Targeted Inhibition of Ca2+/Calmodulin-dependent Protein Kinase II in Cardiac Longitudinal Sarcoplasmic Reticulum Results in Decreased Phospholamban Phosphorylation at Threonine 17

To investigate the role of Ca2+/calmodulin-dependent kinase II in cardiac sarcoplasmic reticulum function, transgenic mice were designed and generated to target the expression of a Ca2+/calmodulin-dependent kinase II inhibitory peptide in cardiac longitudinal sarcoplasmic reticulum using a truncated phospholamban transmembrane domain. The expressed inhibitory peptide was highly concentrated in cardiac sarcoplasmic reticulum. This resulted in a 59.7 and 73.6% decrease in phospholamban phosphorylation at threonine 17 under basal and β-adrenergic stimulated conditions without changing phospholamban phosphorylation at serine 16. Sarcoplasmic reticulum Ca2+ uptake assays showed that the Vmax was decreased by ∼30% although the apparent affinity for Ca2+ was unchanged in heterozygous hearts. The in vivo measurement of cardiac function showed no significant reductions in positive and negative dP/dt, but a moderate 18% decrease in dP/dt40, indicative of isovolumic contractility, and a 26.1% increase in the time constant of relaxation (τ) under basal conditions. The changes in these parameters indicate a moderate cardiac dysfunction in transgenic mice. Although the 3and 4-month-old transgenic mice displayed no overt signs of cardiac disease, when stressed by gestation and parturition, the 7-month-old female mice develop dilated heart failure, suggesting the important role of Ca2+/calmodulin-dependent kinase II pathway in the development of cardiac disease. To investigate the role of Ca2+/calmodulin-dependent kinase II in cardiac sarcoplasmic reticulum function, transgenic mice were designed and generated to target the expression of a Ca2+/calmodulin-dependent kinase II inhibitory peptide in cardiac longitudinal sarcoplasmic reticulum using a truncated phospholamban transmembrane domain. The expressed inhibitory peptide was highly concentrated in cardiac sarcoplasmic reticulum. This resulted in a 59.7 and 73.6% decrease in phospholamban phosphorylation at threonine 17 under basal and β-adrenergic stimulated conditions without changing phospholamban phosphorylation at serine 16. Sarcoplasmic reticulum Ca2+ uptake assays showed that the Vmax was decreased by ∼30% although the apparent affinity for Ca2+ was unchanged in heterozygous hearts. The in vivo measurement of cardiac function showed no significant reductions in positive and negative dP/dt, but a moderate 18% decrease in dP/dt40, indicative of isovolumic contractility, and a 26.1% increase in the time constant of relaxation (τ) under basal conditions. The changes in these parameters indicate a moderate cardiac dysfunction in transgenic mice. Although the 3and 4-month-old transgenic mice displayed no overt signs of cardiac disease, when stressed by gestation and parturition, the 7-month-old female mice develop dilated heart failure, suggesting the important role of Ca2+/calmodulin-dependent kinase II pathway in the development of cardiac disease. Calcium plays a central role in cardiac excitation-contraction coupling. The sarcoplasmic reticulum (SR) 1The abbreviations used are: SR, sarcoplasmic reticulum; CaMKII, Ca2+/calmodulin-dependent kinase II; SERCA, SR Ca2+-ATPase; TG, transgenic; NTG, non-transgenic; PLB, phospholamban; PKA, cAMP-dependent protein kinase; AIP, autocamitide inhibitory peptide; α-MHC, α-myosin heavy chain; RyR, ryanodine receptor; LV, left ventricle; LVEDP, LV end-diastolic pressure; PP, protein phosphatases. 1The abbreviations used are: SR, sarcoplasmic reticulum; CaMKII, Ca2+/calmodulin-dependent kinase II; SERCA, SR Ca2+-ATPase; TG, transgenic; NTG, non-transgenic; PLB, phospholamban; PKA, cAMP-dependent protein kinase; AIP, autocamitide inhibitory peptide; α-MHC, α-myosin heavy chain; RyR, ryanodine receptor; LV, left ventricle; LVEDP, LV end-diastolic pressure; PP, protein phosphatases. releases Ca2+ to trigger contraction and uptakes Ca2+ to initiate relaxation. Ca2+-induced Ca2+ release occurs via ryanodine-sensitive SR Ca2+ release channels located mainly at junctional SR (“foot” structure), whereas Ca2+ uptake is mediated principally by the SR Ca2+-ATPase (SERCA) which is located in the longitudinal SR. Both Ca2+ release and uptake are proposed to be regulated in a Ca2+-dependent manner via Ca2+/calmodulin-dependent protein kinase II (CaMKII) (1Maier L.S. Bers D.M. J. Mol. Cell. Cardiol. 2002; 34: 919-939Google Scholar). CaMKII is a member of a family of Ca2+/calmodulin-regulated enzymes. Four CaMKII isoforms are derived from four closely related genes, α, β, γ, and δ (2Schuman H. Curr. Opin. Cell Biol. 1993; 5: 247-253Google Scholar). The δ and γ are the primary cardiac CaMKII isoforms expressed in the adult heart (3Tobimatsu T. Fujisawa H. J. Biol. Chem. 1989; 264: 17907-17912Google Scholar, 4Mayer P. Mohlig M. Idlibe D. Pfeiffer A. Basic Res. Cardiol. 1995; 90: 372-379Google Scholar, 5Hagemann D. Hoch B. Krause E.G. Karczewski P. J. Cell. Biochem. 1999; 74: 202-210Google Scholar). CaMKII distributes in distinct compartments of the cardiomyocytes including sarcolemma, cytosol, SR, and nucleus (6Braun A.P. Schulman H. Annu. Rev. Physiol. 1995; 57: 417-445Google Scholar), which may represent its functional relevance in the heart. The link between specific isoforms of CaMKII with particular regulatory properties, intracellular localization, and cellular substrates is not established. The activity of SERCA is primarily regulated by an SR intrinsic protein, phospholamban (PLB). PLB physically interacts with SERCA to inhibit pump activity (7Tada M. Kirchberger M.A. Repke D.L. Katz A.M. J. Biol. Chem. 1974; 249: 6174-6180Google Scholar). The phosphorylation of PLB disrupts the interaction of PLB with the SERCA pump, relieving its inhibitory effect and resulting in an increase in the apparent affinity of SERCA for Ca2+ (7Tada M. Kirchberger M.A. Repke D.L. Katz A.M. J. Biol. Chem. 1974; 249: 6174-6180Google Scholar). cAMP-dependent protein kinase (PKA) mediates PLB phosphorylation at serine 16 (7Tada M. Kirchberger M.A. Repke D.L. Katz A.M. J. Biol. Chem. 1974; 249: 6174-6180Google Scholar, 8Simmerman H.K.B. Collins J.H. Theibert J.L. Wegener A.D. Jones L.R. J. Biol. Chem. 1986; 258: 13587-13591Google Scholar), whereas CaMKII mediates phosphorylation of PLB at threonine 17 (8Simmerman H.K.B. Collins J.H. Theibert J.L. Wegener A.D. Jones L.R. J. Biol. Chem. 1986; 258: 13587-13591Google Scholar, 9Lepeuch C.J. Haiech J. Demaille J.G. Biochemistry. 1979; 18: 5150-5157Google Scholar). In the intact heart, both PLB serine 16 and threonine 17 are phosphorylated by PKA and CaMKII, respectively, in response to β-adrenergic stimulation (10Wegener A.D. Simmermann H.K.B. Lindemann J.P. Jones L.R. J. Biol. Chem. 1989; 264: 11469-11474Google Scholar, 11Talosi L. Edes I. Kranias E.G. Am. J. Physiol. 1993; 264: H791-H797Google Scholar). In vitro studies have indicated that SERCA is also phosphorylated by CaMKII at Ser38, which may enhance the maximal velocity (Vmax) of calcium uptake (12Hawkins C. Xu A. Narayanan N. J. Biol. Chem. 1994; 269: 31198-31206Google Scholar, 13Toyofuku T. Kurzydloski K. Narayanan N. MacLennan D.H. J. Biol. Chem. 1994; 269: 26492-26496Google Scholar, 14Xu A. Narayanan N. J. Biol. Chem. 2000; 275: 4407-4416Google Scholar). However, this pathway remains controversial (15Reddy L.G. Jones L.R. Pace R.C. Stockes D.L. J. Biol. Chem. 1996; 271: 14964-14970Google Scholar, 16Odermatt A. Kazimieez K. MacLennan D.H. J. Biol. Chem. 1996; 271: 14206-14213Google Scholar), due to a lack of in vivo evidence. In addition to Ca2+ uptake, CaMKII may also play a role in regulating Ca2+-induced Ca2+ release by phosphorylation of serine 2809 of the cardiac SR calcium release channel (ryanodine receptor, RyR) which can also be a target for PKA (17Witcher D.R. Kovacs R.J. Schulman H. Cefali D.C. Jones L.R. J. Biol. Chem. 1991; 266: 11144-11152Google Scholar, 18Hain J. Onoue H. Mayrleitner M. Fleischer S. Schindler H. J. Biol. Chem. 1995; 270: 2074-2081Google Scholar). However, whether the phosphorylation of RyR leads to opening or closing the Ca2+ release channel remains undefined (17Witcher D.R. Kovacs R.J. Schulman H. Cefali D.C. Jones L.R. J. Biol. Chem. 1991; 266: 11144-11152Google Scholar, 18Hain J. Onoue H. Mayrleitner M. Fleischer S. Schindler H. J. Biol. Chem. 1995; 270: 2074-2081Google Scholar, 19Lokuta A.J. Roger T.B. Lederer W.J. Valdivia H.H. J. Physiol. (Lond.). 1995; 487: 609-622Google Scholar). Therefore, the role of CaMKII in the regulation of RyR in intact heart is not clear. Recently, several studies (20Hoch B. Meyer R. Hetzer R. Krause E.G. Karczewski P. Circ. Res. 1999; 84: 713-721Google Scholar) have shown that the level of cardiac isoform CaMKII δ3 is significantly increased in dilated cardiomyopathy patients. Currie (21Currie S. Smith G.L. FEBS Lett. 1999; 459: 244-248Google Scholar) and Kirchhefer et al. (22Kirchhefer U. Schmitz W. Scholz H. Neumann J. Cardiovasc. Res. 1999; 42: 254-261Google Scholar) have independently reported that the activity of CaMKII is significantly increased in hypertrophied animal models as well as in human heart failure. However, Netticadan et al. (23Netticadan T. Temsah R. Osada M. Dhallar N.S. Am. J. Physiol. 1999; 277: C384-C391Google Scholar, 24Netticadan T. Temsah R. Kawabata K. Dhalla N.S. Circ. Res. 2000; 86: 596-605Google Scholar) showed that the endogenous SR-associated CaMKII-mediated phosphorylation of SR Ca2+-handling proteins is depressed in heart failure due to either ischemia-reperfusion or myocardial infarction. These data, taken together, suggest that SR CaMKII is associated with the abnormal Ca2+ handling of the SR in cardiomyocytes. However, the role of CaMKII in the regulation of SR Ca2+-handling proteins under pathophysiological conditions is not clear. In order to investigate the role of SR-associated CaMKII in the regulation of SR Ca2+ transport, as well as in regulating cardiac function, transgenic mice were generated to target specifically the expression of CaMKII inhibitory peptide to cardiac longitudinal SR. The SR targeting sequence is defined by the transmembrane portion of the PLB protein, encoding a double mutant that obviates PLB function. We demonstrate that targeted inhibition of SR CaMKII activity results in a significant decrease in PLB phosphorylation at threonine 17. When stressed, the transgenic mice develop dilated heart failure, suggesting the important role of CaMKII pathway in the development of cardiac disease. Generation of CaMKII-AIP4Transgenic Mice—A synthetic gene expression unit was engineered to encode three functional using by the at the of at the the expression unit that encode a CaMKII a and an SR The CaMKII of encoding a of the CaMKII autocamitide inhibitory peptide this sequence is to be a highly specific and of CaMKII A. I. S. T. Fujisawa H. Biochem. Res. 1995; Scholar). encoding a were of the In a synthetic gene encoding a truncated PLB transmembrane was to as SR with at and studies have shown that and in of the PLB inhibition to SERCA activity K. M. MacLennan D.H. J. Biol. Chem. Scholar). This expression unit was a between the α-myosin heavy from J. and an The were from and were by The was of by the at of The were for the of the by using an and an The mice were by mice were with mice. from at the were used for with as were in with the for the and of and by the and was to the to the of of of of was to the that as of from was with and The were with the to of the The of the were using a and were with the to of and as well as were as with a M. Biochem. 1999; 269: Scholar). In an heart was used for of cardiac with of and SR were were at for the was and the resulting was in and as The from were with as that was by to the of and at for The was concentrated and was as a The resulting was in and at for an The was used as an SR was by used for were as and and and The or were from was using the protein was used as The were using were taken from and mice. was to the a and the were in were as Xu M. A. M. Scholar). The expression was using by The was used to protein using from in by The were by a of of PLB and the basal phosphorylation level of PLB and RyR, a PLB peptide phosphorylated at or at and phosphorylated were The were and using with to and of the cardiac were and the were to the Xu D.L. N. M. J. Biol. Chem. 2000; 275: Scholar). In vitro phosphorylation was using cardiac from heart, as W. Kranias E.G. J. Biol. Chem. Scholar). endogenous CaMKII of the cardiac was to of and protein kinase peptide were The was at PKA phosphorylation of the cardiac was in the with of the PKA were with of a and (PKA) which was associated with in of protein was to by the of PLB and PLB using the phosphorylated PLB as In vivo phosphorylation was by with from to or by Ca2+ via left The were with and of cardiac protein was used for the of PLB and PLB as To phosphorylated RyR, of protein was to by with the the was used to with to the of the protein the Ca2+ uptake SR in cardiac was by the an T. Jones L.R. M. Am. J. Physiol. 1999; Scholar). were in uptake of to Ca2+ as by the S. A. in Scholar). The was by the addition of The of Ca2+ uptake was by of uptake at and were by SR SR protein activity was with the from The was by SR or proteins as to with and without the synthetic in a of for The was by the addition of of The was at to the The activity was by the in the of the from in its T. Temsah R. Kawabata K. Dhalla N.S. Circ. Res. 2000; 86: 596-605Google Scholar). function was as J. Am. J. Physiol. Scholar). were with and and a the and were with The was to a for measurement of and the was to a pump for the of was the and the and the left of the were to for to of as a constant a were were to to for between of the was at a of and of heart function were To left function, the and were the maximal positive of LV an of myocardial the of LV increase at an LV of an indicative of isovolumic Bers D.M. J. 2000; Scholar, W. S. M. R.J. 1995; the time constant of relaxation a of LV from the of the LV to a end-diastolic an of LV and C. A. J. C. I. H. P. J. Scholar). were and using a and were for at the of and are reported as the The was using or or of cardiac function of was and of CaMKII-AIP4Transgenic a peptide is an for AIP, the sequence is a and specific for CaMKII A. I. S. T. Fujisawa H. Biochem. Res. 1995; Scholar). In this the cardiac specific was used to the to the heart. In order to target the expression of to the longitudinal SR, a truncated PLB transmembrane was used as SR To the inhibition of PLB transmembrane SERCA were at and K. M. MacLennan D.H. J. Biol. Chem. Scholar). the was to the targeted expression of in cardiac SR. The for the is shown in mice were generated as under transgenic were by and of transgenic three and the to and not the to and to an of and To the protein cardiac were to by using shown in the protein is expressed in hearts. of the protein that the expression level in is in was for the functional To the of expression cardiac SR, was The from using both and showed the of This indicated that the protein as its SR and highly concentrated in the SR. by using SR and showed the was expressed in cardiac SR in transgenic mice To whether expression CaMKII expression in cardiac SR, SR proteins were and with is no significant in CaMKII protein level between and mice in in TG, and protein in SR and from and CaMKII protein were in cardiac SR, and from and mice by this δ were which may represent δ and γ the are as the for The mice were of that no overt of a was no of cardiac in the mice. The heart of the mice used for the of function, PLB and SR Ca2+ uptake was for and for in of in of is regulated by phosphorylation of by CaMKII and of by PKA (7Tada M. Kirchberger M.A. Repke D.L. Katz A.M. J. Biol. Chem. 1974; 249: 6174-6180Google Scholar, 8Simmerman H.K.B. Collins J.H. Theibert J.L. Wegener A.D. Jones L.R. J. Biol. Chem. 1986; 258: 13587-13591Google Scholar, 9Lepeuch C.J. Haiech J. Demaille J.G. Biochemistry. 1979; 18: 5150-5157Google Scholar). To the effect of expression of PLB the PLB phosphorylation under either basal or stimulated conditions were shown in the basal of both and of PLB were significantly decreased by in from in in TG, However, PLB in mice was not significantly in in TG, results indicate that functional expression of is by the CaMKII the PLB phosphorylation under in vitro stimulated conditions. from and were with in the of Ca2+ under conditions and to by using shown in in the and of and PLB, were decreased by in in TG, and in in TG, respectively, in with the endogenous CaMKII activity in is the phosphorylation of PLB was not significantly as by the of PLB in the of PKA between by and by This that the PKA pathway is not by expression SR. reported that β-adrenergic stimulation was associated with significant PLB phosphorylation at both and To the effect of β-adrenergic stimulation phosphorylation of PLB, the and were with from to were to the PLB phosphorylation with to phosphorylated and The maximal phosphorylation level of the PKA in response to β-adrenergic stimulation as by PLB in mice However, the level of PLB was decreased by 73.6% in mice β-adrenergic stimulation maximal PLB phosphorylation of the CaMKII not be in the These results demonstrate that due to expression of the SR CaMKII pathway is significantly resulting in a significant decrease in PLB phosphorylation at reported that Ca2+ Ca2+ channels the of CaMKII pathway in cardiomyocytes H. Lederer W.J. T. E.G. U. S. A. 1994; Scholar). To the stimulation of CaMKII pathway by Ca2+ was in vivo the cardiac function from the response to β-adrenergic The maximal level of PLB was decreased by in in TG, in mice of the using not the of the PLB protein in mice not but of for is well that SR Ca2+ uptake is regulated by PLB phosphorylation disrupts the inhibitory of PLB with SERCA, a in SERCA affinity for several studies have shown that SERCA can be phosphorylated by CaMKII resulting in an increase in the Vmax of Ca2+ uptake (12Hawkins C. Xu A. Narayanan N. J. Biol. Chem. 1994; 269: 31198-31206Google Scholar, 13Toyofuku T. Kurzydloski K. Narayanan N. MacLennan D.H. J. Biol. Chem. 1994; 269: 26492-26496Google Scholar, 14Xu A. Narayanan N. J. Biol. Chem. 2000; 275: 4407-4416Google Scholar). To whether the decreased phosphorylation of PLB due to inhibition of SR CaMKII SR Ca2+ uptake, SR Ca2+ uptake was using cardiac shown in and the Vmax of Ca2+ uptake in was decreased by ∼30% in NTG, for TG, However, the apparent affinity for as was in in TG, results indicate that decreased PLB phosphorylation at not SERCA affinity for suggesting that the intact PKA pathway is for the PLB regulation of SERCA under the conditions. However, an ∼30% decrease in the Vmax of SR Ca2+ uptake function was in mice. is that the decreased Vmax of SR Ca2+ uptake may be due to the changes of SERCA pump To this was to the protein level using both cardiac and SR. shown in showed that protein was not significantly in mice in with mice in in TG, for and in in TG, for These results suggest a inhibition of the Vmax of SR Ca2+ uptake by expression in cardiac SR. of of of the can be phosphorylated at serine 2809 by either PKA or CaMKII in cardiac SR (17Witcher D.R. Kovacs R.J. Schulman H. Cefali D.C. Jones L.R. J. Biol. Chem. 1991; 266: 11144-11152Google Scholar, 18Hain J. Onoue H. Mayrleitner M. Fleischer S. Schindler H. J. Biol. Chem. 1995; 270: 2074-2081Google Scholar). To whether the expression the phosphorylation level of RyR in junctional SR, was used to the phosphorylated RyR under basal conditions as To the of protein as RyR, the was and with shown in the basal phosphorylation level of RyR was decreased by for and for TG, in mice when by RyR protein suggest that RyR phosphorylation was not significantly by expression in SR, the of SR CaMKII pathway to longitudinal SR. SR in are between protein and protein in cellular Therefore, the decrease in SR CaMKII activity resulting from expression in mice the SR protein and protein level as well as protein activity were in SR from and hearts. shown in and protein were not significantly in mice the protein protein and activity in and protein protein in a in CaMKII both basal and stimulated heart was significantly in mice with and the maximal heart response to β-adrenergic stimulation was in the to be increased in the mice with but the was not significant not However, as shown in LV was in the with mice and in response to LV increased in the but not in mice Although LV was not significantly between the of as shown in the dP/dt40, an of that to for the in a in LV function in the mice. shown in was by the and the of the response to was in the mice as indicated by the time constant of was significantly by 26.1% in at and the of β-adrenergic stimulation was significantly in the with mice under conditions but was no β-adrenergic stimulation not of Ca2+ to Ca2+ which is to CaMKII activity of showed a of as in of SR the to the mice not of cardiac 7-month-old female mice that at three cardiac and dilated heart failure. shown in the and the in mice were increased by respectively, in with mice. In the mice a of heart failure. studies showed a of the left as well as left showed that were in the of the left with and of the These displayed with in the left in female and mice to as with as with as with as with as with in a The functional relevance of of CaMKII in cardiomyocytes is not clear. In this the and SR to target the expression of a CaMKII inhibitory cardiac longitudinal SR. This to the function of CaMKII in the hearts. of CaMKII, and cellular function by CaMKII in of the are used to specifically a CaMKII but the the The of inhibitory to the role of CaMKII is that specific expression can be by the gene and intracellular can be the of targeting This was to inhibitory peptide in in J. B. M.A. Am. J. Physiol. 1996; 271: Scholar). In the a targeted expression of cardiac SR. a decrease in basal phosphorylation of PLB threonine 17 in longitudinal SR, but a decrease in the phosphorylation of RyR serine 2809 in junctional SR, that the expressed is mainly functional at longitudinal SR. The 3and 4-month-old displayed a moderate decrease in both heart and the maximal of contraction under both basal and stimulated conditions and was a significant increase in the relaxation time constant (τ) and an in basal LVEDP, a in myocardial function. these to be in the These results indicate the contraction and relaxation function of was due to targeted inhibition of CaMKII in longitudinal SR. the the response to β-adrenergic stimulation was in suggesting that PLB phosphorylation plays an important role in the regulation of heart function under β-adrenergic stimulated conditions in intact hearts. the decrease at PLB results the that the effect of PLB phosphorylation is not in the regulation of cardiac function under conditions (7Tada M. Kirchberger M.A. Repke D.L. Katz A.M. J. Biol. Chem. 1974; 249: 6174-6180Google Scholar, W. J. Kranias E.G. J. Biol. Chem. 2000; 275: Scholar). by et al. W. J. Kranias E.G. J. Biol. Chem. 2000; 275: Scholar), using PLB mutant mice under PLB which the PLB phosphorylation by CaMKII, that PLB phosphorylation of is for regulating SERCA activity and the maximal cardiac to β-adrenergic stimulation in cardiomyocytes. the of which is to Ca2+ CaMKII activity of showed a of the response and a decrease in PLB phosphorylation in the mice. This the important role of SR CaMKII pathway in the cardiac response to Ca2+ Although the 4-month-old mice not develop overt cardiac both mice and 7-month-old female mice showed a of dilated heart failure, by a of the left and a the and were increased by the significant decrease in cardiac in mice. The cardiac in mice that the of the SR CaMKII pathway leads to cardiac and heart failure under The of the development of heart failure in mice is is that inhibition of SR CaMKII results in of the SR Ca2+ handling function, which shown to be closely related to the development of cardiac and heart failure J.P. N. J. 1991; Scholar, M. H. M. Circ. Res. 1994; 74: Scholar). In this in addition to the significant decrease at PLB an ∼30% decrease in the Vmax of SR Ca2+ uptake without changing the apparent affinity for Ca2+ in mice. The decreased Vmax of SR Ca2+ uptake be due to the inhibition of SERCA activity by targeted expression of CaMKII inhibitory peptide in cardiac SR, is no decrease in SERCA protein level in mice. the unchanged apparent affinity for Ca2+ that the decreased SR Ca2+ uptake may not be due to the expression of the mutant PLB transmembrane which shown to its inhibitory effect SERCA activity by double K. M. MacLennan D.H. J. Biol. Chem. Scholar). studies also of regulation of SERCA activity by CaMKII A. Bers D.M. Am. J. Physiol. 1995; Scholar, L. Kranias E.G. Bers D.M. Am. J. Physiol. Scholar). However, to be reported that transgenic mice CaMKII in cardiac and decreased function T. T.B. Schulman H. J. J.H. J. Biol. Chem. 2002; 277: Scholar). In decreased PLB phosphorylation at both and as well as SR Ca2+ uptake were and the was to be associated with the increase in activity T. T.B. Schulman H. J. J.H. J. Biol. Chem. 2002; 277: Scholar). In showed that targeted inhibition of SR CaMKII activity leads to a decrease in PLB phosphorylation at without changing the is associated with the inhibition of SR CaMKII In the functional relevance of SR CaMKII in the regulation of SR Ca2+ handling under and conditions to the The transgenic mice be the in which SR CaMKII activity is specifically as a of targeted expression of CaMKII inhibitory peptide in the longitudinal SR. This a to the role of SR CaMKII in the regulation of excitation-contraction and in the development of heart failure. We L. for with measurement and for with and of transgenic