Cardiac Type 2 Inositol 1,4,5-Trisphosphate Receptor

Abstract

The type 2 inositol 1,4,5-trisphosphate receptor (InsP3R2) was identified previously as the predominant isoform in cardiac ventricular myocytes. Here we reported the subcellular localization of InsP3R2 to the cardiomyocyte nuclear envelope (NE). The other major known endo/sarcoplasmic reticulum calcium-release channel (ryanodine receptor) was not localized to the NE, indicating functional segregation of these channels and possibly a unique role for InsP3R2 in regulating nuclear calcium dynamics. Immunoprecipitation experiments revealed that the NE InsP3R2 associates with Ca2+/calmodulin-dependent protein kinase IIδ (CaMKIIδ), the major isoform expressed in cardiac myocytes. Recombinant InsP3R2 and CaMKIIδB also co-immunoprecipitated after co-expression in COS-1 cells. Additionally, the amino-terminal 1078 amino acids of the InsP3R2 were sufficient for interaction with CaMKIIδB and associated upon mixing following separate expression. CaMKII can also phosphorylate InsP3R2, as demonstrated by 32P labeling. Incorporation of CaMKII-treated InsP3R2 into planar lipid bilayers revealed that InsP3-mediated channel open probability is significantly reduced (∼11 times) by phosphorylation via CaMKII. We concluded that the InsP3R2 and CaMKIIδ likely represent two central components of a multiprotein signaling complex, and this raises the possibility that calcium release via InsP3R2 in the myocyte NE may activate local CaMKII signaling, which may feedback on InsP3R2 function. The type 2 inositol 1,4,5-trisphosphate receptor (InsP3R2) was identified previously as the predominant isoform in cardiac ventricular myocytes. Here we reported the subcellular localization of InsP3R2 to the cardiomyocyte nuclear envelope (NE). The other major known endo/sarcoplasmic reticulum calcium-release channel (ryanodine receptor) was not localized to the NE, indicating functional segregation of these channels and possibly a unique role for InsP3R2 in regulating nuclear calcium dynamics. Immunoprecipitation experiments revealed that the NE InsP3R2 associates with Ca2+/calmodulin-dependent protein kinase IIδ (CaMKIIδ), the major isoform expressed in cardiac myocytes. Recombinant InsP3R2 and CaMKIIδB also co-immunoprecipitated after co-expression in COS-1 cells. Additionally, the amino-terminal 1078 amino acids of the InsP3R2 were sufficient for interaction with CaMKIIδB and associated upon mixing following separate expression. CaMKII can also phosphorylate InsP3R2, as demonstrated by 32P labeling. Incorporation of CaMKII-treated InsP3R2 into planar lipid bilayers revealed that InsP3-mediated channel open probability is significantly reduced (∼11 times) by phosphorylation via CaMKII. We concluded that the InsP3R2 and CaMKIIδ likely represent two central components of a multiprotein signaling complex, and this raises the possibility that calcium release via InsP3R2 in the myocyte NE may activate local CaMKII signaling, which may feedback on InsP3R2 function. The inositol 1,4,5-trisphosphate receptor (InsP3R) 1The abbreviations used are: InsP3R, inositol 1,4,5-trisphosphate receptor; InsP3R2, inositol 1,4,5-trisphosphate receptor type 2; NE, nuclear envelopes; RyR, ryanodine receptor; CaMKIIδ, calcium/calmodulin-dependent protein kinase IIδ; DTT, dithiothreitol; PMSF, phenylmethanesulfonyl fluoride; SR, sarcoplasmic reticulum; NS, nuclei storage; HA, hemagglutinin; IP, immunoprecipitation; T2NH, amino-terminal anti-InsP3R2 antibody; T2C, carboxyl-terminal anti-InsP3R2 antibody; NP, nucleoplasm; HDAC, histone deacetylase; InsP3inositol 1,4,5-trisphosphate. family of calcium-release channels occupies a central role in the initiation and propagation of intracellular calcium release events. These release events regulate a plethora of cellular signaling processes (reviewed in Ref. 1Furuichi T. Michikawa T. Mikoshiba K. Carafoli E. Klee C.B. Calcium as a Cellular Regulator. Oxford University Press, New York1999: 200-248Google Scholar). The type 2 InsP3R (InsP3R2) isoform has been identified previously as the predominant isoform expressed in ventricular and atrial cardiomyocytes (2Moschella M.C. Marks A.R. J. Cell Biol. 1993; 120: 1137-1146Crossref PubMed Scopus (122) Google Scholar, 3Perez P.J. Ramos-Franco J. Fill M. Mignery G.A. J. Biol. Chem. 1997; 272: 23961-23969Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar, 4Lipp P. Laine M. Tovey S.C. Burrell K.M. Berridge M.J. Li W. Bootman M.D. Curr. Biol. 2000; 10: 939-942Abstract Full Text Full Text PDF PubMed Scopus (204) Google Scholar). Characterization of InsP3R2 channels from ventricular myocytes in planar lipid bilayers revealed that the permeation properties are very similar to the type 1 receptor (InsP3R1) from cerebellum but appear to be regulated differently. The InsP3R2 channel is more sensitive to InsP3 compared with InsP3R1, and although calcium-dependent, the channel can be fully activated by InsP3 at a [Ca2+] above 100 nm (5Südhof T.C. Newton C.L. Archer B.T. II I Ushkaryov Y.A. Mignery G.A. EMBO J. 1991; 10: 3199-3206Crossref PubMed Scopus (320) Google Scholar, 6Newton C.L. Mignery G.A. Südhof T.C. J. Biol. Chem. 1994; 269: 28613-28619Abstract Full Text PDF PubMed Google Scholar, 7Ramos-Franco J. Fill M. Mignery G.A. Biophys. J. 1998; 75: 834-839Abstract Full Text Full Text PDF PubMed Scopus (154) Google Scholar, 8Ramos-Franco J. Bare D. Caenepeel S. Nani A. Fill M. Mignery G. Biophys. J. 2000; 79: 1388-1399Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar). InsP3R2 does not play a major regulatory role in global [Ca2+] homeostasis in ventricular myocytes, and its functional significance is unclear (9Marks A.R. Circ. Res. 2000; 87: 8-11Crossref PubMed Scopus (194) Google Scholar). However, in atrial myocytes InsP3R2 has been reported to be localized to junctional SR and may participate in arrhythmogenesis (10Mackenzie L. Bootman M.D. Laine M. Berridge M.J. Thuring J. Holmes A. Wen-Hong L. Lipp P. J. Physiol. (Lond.). 2002; 541: 395-409Crossref Scopus (195) Google Scholar, 11Zima A.V. Blatter L.A. J. Physiol. (Lond.). 2004; 555: 607-615Crossref Scopus (168) Google Scholar). For example, calcium released by atrial InsP3 may trigger calcium-induced calcium release via RyR, thereby accentuating calcium oscillations (11Zima A.V. Blatter L.A. J. Physiol. (Lond.). 2004; 555: 607-615Crossref Scopus (168) Google Scholar). A role for InsP3R in cardiac dysfunction has been suggested by alterations in expression levels of the failing heart in which there appears to be an up-regulation of InsP3R while RyR2 levels decrease (12Go L.O. Moschella M.C. Watras J. Handa K.K. Fyfe B.S. Marks A.R. J. Clin. Investig. 1995; 95: 888-894Crossref PubMed Google Scholar). Also, significant evidence has been presented for the involvement of InsP3R in pathologies leading to calcium overload and apoptotic cardiomyocyte death, as well as implications in the pathophysiology of cardiac arrhythmia (10Mackenzie L. Bootman M.D. Laine M. Berridge M.J. Thuring J. Holmes A. Wen-Hong L. Lipp P. J. Physiol. (Lond.). 2002; 541: 395-409Crossref Scopus (195) Google Scholar, 13Felzen B. Shilkrut S. Less H. Sarapov I. Maor G. Coleman R. Robinson R.B. Berke G. Binah O. Circ. Res. 1998; 82: 438-450Crossref PubMed Scopus (70) Google Scholar). In this study, we have identified the nuclear envelope as the primary site for subcellular localization of the type 2 InsP3Rin ventricular myocytes. This highly localized expression may support the functional encoding of InsP3-mediated calcium release in calcium-dependent nuclear specific signaling. Consistent with this view, immunoprecipitation experiments revealed that the type 2 InsP3R in isolated cardiac nuclei associates with Ca2+/calmodulin-dependent protein kinase II (CaMKII). Co-expression of CaMKIIδB and InsP3R2 followed by immunoprecipitation verifies the interaction and that the InsP3R2 is phosphorylated by CaMKII. Additionally, CaMKII-mediated phosphorylation of InsP3R2 results in a dramatic decline in channel open probability, suggesting a feedback mechanism. Whole Heart Subcellular Fractionation—The heart fractionation procedure used was a modified procedure of Kapiloff et al. (14Kapiloff M.S. Jackson N. Airhart N. J. Cell Sci. 2001; 114: 3167-3176Crossref PubMed Google Scholar). Frozen adult rat hearts (Pel-Freez Biologicals, Rogers, AR) were homogenized in Buffer A (10 1 phenylmethanesulfonyl 1 and with a and and The was for at and the was for the of The was to a followed by a The was and on a for at The were at the and in Buffer A. from above was in Buffer nuclei were at for The nuclei were in nuclei 1 1 DTT, PMSF, 1 and and at the the of SR in the and an SR was the of the the was and with and the were isolated via for at The was and in was by of and were a of the by et al. Cell Res. PubMed Scopus Google and et al. B. H. J. Full Text PDF PubMed Scopus Google Scholar). were on and in I and A were to for and The nuclei were in were from by The was and the nuclear were in and at COS-1 Cell were with expression for rat type 2 J. Bare D. Caenepeel S. Nani A. Fill M. Mignery G. Biophys. J. 2000; 79: 1388-1399Abstract Full Text Full Text PDF PubMed Scopus (72) Google 1078 amino acids of rat type 2 (5Südhof T.C. Newton C.L. Archer B.T. II I Ushkaryov Y.A. Mignery G.A. EMBO J. 1991; 10: 3199-3206Crossref PubMed Scopus (320) Google CaMKIIδB M. H. J. Cell Biol. 1994; PubMed Scopus Google and M. H. J. Cell Biol. 1994; PubMed Scopus Google a as previously G.A. Newton C.L. Archer B.T. II I Südhof T.C. J. Biol. Chem. Full Text PDF PubMed Google Scholar). from COS-1 were as previously J. Bare D. Caenepeel S. Nani A. Fill M. Mignery G. Biophys. J. 2000; 79: 1388-1399Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar, G.A. Newton C.L. Archer B.T. II I Südhof T.C. J. Biol. Chem. Full Text PDF PubMed Google Scholar). for in experiments were as by et al. E. I. J. Physiol. 1998; PubMed Scopus Google Scholar). COS-1 were in 1 PMSF, 100 and with a of of was and the was and for at The the was and and were to a of and The was homogenized and for at a the were for at and the was the were at for in and in ventricular cardiomyocytes were isolated as by et al. J. Physiol. (Lond.). Scopus Google with and for as previously J. Bare D. Caenepeel S. Nani A. Fill M. Mignery G. Biophys. J. 2000; 79: 1388-1399Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar). were used of the was a for and a and Immunoprecipitation of from rat and from COS-1 were homogenized and in immunoprecipitation 1 100 and followed by of by at The was and primary were and was and were with and in and The were by T2NH, and used in this have been previously J. Fill M. Mignery G.A. Biophys. J. 1998; 75: 834-839Abstract Full Text Full Text PDF PubMed Scopus (154) Google Scholar, 8Ramos-Franco J. Bare D. Caenepeel S. Nani A. Fill M. Mignery G. Biophys. J. 2000; 79: 1388-1399Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar, J. Caenepeel S. Fill M. Mignery G.A. Biophys. J. 1998; 75: Full Text Full Text PDF PubMed Scopus Google Scholar). receptor was and a a RyR2 protein G. A. S. M. J. Cell Biol. 1995; PubMed Scopus Google Scholar). Ca2+/calmodulin-dependent protein kinase II and used for and a were from and and were as previously G.A. Newton C.L. Archer B.T. II I Südhof T.C. J. Biol. Chem. Full Text PDF PubMed Google and was CaMKII of InsP3R2 were from COS-1 and used as for in CaMKII phosphorylation as for 32P a were for at with CaMKII by in the of the specific from isolated cardiac nuclei were used as a in the in phosphorylation with and CaMKII. The were on to to and with CaMKII of InsP3R2 were with InsP3R2, and were as for planar lipid were for at in the of CaMKII with as well as kinase as a The specific CaMKII used was also the of the were as for the in phosphorylation of InsP3R2 the of with to to CaMKII were in CaMKII phosphorylation with to and PMSF, 100 and at for The were by at and in the to at for with CaMKII In the protein was in CaMKII phosphorylation the were in CaMKII and The were in in and for into planar lipid channel of InsP3R type 2 were by into planar lipid were a in the of a a lipid of and The two and The were to the of the 1 and in the calcium was and with a 2 InsP3 and ryanodine were The was at The channels were identified by to InsP3 and probability was by the from at were a The was at with a and at 1 with a and were and Subcellular of the 2 InsP3R to the of InsP3 receptor (InsP3R) isoform expression in cardiac demonstrated that the predominant isoform expressed in the ventricular cardiomyocyte is InsP3R2 P.J. Ramos-Franco J. Fill M. Mignery G.A. J. Biol. Chem. 1997; 272: 23961-23969Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar, 8Ramos-Franco J. Bare D. Caenepeel S. Nani A. Fill M. Mignery G. Biophys. J. 2000; 79: 1388-1399Abstract Full Text Full Text PDF PubMed Scopus (72) Google Scholar). the role of InsP3R2 in the we a subcellular fractionation of rat heart a modified procedure of Kapiloff et al. (14Kapiloff M.S. Jackson N. Airhart N. J. Cell Sci. 2001; 114: 3167-3176Crossref PubMed Google Scholar). Heart were to to separate SR, from components as and nuclei The was and by The nuclei were from the by were isolated from the by These of protein were with and carboxyl-terminal of the type 1 and and InsP3R and Additionally, an the A and was in the the nuclei was the significant to the and A and This with the InsP3R2 in the A and type 1 receptor was in the nuclei the and not The of nuclei was by the of the type A A and This a in the and and to a a The of was demonstrated in the isolated nuclei The and revealed a following significantly suggesting a of from This fractionation results suggesting that the primary subcellular site for InsP3R2 localization appears to be the with to other cardiomyocyte reported by Kapiloff et al. (14Kapiloff M.S. Jackson N. Airhart N. J. Cell Sci. 2001; 114: 3167-3176Crossref PubMed Google fractionation was to significant nuclear the and the mixing of NE and SR In a of nuclear and SR was suggested by the with the in the However, revealed that a of SR with the in the In to SR the nuclei were isolated via the was and with and the SR were by A of of the SR with NE was via the InsP3R2 and from the in a in the NE and a in the This SR be to a of levels of InsP3R2 expressed in SR as reported by Lipp et al. P. Laine M. Tovey S.C. Burrell K.M. Berridge M.J. Li W. Bootman M.D. Curr. Biol. 2000; 10: 939-942Abstract Full Text Full Text PDF PubMed Scopus (204) Google for atrial myocytes and from nuclear as a of other The a in the SR and a in the NE with SR were used as a in of the InsP3R2 in T. Mikoshiba K. 2000; PubMed Scopus Google Scholar, Biophys. Res. 2000; PubMed Scopus Google Scholar, K. O. G. J. 2002; PubMed Scopus Google Scholar, 555: PubMed Scopus Google the localization of InsP3R2 in as a nuclear that appears to be localized the and results the of InsP3R2 in the nuclear envelope of we on isolated rat ventricular myocytes. were and with type amino-terminal in a very the is A and a of InsP3R2 in the cardiomyocyte NE as by the the of appears to be Consistent with this are the results of Lipp et al. P. Laine M. Tovey S.C. Burrell K.M. Berridge M.J. Li W. Bootman M.D. Curr. Biol. 2000; 10: 939-942Abstract Full Text Full Text PDF PubMed Scopus (204) Google in which the type 2 receptor was at levels in the junctional SR of atrial myocytes that were also to more receptor ventricular myocytes. In to the NE were which in the of of and A.V. J. R. Blatter L.A. J. Physiol. (Lond.). Scopus Google Scholar). the myocyte nuclei was in and We were not to type 2 in the SR of the ventricular myocytes. with the InsP3R and these revealed a similar to previously for the type 2 The was to with an protein and was not used for of ventricular myocytes for the ryanodine receptor in from SR components and was not in the nuclear envelope and This localization is with the of et al. J. Bare Mignery G.A. Blatter L.A. Biophys. J. 2001; Full Text Full Text PDF PubMed Scopus Google were in the nuclear envelope of atrial myocytes. These results that the two intracellular release channels are in the myocyte and the possibility that the InsP3R2 is in local nuclear signaling events. of the results in cardiac with the amino-terminal and carboxyl-terminal InsP3R2 we isolated nuclei into NE and and the for InsP3R2 expression. of NE, and were on and with the and in the results that InsP3R2 is to the in nuclear with the InsP3R a This is demonstrated by the of the type 2 receptor in NE as compared with nuclei and and by the of in following the protein in nuclei and the The of the InsP3R2 in the nuclear envelope is not known and is the NE of InsP3R2 and of a of nuclear localization of (14Kapiloff M.S. Jackson N. Airhart N. J. Cell Sci. 2001; 114: 3167-3176Crossref PubMed Google we similar with an to and significant was The of localized to the NE is with as well as the of J. Bare Mignery G.A. Blatter L.A. Biophys. J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar, W. J. Cell Biol. 1993; 120: PubMed Scopus Google Scholar, G. S. E. 1997; PubMed Scopus Google Scholar, J. M. J. Cell Biol. 1998; PubMed Scopus Google Scholar). were also with the of these with the NE and the of nuclear in the These results that InsP3R2 is to the cardiomyocyte nuclear and as expression is to the SR 2 and with functional for these two intracellular calcium-release channels in the ventricular of InsP3R2 with CaMKIIδ as a localization of the type 2 InsP3 receptor to the NE of myocytes a in the of local nuclear signaling events in the cardiac myocyte as the of The of local calcium the of the has been suggested for P. G. Sci. S. A. 87: PubMed Scopus Google Scholar, L. A. 1995; Full Text PDF PubMed Scopus Google Scholar, L. 1995; PubMed Scopus Google Scholar). and H. 1998; PubMed Scopus Google into a for CaMKII as a of intracellular calcium into of kinase experiments were at the type 2 InsP3R on the NE participate in local CaMKII signaling from heart were followed by immunoprecipitation with the and the was with as well as with a CaMKII in the type 2 InsP3R and with the CaMKII revealed an with a of which is with that of the predominant cardiac isoform of In a were with an CaMKII the with also that CaMKIIδ was with InsP3R2 The of the InsP3R2 was by not These results that InsP3R2 and CaMKIIδ are associated and may the of a signaling of InsP3R2 and CaMKIIδ by immunoprecipitation from cardiac nuclei of CaMKIIδ with the InsP3R2 was expression in COS-1 cells. InsP3R2 was with CaMKIIδB M. H. J. Cell Biol. 1994; PubMed Scopus Google in were and into and the InsP3R2 and associated CaMKII were with the InsP3R2 and and the an The from were for the InsP3R2 as well as in the in the of InsP3R2 and in the In a InsP3R2 co-immunoprecipitated the of with InsP3R2 and InsP3R2 were not Co-expression experiments a carboxyl-terminal of the InsP3R2 with were to the amino-terminal of the receptor with The the amino-terminal 1078 of the receptor and the as well as of the and is a protein (5Südhof T.C. Newton C.L. Archer B.T. II I Ushkaryov Y.A. Mignery G.A. EMBO J. 1991; 10: 3199-3206Crossref PubMed Scopus (320) Google Scholar). protein from COS-1 the two were to immunoprecipitation with and in the and expression as a The to the in the of the expression Additionally, a mixing from that were with was also from the in of the not These results from cardiac nuclei and that at site of interaction the two is localized to the InsP3R2 amino-terminal 1078 the of the interaction with the receptor is not results the amino-terminal 1078 that receptor with CaMKII and that as a of receptor likely does not this site for CaMKII of InsP3R2 by that InsP3-mediated calcium release is in the of nuclear signaling and the two a raises the possibility that InsP3R2 may be a for CaMKII phosphorylation events. In experiments at CaMKII the InsP3R2, we expressed the type 2 receptor in COS-1 cells. were and the receptor was The was used as for in kinase with CaMKII New The was from the kinase and on to and in of CaMKII results in the phosphorylation of the InsP3R2, and this of 32P can be by CaMKII was from the we a suggesting that InsP3R2 was phosphorylated by associated We the of 32P as a of the expressed receptor with levels of CaMKII to COS-1 associated This with of the InsP3R2 was phosphorylated to a in the of CaMKII This was by of the CaMKII and demonstrated specific phosphorylation of to the was with T2NH, and the from the and that the was the InsP3R2 and that the similar of Additionally, in CaMKII in the phosphorylation of an amino-terminal of the 1078 from COS-1 The of 32P that of the type 2 CaMKII in significant of the InsP3R2 and this be by the of and in the kinase CaMKII and revealed levels of 32P These that in to with this of the InsP3R2, CaMKII also the receptor this that CaMKII phosphorylation This does not the possibility that CaMKII and phosphorylation are the to this The results were from receptor and CaMKII. a that this phosphorylation in the of the localized in cardiac we the from isolated cardiac nuclei and in kinase in the receptor from nuclei 32P in to in the CaMKII. This is significantly reduced in the of the The levels of 32P in the of this as well as expressed InsP3R2, are to as a of CaMKII that are not sensitive to the is also that other associated are to the labeling. These results revealed that InsP3R2 is phosphorylated in by CaMKII. local InsP3-mediated calcium signaling may not be in the of CaMKII-mediated signaling but CaMKII phosphorylation of the receptor may InsP3R2 channel by of InsP3R2 by CaMKII the phosphorylation of InsP3R2 by CaMKII an on channel we used of CaMKII to the The used from COS-1 that were in the kinase in the of CaMKII as well as CaMKII in the of the These were into planar lipid bilayers for channel presented in the revealed channels with a open probability of In with CaMKII in channels that an in open probability However, the with CaMKII in the of the open similar to of the and In a from COS-1 with InsP3R2 were with These were in the in kinase in the of CaMKII with The channels were in planar lipid in the channels of the similar open as the of the indicating that the and in the kinase on the channel However, the with CaMKII a significant times) in the open probability compared with the The with the in channels with open that are similar to of the with this channel was which is likely a of the to channel the results the were very similar to the in a COS-1 were with InsP3R2 and from these and from with InsP3R2 were into in the channels from of the a very open probability compared with the expressed InsP3R2 These results are with the results from the two other of CaMKII to the InsP3R2 and that CaMKII-mediated phosphorylation of InsP3R2 results in a dramatic of the calcium-release channel open The channel of CaMKII-treated was by open probability, and and compared with InsP3R2 and the the for these events is not This may represent a feedback for InsP3-mediated calcium release and signaling events. The of this of InsP3R2 channel as of InsP3 and calcium is The type 2 InsP3R isoform has been identified previously as the predominant isoform expressed in the ventricular The channel is activated by levels of InsP3 and is to by [Ca2+] P.J. Ramos-Franco J. Fill M. Mignery G.A. J. Biol. Chem. 1997; 272: 23961-23969Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar, 7Ramos-Franco J. Fill M. Mignery G.A. Biophys. J. 1998; 75: 834-839Abstract Full Text Full Text PDF PubMed Scopus (154) Google Scholar). In this study, we that the nuclear envelope is the primary site of subcellular localization of the InsP3R2 in the adult Additionally, we that the receptor a with and is a for CaMKII. These that the InsP3R2 is in nuclear calcium signaling in the cardiac The role of the InsP3R2 in myocytes has A significant role for calcium release from nuclear envelope in the of local calcium the of the has been for P. G. Sci. S. A. 87: PubMed Scopus Google Scholar, L. A. 1995; Full Text PDF PubMed Scopus Google Scholar, L. 1995; PubMed Scopus Google Scholar). have that an calcium-release channel of nuclear calcium appears (reviewed in Ref. M.D. D. Tovey S.C. Berridge M.J. Lipp P. Sci. 2000; PubMed Scopus Google Scholar). the of localized calcium release in the after and the of InsP3 with has demonstrated regulated nuclear calcium that appear to be into the via a reticulum that is with the envelope W. Cell Biol. PubMed Scopus Google Scholar). The of the NE of the InsP3R2 a calcium-release channel at a subcellular site that be in the of signaling from calcium In support of isolated nuclei used in this with calcium and have been to release calcium in an In the of nuclei with InsP3 raises nuclear calcium by The possibility that the myocyte nuclear envelope a significant calcium is demonstrated by the of in to InsP3 and A. Bare Mignery G.A. Blatter L.A. Biophys. J. Scholar). has been presented suggesting that CaMKII is of the of calcium into of nuclear kinase H. 1998; PubMed Scopus Google and that CaMKII is in calcium-dependent of nuclear in cardiac myocyte signaling H. J. Biol. Chem. 1997; 272: Full Text Full Text PDF PubMed Scopus Google Scholar). The possibility that nuclear InsP3-mediated signaling is in the of CaMKII is with a by et al. J. T. T. 2004; Google the of ventricular myocytes by in the nuclear of histone isoform This was by and the kinase suggesting the involvement of InsP3R, and CaMKII. In the InsP3R similar of and activated CaMKII In this we have that InsP3R2 as a in with CaMKII from cardiac nuclei as well as InsP3R2 and CaMKIIδ expressed in COS-1 cells. localization the of CaMKII interaction that the amino-terminal 1078 are sufficient for the Additionally, the of CaMKII to the InsP3R2 by phosphorylation was We that from isolated cardiac nuclei 32P upon in kinase and that expressed InsP3R2 an amino-terminal was phosphorylated by CaMKII. The phosphorylation of the receptor by CaMKII may represent a which activated CaMKII feedback the InsP3R2 to the calcium for events. Consistent with this are results that CaMKII the open probability of InsP3R2 channels upon phosphorylation in planar lipid These results are with that CaMKII is with InsP3-mediated calcium release in and G. T. 2001; PubMed Scopus Google Scholar, D. E. PubMed Scopus Google Scholar). In we have identified the NE as the primary site of InsP3R2 in cardiac myocytes and that the receptor a with CaMKII. This the two signaling that local InsP3-mediated calcium release can activate CaMKII events. results that InsP3R2 as a for phosphorylation and these in the of channel We J. of University of for the CaMKIIδ expression and of University of for the of the