstract

Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995)J. Biol. Chem. 270, 30787–30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into “KEKE” motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027–9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release. Several key proteins have been localized to junctional sarcoplasmic reticulum which are important for Ca2+ release. These include the ryanodine receptor, triadin, and calsequestrin, which may associate into a stable complex at the junctional membrane. We recently purified and cloned a fourth component of this complex, junctin, which exhibits homology with triadin and is the major 125I-calsequestrin-binding protein detected in cardiac sarcoplasmic reticulum vesicles (Jones, L. R., Zhang, L., Sanborn, K., Jorgensen, A. O., and Kelley, J. (1995)J. Biol. Chem. 270, 30787–30796). In the present study, we have examined the binding interactions between the cardiac forms of these four proteins with emphasis placed on the role of junctin. By a combination of approaches including calsequestrin-affinity chromatography, filter overlay, immunoprecipitation assays, and fusion protein binding analyses, we find that junctin binds directly to calsequestrin, triadin, and the ryanodine receptor. This binding interaction is localized to the lumenal domain of junctin, which is highly enriched in charged amino acids organized into “KEKE” motifs. KEKE repeats are also found in the common lumenal domain of triadin, which likewise is capable of binding to calsequestrin and the ryanodine receptor (Guo, W., and Campbell, K. P. (1995) J. Biol. Chem. 270, 9027–9030). It appears that junctin and triadin interact directly in the junctional sarcoplasmic reticulum membrane and stabilize a complex that anchors calsequestrin to the ryanodine receptor. Taken together, these results suggest that junctin, calsequestrin, triadin, and the ryanodine receptor form a quaternary complex that may be required for normal operation of Ca2+ release. Ultrastructural and biochemical evidence suggests that a protein complex exists at the junctional SR 1The abbreviations used are: SR, sarcoplasmic reticulum; PAGE, polyacrylamide gel electrophoresis; MOPS, 3-(N-morpholino)propanesulfonic acid; CHAPS, 3-[(3-cholamidopropyl)dimethylammino]-1-propanesulfonic acid; GST, glutathione S-transferase; N-junctin, GST fusion protein containing N-terminal, cytoplasmic domain of junctin; C-junctin, GST fusion protein containing C-terminal, lumenal domain of junctin; H-triadin, GST fusion protein containing homologous lumenal domain of triadin. membrane in cardiac and skeletal muscle to facilitate the release of Ca2+ which occurs during muscle contraction (1Johnson E.A. Sommer J.R. J. Cell Biol. 1967; 33: 103-129Crossref PubMed Scopus (113) Google Scholar, 2Fleischer S. Inui M. Annu. Rev. Biophys. Biophys. Chem. 1989; 18: 333-364Crossref PubMed Scopus (444) Google Scholar, 3Meissner G. Annu. Rev. Physiol. 1994; 56: 485-508Crossref PubMed Scopus (842) Google Scholar, 4Franzini-Armstrong C. Jorgensen A.O. Annu. Rev. Physiol. 1994; 56: 509-534Crossref PubMed Scopus (345) Google Scholar). Components of this protein complex identified to date include the ryanodine receptor or Ca2+ release channel, which is visualized by electron microscopy as projecting feet on the cytoplasmic surface of the junctional membrane (2Fleischer S. Inui M. Annu. Rev. Biophys. Biophys. Chem. 1989; 18: 333-364Crossref PubMed Scopus (444) Google Scholar, 4Franzini-Armstrong C. Jorgensen A.O. Annu. Rev. Physiol. 1994; 56: 509-534Crossref PubMed Scopus (345) Google Scholar); calsequestrin, a high capacity Ca2+-binding protein located in the junctional SR lumen, which buffers the calcium that is released during muscle contraction (5Yano K. Zarain-Herzberg A. Mol. Cell. Biochem. 1994; 135: 61-70Crossref PubMed Scopus (139) Google Scholar, 6Fliegel L. Ohnishi M. Carpenter M.R. Khanna V.K. Reithmeier R.A.F. 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PubMed Scopus Google Scholar). evidence suggests that calsequestrin in muscle contraction by the of by the ryanodine receptor M. M. 1989; PubMed Scopus Google Scholar, B. M. PubMed Scopus Google Scholar, M. Biochem. Biophys. 1994; PubMed Scopus Google Scholar, P. M. C. PubMed Scopus Google Scholar). This may be by proteins as triadin J. Biol. Chem. Full Text Full Text PubMed Scopus Google Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google and junctin Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google and triadin J. Biol. Chem. Full Text PubMed Google Scholar, Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google are membrane proteins and amino which with the ryanodine receptor and calsequestrin at the junctional SR membrane in cardiac and skeletal identified as a protein in cardiac and skeletal muscle junctional SR with of filter the protein detected Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar). on this that junctin of the proteins calsequestrin to the SR membrane. junctin purified and cloned and to be in skeletal and cardiac muscle L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). identified as a protein in skeletal muscle junctional SR vesicles S. PubMed Scopus Google Scholar, A.O. J. Biol. Chem. Full Text PubMed Google Scholar). skeletal muscle J. Biol. Chem. Full Text PubMed Google and cardiac Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google triadin cardiac triadin of and on triadin junctin and triadin are the of proteins have membrane that are located in the and which and are highly charged and J. Biol. Chem. Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar, Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). in the lumenal of proteins is the of of and charged which are enriched in and of this have been by as “KEKE” and are to be in the of of binding interactions C. M. 1994; PubMed Scopus Google Scholar, C. M. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar, M. Sci. 1994; PubMed Scopus Google Scholar). GST fusion protein containing the common lumenal domain of triadin, of and the KEKE recently to interact directly with calsequestrin and the ryanodine receptor in skeletal muscle J. Biol. Chem. Full Text Full Text PubMed Scopus Google and cardiac Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). These results evidence for role for triadin, in which calsequestrin to the ryanodine receptor at the junctional SR membrane in cardiac and skeletal muscle J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar, Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). between junctin and triadin suggest that proteins have to date is the role of junctin and in junctin, triadin, is capable of directly with the ryanodine receptor, as as with calsequestrin Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). for triadin of a complex between the ryanodine receptor and calsequestrin been by of fusion protein binding for triadin or with of purified In the we have for the of junctin to to junctional SR proteins important for Ca2+ with on of cardiac membrane we that junctin binds to calsequestrin and the ryanodine receptor also directly with triadin. results for a in which a quaternary protein complex exists between junctin, triadin, calsequestrin, and the ryanodine receptor at the junctional SR membrane. This complex may be important for operation of Ca2+ release during in cardiac and skeletal enriched in junctional SR vesicles by as Jones L.R. 1989; PubMed Scopus Google Scholar). by the of J. Biol. Chem. Full Text PubMed Google Scholar). of cardiac calsequestrin purified by Jones L.R. J. Biol. Chem. Full Text PubMed Google to of to the calsequestrin-affinity chromatography, of cardiac junctional SR vesicles for at in of containing and for at in a in MOPS, and of the calsequestrin-affinity with four of MOPS, and by four with the with and with of the with the to by by with to junctin and triadin. protein with the proteins with in and with a to in polyacrylamide Biophys. PubMed Scopus Google Scholar). cardiac junctin L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google by with and into the of the protein in the as recently for H.K.B. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). junctin with to a highly enriched in the protein Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). of junctin, the protein the and and purified by L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). protein the in and as recently for junctin L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google and found to be by of this of junctin to we to junctin by a with junctin purified by as and the to protein Scholar). junctin in the purified to in by as recently for of Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). of junctin purified of of the protein by and by amino L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). junctin found to be the as recently for junctin L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google and also for triadin J. Biol. Chem. Full Text PubMed Google Scholar). filter used to binding a for proteins Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). of purified junctin with to the binding of cardiac junctional SR vesicles on gel Biophys. PubMed Scopus Google and to for with in MOPS, by for with the in the of of in a of to the in of containing MOPS, and and the for at the of the for with the and binding proteins identified by cardiac junctional SR vesicles for on at a protein of in containing CHAPS, and and proteins by with a at for cardiac junctional SR as in and to the high and also or the in the of with of protein for at with and to to of the and the for at by with of protein for four with containing and including or proteins the by in containing and to L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). binding to proteins to as GST fusion proteins used in this is the GST protein containing the N-terminal, cytoplasmic domain of junctin is the GST fusion protein containing the C-terminal, lumenal domain of junctin and Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google is the GST fusion protein containing the homologous lumenal domain of triadin with the for by K. Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). of N-junctin, of junctin by the cardiac L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google and into of and fusion proteins and purified We that in of C-junctin, of junctin L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google by and in into the of a containing a and a GST of the in and purified with of by S. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). by of GST fusion proteins on of for at containing junctional SR proteins with for at to with the for at in the used for immunoprecipitation and proteins by in and to and binding to proteins to as of cardiac junctional SR vesicles on in of containing CHAPS, and at for in a and the on the of a in a the with the to and the to for at of and to and for binding Jones L.R. 1989; PubMed Scopus Google Scholar). the purified ryanodine receptor used in the fusion protein binding the the in to the to binding to the ryanodine receptor or to to in and binding in the of with to Jones L.R. 1989; PubMed Scopus Google Scholar). containing to and Biophys. PubMed Scopus Google or PubMed Scopus Google Scholar). of ryanodine polyacrylamide and for of polyacrylamide with as to proteins L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). used in this a K. This triadin Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). to the of cardiac triadin as as to the purified protein also in and to junctin L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google and calsequestrin L. Jones L.R. J. Biol. Chem. Full Text PubMed Google in to the cardiac proteins and purified as J. Biol. Chem. Full Text PubMed Google Scholar). to junctin as the cardiac of the ryanodine receptor to a fusion protein containing amino acids of the cardiac ryanodine receptor Jones Biophys. J. Scholar). In we identified junctin as the major protein in cardiac skeletal junctional SR vesicles by of a filter in which with SR proteins to Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). interaction between calsequestrin and junctin by Ca2+ binding to calsequestrin, with at Several proteins also detected in including triadin and a protein the Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). the filter the of we to for proteins calsequestrin-affinity cardiac junctional SR vesicles with and on the in proteins in containing a calcium to calsequestrin of the with triadin that triadin to the and as a in of triadin detected in by a triadin Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google as as to the of cardiac triadin This suggests that the cardiac triadin and Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google detected in in cardiac SR of the junctin to the as by of the purified junctin the triadin. protein by the junctin which the is to be a junctin L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google and is with Ca2+ in and by a of junctin and triadin in the L. Jones L.R. J. Biol. Chem. Full Text PubMed Google that junctin and triadin in these purified and junctional SR These results that junctin and triadin are major proteins and that the binding interaction of calsequestrin with these proteins is for interactions between junctin and calsequestrin, immunoprecipitation with junctional SR proteins or cardiac SR vesicles by purified In with results the Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Ca2+ the binding interaction between calsequestrin and junctin, with between and immunoprecipitation with calsequestrin, and the binding interaction a Ca2+ These results that junctin binds to calsequestrin and that the interaction between these proteins occurs in on these a of calsequestrin is to to junctin at the Ca2+ to in junctional SR in cardiac muscle C. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). the calsequestrin interaction domain of junctin, binding with GST fusion proteins and with cardiac junctional SR proteins in to to or GST These results that the domain of junctin is localized to the lumenal of the with immunoprecipitation the binding interaction between and calsequestrin in the of Ca2+ at proteins by the the are the proteins Jones L.R. J. Biol. Chem. Full Text PubMed Google which in these These proteins with calsequestrin and of amino with calsequestrin B.T. Jones L.R. Cell Biol. Google Scholar). proteins to be of calsequestrin B.T. Jones L.R. Cell Biol. Google Scholar). proteins also in the in and are in these for immunoprecipitation of cardiac junctional SR as in the to this purified to cardiac triadin with triadin calsequestrin and junctin of the to proteins Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google to the to binding this junctin in purified by chromatography, and that purified junctin as a on as visualized by or by of the protein with the junctin that junctin on as junctin in cardiac SR cardiac SR proteins directly with junctin, junctional SR proteins by to and with that of the proteins present in SR vesicles visualized by a proteins calsequestrin, the triadin, and junctin of these proteins by with calsequestrin, triadin, and junctin protein the is the protein in which is by the junctin binding of junctin to triadin detected by filter that junctin in SR interact directly with triadin. this cardiac SR proteins with the triadin This junctin and calsequestrin, the the interaction the interaction also by Ca2+ of junctin by triadin that the binding interaction between junctin and triadin of for for a interaction between junctin and triadin, These containing the common lumenal domain of triadin, junctin cardiac SR vesicles with immunoprecipitation of the proteins triadin binding to junctin by the Ca2+ also purified junctin a interaction between these the binding interaction for junctin binding to the cardiac ryanodine receptor, we cardiac junctional SR proteins in with junctin to protein for ryanodine by with a ryanodine receptor and also by binding that a of the ryanodine with of the junctin ryanodine binding that of the by the junctin that junctin to of the junctin. These results suggest that a of the cardiac ryanodine to junctin in the junctin of ryanodine in or the of junctin with the cardiac ryanodine receptor, and with cardiac junctional SR proteins in fusion which been to to the ryanodine receptor Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google as a and of the ryanodine as detected by binding or by or GST to interact with ryanodine binding of the lumenal domain of junctin to the ryanodine receptor at of junctin fusion protein of junctional SR protein This suggests that the of the immunoprecipitation in the of the junctin to of the ryanodine a of for of ryanodine of cardiac junctional SR vesicles with of fusion protein to glutathione and ryanodine by and in the by binding In ryanodine to the detected by with ryanodine receptor that junctin binds directly to the ryanodine receptor and to the of binding proteins triadin, we purified the cardiac ryanodine receptor to and for to interact with binding to the purified ryanodine receptor also purified cardiac ryanodine receptor a binding of protein and as high protein on calsequestrin, junctin, or triadin detected in the purified by with these of the binding by the or which by with the ryanodine receptor the lumenal of junctin and triadin directly to the ryanodine receptor with proteins the interaction of junctin with triadin, binding of these fusion proteins to the purified ryanodine receptor It been that junctin and triadin are for the proteins calsequestrin to the ryanodine receptor at the junctional membrane Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar, J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar, Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). evidence to date this been for role as protein to in filter Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar, C. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google evidence for role as protein the of fusion proteins to calsequestrin and the ryanodine receptor in binding J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar, Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). between junctin and triadin and proteins to SR have been by a combination of approaches including chromatography, immunoprecipitation protein and fusion protein we have for a for junctin and triadin at the junctional SR membrane. results a complex of proteins that may be required for of Ca2+ release in cardiac We the of junctin to interact directly with calsequestrin, triadin, and the ryanodine receptor in of these interactions is in identified as a major protein of junctional SR vesicles by of a filter binding Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). In the present we the binding of junctin to calsequestrin calsequestrin-affinity of junctin to the in as with of the Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, A. Biochem. Biophys. PubMed Scopus Google Scholar). of the junctin cardiac SR vesicles to the the triadin is to this of junctin or triadin binds to This is of the calsequestrin in cardiac SR vesicles the with the junctin that This junctin have been with the of junctin the calsequestrin-affinity in with the of triadin suggests that junctin may to calsequestrin triadin. with the purified proteins be required to the of triadin for the of junctin with calsequestrin at the junctional SR membrane in this study, with SR vesicles that the is protein localized the domain of junctin to amino the containing charged also a high of charged B.T. Simmerman H.K.B. Collins J.H. Nadal-Ginard B. Jones L.R. J. Biol. Chem. 1988; 263: 8958-8964Abstract Full Text PubMed Google that between junctin and calsequestrin may stabilize the binding interaction with of or of the protein interactions of junctin binding to calsequestrin as or Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google Scholar, L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar, A. Biochem. Biophys. PubMed Scopus Google assays, that interactions may be for protein of calsequestrin binding Ca2+ Simmerman H.K.B. Jones L.R. J. Biol. Chem. 1988; 263: Full Text PubMed Google is for of interaction with junctin and triadin at high the Ca2+ in the SR C. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google calsequestrin to junctin and triadin the proteins are into immunoprecipitation or fusion protein binding of and charged amino acids at the of the B.T. Simmerman H.K.B. Collins J.H. Nadal-Ginard B. Jones L.R. J. Biol. Chem. 1988; 263: 8958-8964Abstract Full Text PubMed Google and are in to this in binding to junctin this is that junctin binds directly to triadin. approaches used to this In the the we that junctin directly to the triadin In the triadin junctin junctional SR proteins and this interaction of for In the to junctin SR vesicles as as purified junctin. In to junctin and triadin interactions with calsequestrin, the junctin interaction with triadin junctin binding domain localized to of triadin, the common lumenal domain also with calsequestrin and the ryanodine receptor Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). appears that the highly charged lumenal of junctin and triadin are capable of binding to calsequestrin, to the ryanodine receptor to common to the lumenal of triadin and junctin is highly charged charged with of and charged amino C. M. 1994; PubMed Scopus Google Scholar, C. M. J. Biol. Chem. Full Text Full Text PubMed Scopus Google have that KEKE are present in a of Ca2+-binding proteins and have that charged protein protein as have been by M. Sci. 1994; PubMed Scopus Google Scholar). We that the KEKE by junctin and triadin are required for interactions with as as with junctional SR proteins that a high of charged that that is capable of binding to on as as to the protein of which is to be a junctin L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar). This suggests the for the of protein in the of the SR membrane junctin and triadin are present in high these protein may calsequestrin to the membrane in to the ryanodine receptor. in protein and between triadin and junctin to junctin, triadin J. Biol. Chem. Full Text Full Text PubMed Scopus Google Scholar, Jorgensen A.O. Jones L.R. J. Biol. Chem. Full Text Full Text PubMed Scopus Google is capable of binding to the ryanodine receptor. We this interaction that junctin and triadin directly to the purified ryanodine receptor with proteins with the junctin that junctin in cardiac SR vesicles binds a of the ryanodine the by to of the ryanodine with the junctin be to a of the containing triadin for the junctin binding or to as a binding of the ryanodine receptor for junctin the proteins are the membrane and into the lumenal of junctin and triadin are for binding to the ryanodine receptor, appears that in muscle these proteins interact with the ryanodine receptor the lumenal surface of the SR membrane. of the cardiac and skeletal muscle ryanodine and is highly and is enriched in charged amino acids J. M. S. PubMed Scopus Google Scholar). It is that of the ryanodine receptor binds to KEKE common to triadin and junctin. on biochemical with purified junctin and triadin interactions with the ryanodine receptor are to be In a for the protein interactions at the junctional membrane junctin and triadin highly in the of the membrane and directly with C-terminal, lumenal of junctin and triadin to the binding of calsequestrin, which by is of binding to the ryanodine receptor. lumenal of junctin and triadin also directly to the ryanodine receptor and by this calsequestrin to the membrane in to the Ca2+ release and triadin interactions with the ryanodine receptor are junctin and triadin binding to calsequestrin may be for as the the junctional SR and during muscle and of binding between the four proteins of the complex may be by interactions between KEKE of junctin and triadin and the of these proteins to by charged is a protein component of skeletal and cardiac muscle junctional SR L.R. L. K. Jorgensen A.O. J. J. Biol. Chem. Full Text Full Text PubMed Scopus Google and may be that this protein as to and at the junctional of the of junctin as a to calsequestrin at the SR or also a role in Ca2+ release be that have that Ca2+ binding to calsequestrin directly the of by the ryanodine receptor M. M. 1989; PubMed Scopus Google Scholar, B. M. PubMed Scopus Google Scholar, M. Biochem. Biophys. 1994; PubMed Scopus Google Scholar, P. M. C. PubMed Scopus Google and that have junctin and to which may be junctin in this M. M. 1989; PubMed Scopus Google Scholar, M. Biochem. Biophys. PubMed Scopus Google Scholar, M. Biochem. Biophys. PubMed Scopus Google Scholar). We K. for with the and for the triadin