Molecules and Cells

Korean Society for Molecular and Cellular Biology (한국분자세포생물학회)
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Integrated Quantitative Phosphoproteomics and Cell-Based Functional Screening Reveals Specific Pathological Cardiac Hypertrophy-Related Phosphorylation Sites

  • Kwon, Hye Kyeong (School of Life Sciences, Gwangju Institute of Science and Technology (GIST)) ;
  • Choi, Hyunwoo (School of Life Sciences, Gwangju Institute of Science and Technology (GIST)) ;
  • Park, Sung-Gyoo (School of Life Sciences, Gwangju Institute of Science and Technology (GIST)) ;
  • Park, Woo Jin (School of Life Sciences, Gwangju Institute of Science and Technology (GIST)) ;
  • Kim, Do Han (School of Life Sciences, Gwangju Institute of Science and Technology (GIST)) ;
  • Park, Zee-Yong (School of Life Sciences, Gwangju Institute of Science and Technology (GIST))
  • Received : 2019.01.06
  • Accepted : 2019.01.07
  • Published : 2021.07.31
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Abstract

Cardiac hypertrophic signaling cascades resulting in heart failure diseases are mediated by protein phosphorylation. Recent developments in mass spectrometry-based phosphoproteomics have led to the identification of thousands of differentially phosphorylated proteins and their phosphorylation sites. However, functional studies of these differentially phosphorylated proteins have not been conducted in a large-scale or high-throughput manner due to a lack of methods capable of revealing the functional relevance of each phosphorylation site. In this study, an integrated approach combining quantitative phosphoproteomics and cell-based functional screening using phosphorylation competition peptides was developed. A pathological cardiac hypertrophy model, junctate-1 transgenic mice and control mice, were analyzed using label-free quantitative phosphoproteomics to identify differentially phosphorylated proteins and sites. A cell-based functional assay system measuring hypertrophic cell growth of neonatal rat ventricle cardiomyocytes (NRVMs) following phenylephrine treatment was applied, and changes in phosphorylation of individual differentially phosphorylated sites were induced by incorporation of phosphorylation competition peptides conjugated with cell-penetrating peptides. Cell-based functional screening against 18 selected phosphorylation sites identified three phosphorylation sites (Ser-98, Ser-179 of Ldb3, and Ser-1146 of palladin) displaying near-complete inhibition of cardiac hypertrophic growth of NRVMs. Changes in phosphorylation levels of Ser-98 and Ser-179 in Ldb3 were further confirmed in NRVMs and other pathological/physiological hypertrophy models, including transverse aortic constriction and swimming models, using site-specific phospho-antibodies. Our integrated approach can be used to identify functionally important phosphorylation sites among differentially phosphorylated sites, and unlike conventional approaches, it is easily applicable for large-scale and/or high-throughput analyses.

Keywords

References

  1. Adkins, G.B. and Curtis, M.J. (2015). Potential role of cardiac chloride channels and transporters as novel therapeutic targets. Pharmacol. Ther. 145, 67-75. https://doi.org/10.1016/j.pharmthera.2014.08.002
  2. Bass, G.T., Ryall, K.A., Katikapalli, A., Taylor, B.E., Dang, S.T., Acton, S.T., and Saucerman, J.J. (2012). Automated image analysis identifies signaling pathways regulating distinct signatures of cardiac myocyte hypertrophy. J. Mol. Cell. Cardiol. 52, 923-930. https://doi.org/10.1016/j.yjmcc.2011.11.009
  3. Beck, M.R., Dixon, R.D.S., Goicoechea, S.M., Murphy, G.S., Brungardt, J.G., Beam, M.T., Srinath, P., Patel, J., Mohiuddin, J., Otey, C.A., et al. (2013). Structure and function of Palladin's actin binding domain. J. Mol. Biol. 425, 3325-3337. https://doi.org/10.1016/j.jmb.2013.06.016
  4. Benna, C., Peron, S., Rizzo, G., Faulkner, G., Megighian, A., Perini, G., Tognon, G., Valle, G., Reggiani, C., Costa, R., et al. (2009). Posttranscriptional silencing of the Drosophila homolog of human ZASP: a molecular and functional analysis. Cell Tissue Res. 337, 463-476. https://doi.org/10.1007/s00441-009-0813-y
  5. Brandenburg, S., Arakel, E.C., Schwappach, B., and Lehnart, S.E. (2016). The molecular and functional identities of atrial cardiomyocytes in health and disease. Biochim. Biophys. Acta 1863(7 Pt B), 1882-1893. https://doi.org/10.1016/j.bbamcr.2015.11.025
  6. Byrum, S.D., Larson, S.K., Avaritt, N.L., Moreland, L.E., Mackintosh, S.G., Cheung, W.L., and Tackett, A.J. (2013). Quantitative proteomics identifies activation of hallmark pathways of cancer in patient melanoma. J. Proteomics Bioinform. 6, 43-50. https://doi.org/10.4172/jpb.1000260
  7. Chang, Y.W., Chang, Y.T., Wang, Q., Lin, J.J.C., Chen, Y.J., and Chen, C.C. (2013). Quantitative phosphoproteomic study of pressure-overloaded mouse heart reveals dynamin-related protein 1 as a modulator of cardiac hypertrophy. Mol. Cell. Proteomics 12, 3094-3107. https://doi.org/10.1074/mcp.M113.027649
  8. Chin, Y.R. and Toker, A. (2010). The actin-bundling protein palladin is an Akt1-specific substrate that regulates breast cancer cell migration. Mol. Cell 38, 333-344. https://doi.org/10.1016/j.molcel.2010.02.031
  9. Choi, H., Lee, S., Jun, C.D., and Park, Z.Y. (2011). Development of an off-line capillary column IMAC phosphopeptide enrichment method for label-free phosphorylation relative quantification. J. Chromatogr. B Analyt. Technol. Biomed. Life Sci. 879, 2991-2997. https://doi.org/10.1016/j.jchromb.2011.08.035
  10. Craig, R., Lee, K.H., Mun, J.Y., Torre, I., and Luther, P.K. (2014). Structure, sarcomeric organization, and thin filament binding of cardiac myosinbinding protein-C. Pflugers Arch. 466, 425-431. https://doi.org/10.1007/s00424-013-1426-6
  11. Cui, H., Schlesinger, J., Schoenhals, S., Tonjes, M., Dunkel, I., Meierhofer, D., Cano, E., Schulz, K., Berger, M.F., Haack, T., et al. (2016). Phosphorylation of the chromatin remodeling factor DPF3a induces cardiac hypertrophy through releasing HEY repressors from DNA. Nucleic Acids Res. 44, 2538-2553. https://doi.org/10.1093/nar/gkv1244
  12. Day, E.K., Sosale, N.G., and Lazzara, M.J. (2016). Cell signaling regulation by protein phosphorylation: a multivariate, heterogeneous, and contextdependent process. Curr. Opin. Biotechnol. 40, 185-192. https://doi.org/10.1016/j.copbio.2016.06.005
  13. Di Carlo, M.N., Said, M., Ling, H., Valverde, C.A., De Giusti, V.C., Sommese, L., Palomeque, J., Aiello, E.A., Skapura, D.G., Rinaldi, G., et al. (2014). CaMKII-dependent phosphorylation of cardiac ryanodine receptors regulates cell death in cardiac ischemia/reperfusion injury. J. Mol. Cell. Cardiol. 74, 274-283. https://doi.org/10.1016/j.yjmcc.2014.06.004
  14. Dixon, R.D.S., Arneman, D.K., Rachlin, A.S., Sundaresan, N.R., Costello, M.J., Campbell, S.L., and Otey, C.A. (2008). Palladin is an actin cross-linking protein that uses immunoglobulin-like domains to bind filamentous actin. J. Biol. Chem. 283, 6222-6231. https://doi.org/10.1074/jbc.M707694200
  15. Dobrev, D. and Wehrens, X.H.T. (2014). Role of RyR2 phosphorylation in heart failure and arrhythmias: controversies around ryanodine receptor phosphorylation in cardiac disease. Circ. Res. 114, 1311-1319. https://doi.org/10.1161/CIRCRESAHA.114.300568
  16. Du, J., Wong, W.Y., Sun, L., Huang, Y., and Yao, X. (2012). Protein kinase G inhibits flow-induced Ca2+ entry into collecting duct cells. J. Am. Soc. Nephrol. 23, 1172-1180. https://doi.org/10.1681/ASN.2011100972
  17. Elkins, J.M., Papagrigoriou, E., Berridge, G., Yang, X., Phillips, C., Gileadi, C., Savitsky, P., and Doyle, D.A. (2007). Structure of PICK1 and other PDZ domains obtained with the help of self-binding C-terminal extensions. Protein Sci. 16, 683-694. https://doi.org/10.1110/ps.062657507
  18. Eom, G.H., Cho, Y.K., Ko, J.H., Shin, S., Choe, N., Kim, Y., Joung, H., Kim, H.S., Nam, K.I., Kee, H.J., et al. (2011). Casein kinase-2α1 induces hypertrophic response by phosphorylation of histone deacetylase 2 S394 and its activation in the heart. Circulation 123, 2392-2403. https://doi.org/10.1161/CIRCULATIONAHA.110.003665
  19. Faulkner, G., Pallavicini, A., Formentin, E., Comelli, A., Ievolella, C., Trevisan, S., Bortoletto, G., Scannapieco, P., Salamon, M., Mouly, V., et al. (1999). ZASP: a new Z-band alternatively spliced PDZ-motif protein. J. Cell Biol. 146, 465-475. https://doi.org/10.1083/jcb.146.2.465
  20. Feng, W., Shi, Y., Li, M., and Zhang, M. (2003). Tandem PDZ repeats in glutamate receptor-interacting proteins have a novel mode of PDZ domain-mediated target binding. Nat. Struct. Biol. 10, 972-978. https://doi.org/10.1038/nsb992
  21. Fischer, T.H., Herting, J., Mason, F.E., Hartmann, N., Watanabe, S., Nikolaev, V.O., Sprenger, J.U., Fan, P., Yao, L., Popov, A.F., et al. (2015). Late INa increases diastolic SR-Ca2+-leak in atrial myocardium by activating PKA and CaMKII. Cardiovasc. Res. 107, 184-196. https://doi.org/10.1093/cvr/cvv153
  22. Flashman, E., Redwood, C., Moolman-Smook, J., and Watkins, H. (2004). Cardiac myosin binding protein C: its role in physiology and disease. Circ. Res. 94, 1279-1289. https://doi.org/10.1161/01.RES.0000127175.21818.C2
  23. Frey, N. and Olson, E.N. (2002). Calsarcin-3, a novel skeletal muscle-specific member of the calsarcin family, interacts with multiple Z-disc proteins. J. Biol. Chem. 277, 13998-14004. https://doi.org/10.1074/jbc.M200712200
  24. Fu, Y., Westenbroek, R.E., Scheuer, T., and Catterall, W.A. (2013). Phosphorylation sites required for regulation of cardiac calcium channels in the fight-or-flight response. Proc. Natl. Acad. Sci. U. S. A. 110, 19621-19626. https://doi.org/10.1073/pnas.1319421110
  25. Goicoechea, S.M., Garcia-Mata, R., Staub, J., Valdivia, A., Sharek, L., Mcculloch, C.G., Hwang, R.F., Urrutia, R., Yeh, J.J., Kim, H.J., et al. (2014). Palladin promotes invasion of pancreatic cancer cells by enhancing invadopodia formation in cancer-associated fibroblasts. Oncogene 33, 1265-1273. https://doi.org/10.1038/onc.2013.68
  26. Gokce, E., Shuford, C.M., Franck, W.L., Dean, R.A., and Muddiman, D.C. (2011). Evaluation of normalization methods on GeLC-MS/MS label-free spectral counting data to correct for variation during proteomic workflows. J. Am. Soc. Mass Spectrom. 22, 2199-2208. https://doi.org/10.1007/s13361-011-0237-2
  27. Gresham, K.S. and Stelzer, J.E. (2016). The contributions of cardiac myosin binding protein C and troponin I phosphorylation to β-adrenergic enhancement of in vivo cardiac function. J. Physiol. 594, 669-686. https://doi.org/10.1113/JP270959
  28. Griggs, R., Vihola, A., Hackman, P., Talvinen, K., Haravuori, H., Faulkner, G., Eymard, B., Richard, I., Selcen, D., Engel, A., et al. (2007). Zaspopathy in a large classic late-onset distal myopathy family. Brain 130, 1477-1484. https://doi.org/10.1093/brain/awm006
  29. Guerra-Castellano, A., Diaz-Moreno, I., Velazquez-Campoy, A., De La Rosa, M.A., and Diaz-Quintana, A. (2016). Structural and functional characterization of phosphomimetic mutants of cytochrome c at threonine 28 and serine 47. Biochim. Biophys. Acta 1857, 387-395. https://doi.org/10.1016/j.bbabio.2016.01.011
  30. Heineke, J. and Molkentin, J.D. (2006). Regulation of cardiac hypertrophy by intracellular signalling pathways. Nat. Rev. Mol. Cell Biol. 7, 589-600. https://doi.org/10.1038/nrm1983
  31. Hong, C.S., Cho, M.C., Kwak, Y.G., Song, C.H., Lee, Y.H., Lim, J.S., Kwon, Y.K., Chae, S.W., and Kim, D.H. (2002). Cardiac remodeling and atrial fibrillation in transgenic mice overexpressing junctin. FASEB J. 16, 1310-1312. https://doi.org/10.1096/fj.01-0908fje
  32. Hong, C.S., Kwak, Y.G., Ji, J.H., Chae, S.W., and Kim, D.H. (2001). Molecular cloning and characterization of mouse cardiac junctate isoforms. Biochem. Biophys. Res. Commun. 289, 882-887. https://doi.org/10.1006/bbrc.2001.6056
  33. Hong, C.S., Kwon, S.J., Cho, M.C., Kwak, Y.G., Ha, K.C., Hong, B., Li, H., Chae, S.W., Chai, O.H., Song, C.H., et al. (2008). Overexpression of junctate induces cardiac hypertrophy and arrhythmia via altered calcium handling. J. Mol. Cell. Cardiol. 44, 672-682. https://doi.org/10.1016/j.yjmcc.2008.01.012
  34. Huang, C., Zhou, Q., Liang, P., Hollander, M.S., Sheikh, F., Li, X., Greaser, M., Shelton, G.D., Evans, S., and Chen, J. (2003). Characterization and in vivo functional analysis of splice variants of cypher. J. Biol. Chem. 278, 7360-7365. https://doi.org/10.1074/jbc.M211875200
  35. Huang, D.W., Sherman, B.T., and Lempicki, R.A. (2009). Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. Nat. Protoc. 4, 44-57. https://doi.org/10.1038/nprot.2008.211
  36. Jentzsch, C., Leierseder, S., Loyer, X., Flohrschutz, I., Sassi, Y., Hartmann, D., Thum, T., Laggerbauer, B., and Engelhardt, S. (2012). A phenotypic screen to identify hypertrophy-modulating microRNAs in primary cardiomyocytes. J. Mol. Cell. Cardiol. 52, 13-20. https://doi.org/10.1016/j.yjmcc.2011.07.010
  37. Jia, W., Shaffer, J.F., Harris, S.P., and Leary, J.A. (2010). Identification of novel protein kinase A phosphorylation sites in the M-domain of human and murine cardiac myosin binding protein-C using mass spectrometry analysis. J. Proteome Res. 9, 1843-1853. https://doi.org/10.1021/pr901006h
  38. Jin, L. (2011). The actin associated protein palladin in smooth muscle and in the development of diseases of the cardiovasculature and in cancer. J. Muscle Res. Cell Motil. 32, 7-17. https://doi.org/10.1007/s10974-011-9246-9
  39. Kall, L., Canterbury, J.D., Weston, J., Noble, W.S., and MacCoss, M.J. (2007). Semi-supervised learning for peptide identification from shotgun proteomics datasets. Nat. Methods 4, 923-925. https://doi.org/10.1038/nmeth1113
  40. Kho, C., Lee, A., Jeong, D., Oh, J.G., Chaanine, A.H., Kizana, E., Park, W.J., and Hajjar, R.J. (2011). SUMO1-dependent modulation of SERCA2a in heart failure. Nature 477, 601-605. https://doi.org/10.1038/nature10407
  41. Klaavuniemi, T., Alho, N., Hotulainen, P., Kelloniemi, A., Havukainen, H., Permi, P., Mattila, S., and Ylanne, J. (2009). Characterization of the interaction between Actinin-Associated LIM Protein (ALP) and the rod domain of α-actinin. BMC Cell Biol. 10, 22. https://doi.org/10.1186/1471-2121-10-22
  42. Klaavuniemi, T., Kelloniemi, A., and Ylanne, J. (2004). The ZASP-like motif in actinin-associated LIM protein is required for interaction with the α-actinin rod and for targeting to the muscle Z-line. J. Biol. Chem. 279, 26402-26410. https://doi.org/10.1074/jbc.M401871200
  43. Klaavuniemi, T. and Ylanne, J. (2006). Zasp/Cypher internal ZM-motif containing fragments are sufficient to co-localize with α-actinin--analysis of patient mutations. Exp. Cell Res. 312, 1299-1311. https://doi.org/10.1016/j.yexcr.2005.12.036
  44. Kohli, S., Ahuja, S., and Rani, V. (2011). Transcription factors in heart: promising therapeutic targets in cardiac hypertrophy. Curr. Cardiol. Rev. 7, 262-271. https://doi.org/10.2174/157340311799960618
  45. Kooij, V., Saes, M., Jaquet, K., Zaremba, R., Foster, D.B., Murphy, A.M., dos Remedios, C., van der Velden, J., and Stienen, G.J.M. (2010). Effect of troponin I Ser23/24 phosphorylation on Ca2+-sensitivity in human myocardium depends on the phosphorylation background. J. Mol. Cell. Cardiol. 48, 954-963. https://doi.org/10.1016/j.yjmcc.2010.01.002
  46. Kranias, E.G. and Hajjar, R.J. (2012). Modulation of cardiac contractility by the phopholamban/SERCA2a regulatome. Circ. Res. 110, 1646-1660. https://doi.org/10.1161/CIRCRESAHA.111.259754
  47. Kuzmanov, U., Guo, H., Buchsbaum, D., Cosme, J., Abbasi, C., Isserlin, R., Sharma, P., Gramolini, A.O., and Emili, A. (2016). Global phosphoproteomic profiling reveals perturbed signaling in a mouse model of dilated cardiomyopathy. Proc. Natl. Acad. Sci. U. S. A. 113, 12592-12597. https://doi.org/10.1073/pnas.1606444113
  48. Kwon, S.J. and Kim, D.H. (2009). Characterization of junctate-SERCA2a interaction in murine cardiomyocyte. Biochem. Biophys. Res. Commun. 390, 1389-1394. https://doi.org/10.1016/j.bbrc.2009.10.165
  49. Li, J., Imtiaz, M.S., Beard, N.A., Dulhunty, A.F., Thorne, R., VanHelden, D.F., and Laver, D.R. (2013). β-Adrenergic stimulation increases RyR2 activity via intracellular Ca2+ and Mg2+ regulation. PLoS One 8, e58334. https://doi.org/10.1371/journal.pone.0058334
  50. Liao, K.A., Gonzalez-Morales, N., and Schock, F. (2016). Zasp52, a core Z-disc protein in Drosophila indirect flight muscles, interacts with α-actinin via an extended PDZ domain. PLoS Genet. 12, e1006400. https://doi.org/10.1371/journal.pgen.1006400
  51. Lin, C., Guo, X., Lange, S., Liu, J., Ouyang, K., Yin, X., Jiang, L., Cai, Y., Mu, Y., Sheikh, F., et al. (2013). Cypher/ZASP is a novel a-kinase anchoring protein. J. Biol. Chem. 288, 29403-29413. https://doi.org/10.1074/jbc.M113.470708
  52. Lin, X., Ruiz, J., Bajraktari, I., Ohman, R., Banerjee, S., Gribble, K., Kaufman, J.D., Wingfield, P.T., Griggs, R.C., Fischbeck, K.H., et al. (2014). Z-disc-associated, alternatively spliced, PDZ motif-containing protein (ZASP) mutations in the actin-binding domain cause disruption of skeletal muscle actin filaments in myofibrillar myopathy. J. Biol. Chem. 289, 13615-13626. https://doi.org/10.1074/jbc.M114.550418
  53. Lindskog, C., Linne, J., Fagerberg, L., Hallstrom, B.M., Sundberg, C.J., Lindholm, M., Huss, M., Kampf, C., Choi, H., Liem, D.A., et al. (2015). The human cardiac and skeletal muscle proteomes defined by transcriptomics and antibody-based profiling. BMC Genomics 16, 475. https://doi.org/10.1186/s12864-015-1686-y
  54. Long, J., Wei, Z., Feng, W., Yu, C., Zhao, Y.X., and Zhang, M. (2008). Supramodular nature of GRIP1 revealed by the structure of its PDZ12 tandem in complex with the carboxyl tail of Fras1. J. Mol. Biol. 375, 1457-1468. https://doi.org/10.1016/j.jmb.2007.11.088
  55. Lorenz, K., Schmitt, J.P., Schmitteckert, E.M., and Lohse, M.J. (2009). A new type of ERK1/2 autophosphorylation causes cardiac hypertrophy. Nat. Med. 15, 75-83. https://doi.org/10.1038/nm.1893
  56. Lou, Q., Janardhan, A., and Efimov, I.R. (2012). Remodeling of calcium handling in human heart failure. Adv. Exp. Med. Biol. 740, 1145-1174. https://doi.org/10.1007/978-94-007-2888-2_52
  57. Luck, K., Charbonnier, S., and Trave, G. (2012). The emerging contribution of sequence context to the specificity of protein interactions mediated by {PDZ} domains. FEBS Lett. 586, 2648-2661. https://doi.org/10.1016/j.febslet.2012.03.056
  58. Lundby, A., Andersen, M.N., Steffensen, A.B., Horn, H., Kelstrup, C.D., Francavilla, C., Jensen, L.J., Schmitt, N., Thomsen, M.B., and Olsen, J.V. (2013). In vivo phosphoproteomics analysis reveals the cardiac targets of β-adrenergic receptor signaling. Sci. Signal. 6, rs11. https://doi.org/10.1126/scisignal.2003506
  59. Luther, P.K., Winkler, H., Taylor, K., Zoghbi, M.E., Craig, R., Padron, R., Squire, J.M., and Liu, J. (2011). Direct visualization of myosin-binding protein C bridging myosin and actin filaments in intact muscle. Proc. Natl. Acad. Sci. U. S. A. 108, 11423-11428. https://doi.org/10.1073/pnas.1103216108
  60. MacLennan, D.H. and Kranias, E.G. (2003). Phospholamban: a crucial regulator of cardiac contractility. Nat. Rev. Mol. Cell Biol. 4, 566-577. https://doi.org/10.1038/nrm1151
  61. Mamidi, R., Gresham, K.S., Li, J., and Stelzer, J.E. (2017). Cardiac myosin binding protein-C Ser 302 phosphorylation regulates cardiac β-adrenergic reserve. Sci. Adv. 3, e1602445. https://doi.org/10.1126/sciadv.1602445
  62. Martinelli, V.C., Kyle, W.B., Kojic, S., Vitulo, N., Li, Z., Belgrano, A., Maiuri, P., Banks, L., Vatta, M., Valle, G., et al. (2014). ZASP interacts with the mechanosensing protein Ankrd2 and p53 in the signalling network of striated muscle. PLoS One 9, e92259. https://doi.org/10.1371/journal.pone.0092259
  63. Mattiazzi, A. and Kranias, E.G. (2014). The role of CaMKII regulation of phospholamban activity in heart disease. Front. Pharmacol. 5, 5. https://doi.org/10.3389/fphar.2014.00005
  64. McLane, J.S. and Ligon, L.A. (2015). Palladin mediates stiffness-induced fibroblast activation in the tumor microenvironment. Biophys. J. 109, 249-264. https://doi.org/10.1016/j.bpj.2015.06.033
  65. Michalek, A.J., Howarth, J.W., Gulick, J., Previs, M.J., Robbins, J., Rosevear, P.R., and Warshaw, D.M. (2013). Phosphorylation modulates the mechanical stability of the cardiac myosin-binding protein C motif. Biophys. J. 104, 442-452. https://doi.org/10.1016/j.bpj.2012.12.021
  66. Najm, P. and El-Sibai, M. (2014). Palladin regulation of the actin structures needed for cancer invasion. Cell Adh. Migr. 8, 29-35. https://doi.org/10.4161/cam.28024
  67. Nishida, K., Michael, G., Dobrev, D., and Nattel, S. (2010). Animal models for atrial fibrillation: clinical insights and scientific opportunities. Europace 12, 160-172. https://doi.org/10.1093/europace/eup328
  68. Oh, J.G., Kim, J., Jang, S.P., Nguen, M., Yang, D.K., Jeong, D., Park, Z.Y., Park, S.G., Hajjar, R.J., and Park, W.J. (2013). Decoy peptides targeted to protein phosphatase 1 inhibit dephosphorylation of phospholamban in cardiomyocytes. J. Mol. Cell. Cardiol. 56, 63-71. https://doi.org/10.1016/j.yjmcc.2012.12.005
  69. Olsen, J.V., Vermeulen, M., Santamaria, A., Kumar, C., Miller, M.L., Jensen, L.J., Gnad, F., Cox, J., Jensen, T.S., Nigg, E.A., et al. (2010). Quantitative phosphoproteomics reveals widespread full phosphorylation site occupancy during mitosis. Sci. Signal. 3, ra3. https://doi.org/10.1126/scisignal.2000475
  70. Otey, C.A., Rachlin, A., Moza, M., Arneman, D., and Carpen, O. (2005). The palladin/myotilin/myopalladin family of actin-associated scaffolds. Int. Rev. Cytol. 246, 31-58. https://doi.org/10.1016/S0074-7696(05)46002-7
  71. Palermo, J., Gulick, J., Colbert, M., Fewell, J., and Robbins, J. (1996). Transgenic remodeling of the contractile apparatus in the mammalian heart. Circ. Res. 78, 504-509. https://doi.org/10.1161/01.RES.78.3.504
  72. Parast, M.M. and Otey, C.A. (2000). Characterization of palladin, a novel protein localized to stress fibers and cell adhesions. J. Cell Biol. 150, 643-655. https://doi.org/10.1083/jcb.150.3.643
  73. Passier, R., Richardson, J.A., and Olson, E.N. (2000). Oracle, a novel PDZ-LIM domain protein expressed in heart and skeletal muscle. Mech. Dev. 92, 277-284. https://doi.org/10.1016/S0925-4773(99)00330-5
  74. Patel, P.C., Fisher, K.H., Yang, E.C.C., Deane, C.M., and Harrison, R.E. (2009). Proteomic analysis of microtubule-associated proteins during macrophage activation. Mol. Cell. Proteomics 8, 2500-2514. https://doi.org/10.1074/mcp.M900190-MCP200
  75. Peter, A.K., Bjerke, M.A., and Leinwand, L.A. (2016). Biology of the cardiac myocyte in heart disease. Mol. Biol. Cell 27, 2149-2160. https://doi.org/10.1091/mbc.E16-01-0038
  76. Pollak, A.J., Haghighi, K., Kunduri, S., Arvanitis, D.A., Bidwell, P.A., Liu, G.S., Singh, V.P., Gonzalez, D.J., Sanoudou, D., Wiley, S.E., et al. (2017). Phosphorylation of serine96 of histidine-rich calcium-binding protein by the Fam20C kinase functions to prevent cardiac arrhythmia. Proc. Natl. Acad. Sci. U. S. A. 114, 9098-9103. https://doi.org/10.1073/pnas.1706441114
  77. Pondugula, S.R., Brimer-Cline, C., Wu, J., Schuetz, E.G., Tyagi, R.K., and Chen, T. (2009). A phosphomimetic mutation at threonine-57 abolishes transactivation activity and alters nuclear localization pattern of human pregnane X receptor. Drug Metab. Dispos. 37, 719-730. https://doi.org/10.1124/dmd.108.024695
  78. Previs, M.J., Mun, J.Y., Michalek, A.J., Previs, S.B., Gulick, J., Robbins, J., Warshaw, D.M., and Craig, R. (2016). Phosphorylation and calcium antagonistically tune myosin-binding protein C's structure and function. Proc. Natl. Acad. Sci. U. S. A. 113, 3239-3244. https://doi.org/10.1073/pnas.1522236113
  79. Rachlin, A.S. and Otey, C.A. (2006). Identification of palladin isoforms and characterization of an isoform-specific interaction between Lasp-1 and palladin. J. Cell Sci. 119, 995-1004. https://doi.org/10.1242/jcs.02825
  80. Reid, B.G., Stratton, M.S., Bowers, S., Cavasin, M.A., Demos-Davies, K.M., Susano, I., and McKinsey, T.A. (2016). Discovery of novel small molecule inhibitors of cardiac hypertrophy using high throughput, high content imaging. J. Mol. Cell. Cardiol. 97, 106-113. https://doi.org/10.1016/j.yjmcc.2016.04.015
  81. Respress, J.L., Van Oort, R.J., Li, N., Rolim, N., Dixit, S.S., Dealmeida, A., Voigt, N., Lawrence, W.S., Skapura, D.G., Skardal, K., et al. (2012). Role of RyR2 phosphorylation at S2814 during heart failure progression. Circ. Res. 110, 1474-1483. https://doi.org/10.1161/CIRCRESAHA.112.268094
  82. Rohini, A., Agrawal, N., Koyani, C.N., and Singh, R. (2010). Molecular targets and regulators of cardiac hypertrophy. Pharmacol. Res. 61, 269-280. https://doi.org/10.1016/j.phrs.2009.11.012
  83. Rosas, P.C., Liu, Y., Abdalla, M.I., Thomas, C.M., Kidwell, D.T., Dusio, G.F., Mukhopadhyay, D., Kumar, R., Baker, K.M., Mitchell, B.M., et al. (2015). Phosphorylation of cardiac myosin-binding protein-C is a critical mediator of diastolic function. Circ. Heart Fail. 8, 582-594. https://doi.org/10.1161/CIRCHEARTFAILURE.114.001550
  84. Rowin, E.J., Hausvater, A., Link, M.S., Abt, P., Gionfriddo, W., Wang, W., Rastegar, H., Estes, N.A.M., Maron, M.S., and Maron, B.J. (2017). Clinical profile and consequences of atrial fibrillation in hypertrophic cardiomyopathy. Circulation 136, 2420-2436. https://doi.org/10.1161/CIRCULATIONAHA.117.029267
  85. Ruppert, C., Deiss, K., Herrmann, S., Vidal, M., Oezkur, M., Gorski, A., Weidemann, F., Lohse, M.J., and Lorenz, K. (2013). Interference with ERKThr188 phosphorylation impairs pathological but not physiological cardiac hypertrophy. Proc. Natl. Acad. Sci. U. S. A. 110, 7440-7445. https://doi.org/10.1073/pnas.1221999110
  86. Ryall, K.A., Bezzerides, V.J., Rosenzweig, A., and Saucerman, J.J. (2014). Phenotypic screen quantifying differential regulation of cardiac myocyte hypertrophy identifies CITED4 regulation of myocyte elongation. J. Mol. Cell. Cardiol. 72, 74-84. https://doi.org/10.1016/j.yjmcc.2014.02.013
  87. Sadoshima, J.I., Jahn, L., Takahashi, T., Kulik, T.J., and Izumo, S. (1992). Molecular characterization of the stretch-induced adaptation of cultured cardiac cells. An in vitro model of load-induced cardiac hypertrophy. J. Biol. Chem. 267, 10551-10560. https://doi.org/10.1016/S0021-9258(19)50052-7
  88. Schechter, M.A., Hsieh, M.K.H., Njoroge, L.W., Thompson, J.W., Soderblom, E.J., Feger, B.J., Troupes, C.D., Hershberger, K.A., Ilkayeva, O.R., Nagel, W.L., et al. (2014). Phosphoproteomic profiling of human myocardial tissues distinguishes ischemic from non-ischemic end stage heart failure. PLoS One 9, e104157. https://doi.org/10.1371/journal.pone.0104157
  89. Scholten, A., Preisinger, C., Corradini, E., Bourgonje, V.J., Hennrich, M.L., van Veen, T.A.B., Swaminathan, P.D., Joiner, M.L., Vos, M.A., Anderson, M.E., et al. (2013). Phosphoproteomics study based on in vivo inhibition reveals sites of calmodulin-dependent protein kinase II regulation in the heart. J. Am. Heart Assoc. 2, e000318. https://doi.org/10.1161/JAHA.113.000318
  90. Seko, Y., Kato, T., Haruna, T., Izumi, T., Miyamoto, S., Nakane, E., and Inoko, M. (2018). Association between atrial fibrillation, atrial enlargement, and left ventricular geometric remodeling. Sci. Rep. 8, 6366. https://doi.org/10.1038/s41598-018-24875-1
  91. Selcen, D. and Engel, A.G. (2005). Mutations in ZASP define a novel form of muscular dystrophy in humans. Ann. Neurol. 57, 269-276. https://doi.org/10.1002/ana.20376
  92. Shan, J., Betzenhauser, M.J., Kushnir, A., Reiken, S., Meli, A.C., Wronska, A., Dura, M., Chen, B.X., and Marks, A.R. (2010a). Role of chronic ryanodine receptor phosphorylation in heart failure and β-adrenergic receptor blockade in mice. J. Clin. Invest. 120, 4375-4387. https://doi.org/10.1172/JCI37649
  93. Shan, J., Kushnir, A., Betzenhauser, M.J., Reiken, S., Li, J., Lehnart, S.E., Lindegger, N., Mongillo, M., Mohler, P.J., and Marks, A.R. (2010b). Phosphorylation of the ryanodine receptor mediates the cardiac fight or flight response in mice. J. Clin. Invest. 120, 4388-4398. https://doi.org/10.1172/JCI32726
  94. Shimizu, I. and Minamino, T. (2016). Physiological and pathological cardiac hypertrophy. J. Mol. Cell. Cardiol. 97, 245-262. https://doi.org/10.1016/j.yjmcc.2016.06.001
  95. Song, H.K., Hong, S.E., Kim, T., and Kim, D.H. (2012). Deep RNA sequencing reveals novel cardiac transcriptomic signatures for physiological and pathological hypertrophy. PLoS One 7, e35552. https://doi.org/10.1371/journal.pone.0035552
  96. Song, J., Gao, E., Wang, J., Zhang, X.Q., Chan, T.O., Koch, W.J., Shang, X., Joseph, J.I., Peterson, B.Z., Feldman, A.M., et al. (2012). Constitutive overexpression of phosphomimetic phospholemman S68E mutant results in arrhythmias, early mortality, and heart failure: potential involvement of Na+/Ca2+ exchanger. Am. J. Physiol. Heart Circ. Physiol. 302, H770-H781. https://doi.org/10.1152/ajpheart.00733.2011
  97. Taus, T., Kocher, T., Pichler, P., Paschke, C., Schmidt, A., Henrich, C., and Mechtler, K. (2011). Universal and confident phosphorylation site localization using phosphoRS. J. Proteome Res. 10, 5354-5362. https://doi.org/10.1021/pr200611n
  98. te Velthuis, A.J.W., Isogai, T., Gerrits, L., and Bagowski, C.P. (2007). Insights into the molecular evolution of the PDZ/LIM family and identification of a novel conserved protein motif. PLoS One 2, e189. https://doi.org/10.1371/journal.pone.0000189
  99. Tham, Y.K., Bernardo, B.C., Ooi, J.Y.Y., Weeks, K.L., and McMullen, J.R. (2015). Pathophysiology of cardiac hypertrophy and heart failure: signaling pathways and novel therapeutic targets. Arch. Toxicol. 89, 1401-1438. https://doi.org/10.1007/s00204-015-1477-x
  100. van Berlo, J.H., Elrod, J.W., Aronow, B.J., Pu, W.T., and Molkentin, J.D. (2011). Serine 105 phosphorylation of transcription factor GATA4 is necessary for stress-induced cardiac hypertrophy in vivo. Proc. Natl. Acad. Sci. U. S. A. 108, 12331-12336. https://doi.org/10.1073/pnas.1104499108
  101. van der Meer, D.L.M., Marques, I.J., Leito, J.T.D., Besser, J., Bakkers, J., Schoonheere, E., and Bagowski, C.P. (2006). Zebrafish cypher is important for somite formation and heart development. Dev. Biol. 299, 356-372. https://doi.org/10.1016/j.ydbio.2006.07.032
  102. Vatta, M., Mohapatra, B., Jimenez, S., Sanchez, X., Faulkner, G., Perles, Z., Sinagra, G., Lin, J.H., Vu, T.M., Zhou, Q., et al. (2003). Mutations in Cypher/ZASP in patients with dilated cardiomyopathy and left ventricular noncompaction. J. Am. Coll. Cardiol. 42, 2014-2027. https://doi.org/10.1016/j.jacc.2003.10.021
  103. Vattepu, R., Yadav, R., and Beck, M.R. (2015). Actin-induced dimerization of palladin promotes actin-bundling. Protein Sci. 24, 70-80. https://doi.org/10.1002/pro.2588
  104. Vizcaino, J.A., Csordas, A., Del-Toro, N., Dianes, J.A., Griss, J., Lavidas, I., Mayer, G., Perez-Riverol, Y., Reisinger, F., Ternent, T., et al. (2016). 2016 update of the PRIDE database and its related tools. Nucleic Acids Res. 44, D447-D456. https://doi.org/10.1093/nar/gkv1145
  105. Wang, C.K., Pan, L., Chen, J., and Zhang, M. (2010). Extensions of PDZ domains as important structural and functional elements. Protein Cell 1, 737-751. https://doi.org/10.1007/s13238-010-0099-6
  106. Wang, H.V. and Moser, M. (2008). Comparative expression analysis of the murine palladin isoforms. Dev. Dyn. 237, 3342-3351. https://doi.org/10.1002/dvdy.21755
  107. Wang, N., Su, P., Zhang, Y., Lu, J., Xing, B., Kang, K., Li, W., and Wang, Y. (2014). Protein kinase D1-dependent phosphorylation of dopamine D1 receptor regulates cocaine-induced behavioral responses. Neuropsychopharmacology 39, 1290-1301. https://doi.org/10.1038/npp.2013.341
  108. Weeland, C.J., van den Hoogenhof, M.M., Beqqali, A., and Creemers, E.E. (2015). Insights into alternative splicing of sarcomeric genes in the heart. J. Mol. Cell. Cardiol. 81, 107-113. https://doi.org/10.1016/j.yjmcc.2015.02.008
  109. Wheeler-Jones, C.P.D. (2005). Cell signalling in the cardiovascular system: an overview. Heart 91, 1366-1374. https://doi.org/10.1136/hrt.2005.072280
  110. Wu, R., Dephoure, N., Haas, W., Huttlin, E.L., Zhai, B., Sowa, M.E., and Gygi, S.P. (2011). Correct interpretation of comprehensive phosphorylation dynamics requires normalization by protein expression changes. Mol. Cell. Proteomics 10, M111.009654. https://doi.org/10.1074/mcp.M111.009654
  111. Wu, Y.B., Dai, J., Yang, X.L., Li, S.J., Zhao, S.L., Sheng, Q.H., Tang, J.S., Zheng, G.Y., Li, Y.X., Wu, J.R., et al. (2009). Concurrent quantification of proteome and phosphoproteome to reveal system-wide association of protein phosphorylation and gene expression. Mol. Cell. Proteomics 8, 2809-2826. https://doi.org/10.1074/mcp.M900293-MCP200
  112. Xi, Y., Ai, T., De Lange, E., Li, Z., Wu, G., Brunelli, L., Kyle, W.B., Turker, I., Cheng, J., Ackerman, M.J., et al. (2012). Loss of function of hNav1.5 by a ZASP1 mutation associated with intraventricular conduction disturbances in left ventricular noncompaction. Circ. Arrhythm. Electrophysiol. 5, 1017-1026. https://doi.org/10.1161/CIRCEP.111.969220
  113. Xing, Y., Ichida, F., Matsuoka, T., Isobe, T., Ikemoto, Y., Higaki, T., Tsuji, T., Haneda, N., Kuwabara, A., Chen, R., et al. (2006). Genetic analysis in patients with left ventricular noncompaction and evidence for genetic heterogeneity. Mol. Genet. Metab. 88, 71-77. https://doi.org/10.1016/j.ymgme.2005.11.009
  114. Yang, L., Dai, D.F., Yuan, C., Westenbroek, R.E., Yu, H., West, N., de la Iglesia, H.O., and Catterall, W.A. (2016). Loss of β-adrenergic-stimulated phosphorylation of Ca V 1.2 channels on Ser1700 leads to heart failure. Proc. Natl. Acad. Sci. U. S. A. 113, E7976-E7985.
  115. Yin, Z., Ren, J., and Guo, W. (2015). Sarcomeric protein isoform transitions in cardiac muscle: a journey to heart failure. Biochim. Biophys. Acta 1852, 47-52. https://doi.org/10.1016/j.bbadis.2014.11.003
  116. Yuan, C.C., Muthu, P., Kazmierczak, K., Liang, J., Huang, W., Irving, T.C., Kanashiro-Takeuchi, R.M., Hare, J.M., and Szczesna-Cordary, D. (2015). Constitutive phosphorylation of cardiac myosin regulatory light chain prevents development of hypertrophic cardiomyopathy in mice. Proc. Natl. Acad. Sci. U. S. A. 112, E4138-E4146.
  117. Zhang, J., Lanham, K.A., Heideman, W., Peterson, R.E., and Li, L. (2013). Statistically enhanced spectral counting approach to TCDD cardiac toxicity in the adult zebrafish heart. J. Proteome Res. 12, 3093-3103. https://doi.org/10.1021/pr400312u
  118. Zhang, J., Petit, C.M., King, D.S., and Lee, A.L. (2011). Phosphorylation of a PDZ domain extension modulates binding affinity and interdomain interactions in postsynaptic density-95 (PSD-95) protein, a membraneassociated guanylate kinase (MAGUK). J. Biol. Chem. 286, 41776-41785. https://doi.org/10.1074/jbc.M111.272583
  119. Zheng, M., Cheng, H., Banerjee, I., and Chen, J. (2010). ALP / Enigma PDZLIM domain proteins in the heart. J. Mol. Cell Biol. 36, 96-102.
  120. Zheng, M., Cheng, H., Li, X., Zhang, J., Cui, L., Ouyang, K., Han, L., Zhao, T., Gu, Y., Dalton, N.D., et al. (2009). Cardiac-specific ablation of Cypher leads to a severe form of dilated cardiomyopathy with premature death. Hum. Mol. Genet. 18, 701-713. https://doi.org/10.1093/hmg/ddn400
  121. Zhou, Q., Chu, P.H., Huang, C., Cheng, C.F., Martone, M.E., Knoll, G., Diane Shelton, G., Evans, S., and Chen, J. (2001). Ablation of Cypher, a PDZ-LIM domain Z-line protein, causes a severe form of congenital myopathy. J. Cell Biol. 155, 605-612. https://doi.org/10.1083/jcb.200107092
  122. Zhou, Q., Ruiz-Lozano, P., Martone, M.E., and Chen, J. (1999). Cypher, a striated muscle-restricted PDZ and LIM domain-containing protein, binds to alpha-actinin-2 and protein kinase C. J. Biol. Chem. 274, 19807-19813. https://doi.org/10.1074/jbc.274.28.19807