© 1995 American Heart Association, Inc.
Articles |
Arachidonic Acid–Dependent Phosphorylation of Troponin I and Myosin Light Chain 2 in Cardiac Myocytes
From the Center for Anesthesiology Research (D.S.D., A.D., W.S., C.S.M.) and the Department of Molecular Cardiology (L.A.M., M.B.), Research Institute, The Cleveland Clinic Foundation, Cleveland, Ohio.
Correspondence to Meredith Bond, PhD, Department of Molecular Cardiology/FF10, The Cleveland Clinic Foundation, 9500 Euclid Ave, Cleveland, OH 44195.
Abstract Recent evidence has suggested that arachidonic acid (AA) may be an important signaling molecule in cardiac excitation-contraction coupling. We previously showed that AA and endothelin-1 (ET) inhibit distinct K+ channels via protein kinase C–dependent pathways in rat ventricular myocytes. In addition, we demonstrated that Ca2+ transients in populations of fura 2–loaded myocytes were potentiated by AA and ET via activation of protein kinase C. In this study, we have used suspensions of [32P]orthophosphate (32Pi)–labeled rat ventricular myocytes to study the effects of AA and ET at the level of the myofilaments. After a 10-minute incubation of the labeled cells with phorbol 12-myristate 13-acetate (PMA), AA, or ET in the presence or absence of the protein kinase C inhibitor calphostin C, the myofibrillar proteins were separated by PAGE. Measurement of unloaded cell shortening using video edge detection in single electrically stimulated myocytes was also used to assess the effects of AA and ET on myocyte contractility. Incubation with either PMA, AA, or ET resulted in similar increases in 32Pi incorporation into troponin I (TnI) and myosin light chain 2 (MLC2), which was inhibited by preincubation with the protein kinase C antagonist calphostin C. In addition, the ability of these agonists to stimulate phosphorylation of TnI or MLC2 did not require extracellular Ca2+ or intact intracellular Ca2+ stores. The effects of AA and ET together on phosphorylation of TnI or MLC2 were not additive. These results show that AA and ET phosphorylate myofibrillar proteins in intact cardiac myocytes via a calphostin C–inhibitable Ca2+-independent pathway and that AA, like ET, stimulates a positive inotropic effect in cardiac myocytes. Thus, phosphorylation of cardiac myofilaments by AA and/or ET, in addition to phosphorylation-dependent inhibition of distinct K+ channels, may represent an important signaling pathway by which AA and ET influence the inotropic state of the heart.
Key Words: myocytes • arachidonic acid • endothelin-1 • phosphorylation • myofilaments
This article has been cited by other articles:
 |
|
 |
|
  A. Molnar, A. Borbely, D. Czuriga, S. M. Ivetta, S. Szilagyi, Z. Hertelendi, E. T. Pasztor, A. Balogh, Z. Galajda, T. Szerafin, et al. Protein Kinase C Contributes to the Maintenance of Contractile Force in Human Ventricular Cardiomyocytes J. Biol. Chem., January 9, 2009; 284(2): 1031 - 1039. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Davis, M. V. Westfall, D. Townsend, M. Blankinship, T. J. Herron, G. Guerrero-Serna, W. Wang, E. Devaney, and J. M. Metzger Designing Heart Performance by Gene Transfer Physiol Rev, October 1, 2008; 88(4): 1567 - 1651. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. Hlavackova, J. Neckar, J. Jezkova, P. Balkova, B. Stankova, O. Novakova, F. Kolar, and F. Novak Dietary Polyunsaturated Fatty Acids Alter Myocardial Protein Kinase C Expression and Affect Cardioprotection Induced by Chronic Hypoxia Experimental Biology and Medicine, June 1, 2007; 232(6): 823 - 832. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. V. Westfall, A. M. Lee, and D. A. Robinson Differential Contribution of Troponin I Phosphorylation Sites to the Endothelin-modulated Contractile Response J. Biol. Chem., December 16, 2005; 280(50): 41324 - 41331. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. D Russell and P. Molenaar Investigation of signaling pathways that mediate the inotropic effect of urotensin-II in human heart Cardiovasc Res, September 1, 2004; 63(4): 673 - 681. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. H. Goldspink, D. E. Montgomery, L. A. Walker, D. Urboniene, R. D. McKinney, D. L. Geenen, R. J. Solaro, and P. M. Buttrick Protein Kinase C{epsilon} Overexpression Alters Myofilament Properties and Composition During the Progression of Heart Failure Circ. Res., August 20, 2004; 95(4): 424 - 432. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. M. Metzger and M. V. Westfall Covalent and Noncovalent Modification of Thin Filament Action: The Essential Role of Troponin in Cardiac Muscle Regulation Circ. Res., February 6, 2004; 94(2): 146 - 158. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 H. Kan, Z. Xie, and M. S. Finkel p38 MAP kinase-mediated negative inotropic effect of HIV gp120 on cardiac myocytes Am J Physiol Cell Physiol, January 1, 2004; 286(1): C1 - C7. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. Pi, D. Zhang, K. R Kemnitz, H. Wang, and J. W Walker Protein kinase C and A sites on troponin I regulate myofilament Ca2+ sensitivity and ATPase activity in the mouse myocardium J. Physiol., November 1, 2003; 552(3): 845 - 857. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. V. Westfall and A. R. Borton Role of Troponin I Phosphorylation in Protein Kinase C-mediated Enhanced Contractile Performance of Rat Myocytes J. Biol. Chem., September 5, 2003; 278(36): 33694 - 33700. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 W. G. Pyle, M. P. Sumandea, R. J. Solaro, and P. P. De Tombe Troponin I serines 43/45 and regulation of cardiac myofilament function Am J Physiol Heart Circ Physiol, September 1, 2002; 283(3): H1215 - H1224. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Pi, K. R. Kemnitz, D. Zhang, E. G. Kranias, and J. W. Walker Phosphorylation of Troponin I Controls Cardiac Twitch Dynamics: Evidence From Phosphorylation Site Mutants Expressed on a Troponin I-Null Background in Mice Circ. Res., April 5, 2002; 90(6): 649 - 656. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
K. Mackay and D. Mochly-Rosen Arachidonic acid protects neonatal rat cardiac myocytes from ischaemic injury through {epsilon} protein kinase C Cardiovasc Res, April 1, 2001; 50(1): 65 - 74. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. T Rapundalo Cardiac protein phosphorylation: functional and pathophysiological correlates Cardiovasc Res, June 1, 1998; 38(3): 559 - 588. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 C. M. Pawloski-Dahm, G. Song, D. L. Kirkpatrick, J. Palermo, J. Gulick, G. W. Dorn II, J. Robbins, and R. A. Walsh Effects of Total Replacement of Atrial Myosin Light Chain-2 With the Ventricular Isoform in Atrial Myocytes of Transgenic Mice Circulation, April 21, 1998; 97(15): 1508 - 1513. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Redaelli, A. Malhotra, B. Li, P. Li, E. H. Sonnenblick, P. A. Hofmann, and P. Anversa Effects of Constitutive Overexpression of Insulin-Like Growth Factor-1 on the Mechanical Characteristics and Molecular Properties of Ventricular Myocytes Circ. Res., March 23, 1998; 82(5): 594 - 603. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. Starkopf, T. V Andreasen, E. Bugge, and K. Ytrehus Lipid peroxidation, arachidonic acid and products of the lipoxygenase pathway in ischaemic preconditioning of rat heart Cardiovasc Res, January 1, 1998; 37(1): 66 - 75. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Malhotra, D. Reich, D. Reich, A. Nakouzi, V. Sanghi, D. L. Geenen, and P. M. Buttrick Experimental Diabetes Is Associated With Functional Activation of Protein Kinase C{epsilon} and Phosphorylation of Troponin I in the Heart, Which Are Prevented by Angiotensin II Receptor Blockade Circ. Res., December 19, 1997; 81(6): 1027 - 1033. [Abstract] [Full Text]
|
|
|
|
|
|
A. Sauvadet, T. Rohn, F. Pecker, and C. Pavoine Arachidonic Acid Drives Mini-glucagon Action in Cardiac Cells J. Biol. Chem., May 9, 1997; 272(19): 12437 - 12445. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Malhotra and V. Sanghi Regulation of contractile proteins in diabetic heart Cardiovasc Res, April 1, 1997; 34(1): 34 - 40. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 X. Huang, Y Pi, A. Lokuta, M. Greaser, and J. Walker Arachidonic acid stimulates protein kinase C-epsilon redistribution in heart cells J. Cell Sci., January 7, 1997; 110(14): 1625 - 1634. [Abstract] [PDF]
|
|
|
|
|
|
B.-X. Zhang, X. Ma, B. K. McConnell, D. S. Damron, and M. Bond Activation of Purinergic Receptors Triggers Oscillatory Contractions in Adult Rat Ventricular Myocytes Circ. Res., July 1, 1996; 79(1): 94 - 102. [Abstract] [Full Text]
|
|
|
|
|
|
Y. Pi, K. R. Kemnitz, D. Zhang, E. G. Kranias, and J. W. Walker Phosphorylation of Troponin I Controls Cardiac Twitch Dynamics: Evidence From Phosphorylation Site Mutants Expressed on a Troponin I-Null Background in Mice Circ. Res., April 5, 2002; 90(6): 649 - 656. [Abstract] [Full Text] [PDF]
|
|