Overexpression of the Sarcolemmal Calcium Pump in the Myocardium of Transgenic Rats

From the Department of Medicine, University of Würzburg (A.H., S.O.-M., T.R., K.N., K.S., C.B., L.N.); Department of Physiology, University of Bonn (F.G., K.W.L., R.M.); Department of Medicine, Cardiology and Angiology, Medical Faculty, Mannheim, University of Heidelberg (K.H., H.H., P.G., G.E.); Max Delbrück Center for Molecular Medicine, Berlin (S.H., U.G., R.V.); and the Institute of Physiology, University of Leipzig (H.G.Z.), Germany. Present affiliation for A.H. is Developmental Genetics, Max Delbrueck Center for Molecular Medicine, Berlin, Germany. Present affiliation for R.V. is Institute of Clinical Pharmacology and Toxicology, Benjamin Franklin Medical Center, Berlin, Germany. Present affiliation for C.B. is Department of Anesthesiology, University of Würzburg, Germany.

Correspondence to Ludwig Neyses, MD, Department of Medicine, University of Würzburg, Josef-Schneider-Str 2, D-97080 Würzburg, Germany.

Abstract—The plasma membrane calmodulin–dependent calcium ATPase (PMCA) is a calcium-extruding enzyme controlling Ca²⁺ homeostasis in nonexcitable cells. However, its function in the myocardium is unclear because of the presence of the Na⁺/Ca²⁺ exchanger. We approached the question of the physiological function of the calcium pump using a transgenic "gain of function" model. Transgenic rat lines carrying the human PMCA 4 cDNA under control of the ventricle-specific myosin light chain-2 promoter were established, and expression in the myocardium was ascertained at the mRNA, protein, and functional levels. In vivo hemodynamic measurements in adult homozygous animals showed no differences in baseline and increased cardiac performance recruited by volume overload compared with controls. No differences between transgenic and control cardiomyocytes were found in patch clamp voltage dependence, activation/inactivation behavior of the L-type Ca²⁺ current, or fast [Ca²⁺]_i transients (assessed by the Fura-2 method). To test whether the PMCA might be involved in processes other than beat-to-beat regulation of contraction/relaxation, we compared growth processes of neonatal transgenic and control cardiomyocytes. A 1.6- and 2.3-fold higher synthesis rate of total protein was seen in cells from transgenic animals compared with controls on incubation with 2% FCS for 24 hours and 36 hours, respectively. An effect of similar magnitude was observed using 20 µmol/L phenylephrine. A 1.4-fold– and 2.0-fold–higher protein synthesis peak was seen in PMCA-overexpressing cardiomyocytes after stimulation with isoproterenol for 12 hours and 24 hours, respectively. Because pivotal parts of the - and ß-adrenergic signal transduction pathways recently have been localized to caveolae, we tested the hypothesis that the PMCA might alter the amplitude of - and ß-adrenergic growth signals by virtue of its localization in caveolae. Biochemical as well as immunocytochemical studies suggested that the PMCA in large part was colocalized with caveolin 3 in caveolae of cardiomyocytes. These results indicate that the sarcolemmal Ca²⁺-pump has little relevance for beat-to-beat regulation of contraction/relaxation in adult animals but likely plays a role in regulating myocardial growth, possibly through modulation of caveolar signal transduction.

Key Words: plasma membrane Ca²⁺-ATPase • myocardium • transgenic rat • contraction • cardiac growth

This article has been cited by other articles:

				T. M. A. Mohamed, D. Oceandy, S. Prehar, N. Alatwi, Z. Hegab, F. M. Baudoin, A. Pickard, A. O. Zaki, R. Nadif, E. J. Cartwright, et al. Specific Role of Neuronal Nitric-oxide Synthase when Tethered to the Plasma Membrane Calcium Pump in Regulating the {beta}-Adrenergic Signal in the Myocardium J. Biol. Chem., May 1, 2009; 284(18): 12091 - 12098. [Abstract] [Full Text] [PDF]

				D. Oceandy, E. J. Cartwright, M. Emerson, S. Prehar, F. M. Baudoin, M. Zi, N. Alatwi, L. Venetucci, K. Schuh, J. C. Williams, et al. Neuronal Nitric Oxide Synthase Signaling in the Heart Is Regulated by the Sarcolemmal Calcium Pump 4b Circulation, January 30, 2007; 115(4): 483 - 492. [Abstract] [Full Text] [PDF]

				J. C. Williams, A. L. Armesilla, T. M. A. Mohamed, C. L. Hagarty, F. H. McIntyre, S. Schomburg, A. O. Zaki, D. Oceandy, E. J. Cartwright, M. H. Buch, et al. The Sarcolemmal Calcium Pump, {alpha}-1 Syntrophin, and Neuronal Nitric-oxide Synthase Are Parts of a Macromolecular Protein Complex J. Biol. Chem., August 18, 2006; 281(33): 23341 - 23348. [Abstract] [Full Text] [PDF]

				R. C. Balijepalli, J. D. Foell, D. D. Hall, J. W. Hell, and T. J. Kamp From the Cover: Localization of cardiac L-type Ca2+ channels to a caveolar macromolecular signaling complex is required for beta2-adrenergic regulation PNAS, May 9, 2006; 103(19): 7500 - 7505. [Abstract] [Full Text] [PDF]

				N. Burkard, J. Becher, C. Heindl, L. Neyses, K. Schuh, and O. Ritter Targeted Proteolysis Sustains Calcineurin Activation Circulation, March 1, 2005; 111(8): 1045 - 1053. [Abstract] [Full Text] [PDF]

				D. Wernicke, C. Thiel, C. M. Duja-Isac, K. V. Essin, M. Spindler, D. J. R. Nunez, R. Plehm, N. Wessel, A. Hammes, R.-J. Edwards, et al. {alpha}-Tropomyosin mutations Asp175Asn and Glu180Gly affect cardiac function in transgenic rats in different ways Am J Physiol Regulatory Integrative Comp Physiol, September 1, 2004; 287(3): R685 - R695. [Abstract] [Full Text] [PDF]

				A. L. Armesilla, J. C. Williams, M. H. Buch, A. Pickard, M. Emerson, E. J. Cartwright, D. Oceandy, M. D. Vos, S. Gillies, G. J. Clark, et al. Novel Functional Interaction between the Plasma Membrane Ca2+ Pump 4b and the Proapoptotic Tumor Suppressor Ras-associated Factor 1 (RASSF1) J. Biol. Chem., July 23, 2004; 279(30): 31318 - 31328. [Abstract] [Full Text] [PDF]

				M. Brede, W. Roell, O. Ritter, F. Wiesmann, R. Jahns, A. Haase, B. K. Fleischmann, and L. Hein Cardiac Hypertrophy Is Associated With Decreased eNOS Expression in Angiotensin AT2 Receptor-Deficient Mice Hypertension, December 1, 2003; 42(6): 1177 - 1182. [Abstract] [Full Text] [PDF]

				K. Schuh, T. Quaschning, S. Knauer, K. Hu, S. Kocak, N. Roethlein, and L. Neyses Regulation of Vascular Tone in Animals Overexpressing the Sarcolemmal Calcium Pump J. Biol. Chem., October 17, 2003; 278(42): 41246 - 41252. [Abstract] [Full Text] [PDF]

				R. Gros, T. Afroze, X.-M. You, G. Kabir, R. Van Wert, W. Kalair, A. E. Hoque, I. N. Mungrue, and M. Husain Plasma Membrane Calcium ATPase Overexpression in Arterial Smooth Muscle Increases Vasomotor Responsiveness and Blood Pressure Circ. Res., October 3, 2003; 93(7): 614 - 621. [Abstract] [Full Text] [PDF]

				J. BORLAK and T. THUM Hallmarks of ion channel gene expression in end-stage heart failure FASEB J, September 1, 2003; 17(12): 1592 - 1608. [Abstract] [Full Text] [PDF]

				K. Schuh, S. Uldrijan, S. Gambaryan, N. Roethlein, and L. Neyses Interaction of the Plasma Membrane Ca2+ Pump 4b/CI with the Ca2+/Calmodulin-dependent Membrane-associated Kinase CASK J. Biol. Chem., March 7, 2003; 278(11): 9778 - 9783. [Abstract] [Full Text] [PDF]

				P. Ratajczak, T. Damy, C. Heymes, P. Oliviero, F. Marotte, E. Robidel, R. Sercombe, J. Boczkowski, L. Rappaport, and J.-L. Samuel Caveolin-1 and -3 dissociations from caveolae to cytosol in the heart during aging and after myocardial infarction in rat Cardiovasc Res, February 1, 2003; 57(2): 358 - 369. [Abstract] [Full Text] [PDF]

				O. Ritter, S. Hack, K. Schuh, N. Rothlein, A. Perrot, K. J. Osterziel, H. D. Schulte, and L. Neyses Calcineurin in Human Heart Hypertrophy Circulation, May 14, 2002; 105(19): 2265 - 2269. [Abstract] [Full Text] [PDF]

				J. Piuhola, A. Hammes, K. Schuh, L. Neyses, O. Vuolteenaho, and H. Ruskoaho Overexpression of sarcolemmal calcium pump attenuates induction of cardiac gene expression in response to ET-1 Am J Physiol Regulatory Integrative Comp Physiol, September 1, 2001; 281(3): R699 - R705. [Abstract] [Full Text] [PDF]

				J. N. Muth, I. Bodi, W. Lewis, G. Varadi, and A. Schwartz A Ca2+-Dependent Transgenic Model of Cardiac Hypertrophy : A Role for Protein Kinase C{{alpha}} Circulation, January 2, 2001; 103(1): 140 - 147. [Abstract] [Full Text] [PDF]

				E. E. Strehler and D. A. Zacharias Role of Alternative Splicing in Generating Isoform Diversity Among Plasma Membrane Calcium Pumps Physiol Rev, January 1, 2001; 81(1): 21 - 50. [Abstract] [Full Text] [PDF]

				Y. Hattori, N. Matsuda, J. Kimura, T. Ishitani, A. Tamada, S. Gando, O. Kemmotsu, and M. Kanno Diminished function and expression of the cardiac Na+-Ca2+ exchanger in diabetic rats: implication in Ca2+ overload J. Physiol., August 15, 2000; 527(1): 85 - 94. [Abstract] [Full Text] [PDF]

				V. O. Rybin, X. Xu, and S. F. Steinberg Activated Protein Kinase C Isoforms Target to Cardiomyocyte Caveolae : Stimulation of Local Protein Phosphorylation Circ. Res., May 14, 1999; 84(9): 980 - 988. [Abstract] [Full Text] [PDF]

				G. Hasenfuss Calcium Pump Overexpression and Myocardial Function : Implications for Gene Therapy of Myocardial Failure Circ. Res., November 2, 1998; 83(9): 966 - 968. [Full Text] [PDF]

				T. de Jager, T. Pelzer, S. Muller-Botz, A. Imam, J. Muck, and L. Neyses Mechanisms of Estrogen Receptor Action in the Myocardium. RAPID GENE ACTIVATION VIA THE ERK1/2 PATHWAY AND SERUM RESPONSE ELEMENTS J. Biol. Chem., July 20, 2001; 276(30): 27873 - 27880. [Abstract] [Full Text] [PDF]

				K. Schuh, S. Uldrijan, M. Telkamp, N. Rothlein, and L. Neyses The plasmamembrane calmodulin-dependent calcium pump: a major regulator of nitric oxide synthase I J. Cell Biol., October 15, 2001; 155(2): 201 - 206. [Abstract] [Full Text] [PDF]