Protein kinase C-α and -ε modulate connexin-43 phosphorylation in human heart

Nancy Bowling, Xiao Di Huang, George E. Sandusky, Rebecca L. Fouts, Karen Mintze, Michail Esterman, Paul D. Allen, Rosemarie Maddi, Eileen McCall, Chris J. Vlahos

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Abstract

We have previously demonstrated that protein kinase C (PKC)-α expression is significantly elevated in failing human left ventricle, with immunostaining showing increased PKC-α localization at the intercalated disks of cardiomyocytes. In the present study we sought to determine, in the failing heart, if PKC-α interacted with connexin-43 (Cx-43) both spatially and functionally, and to compare the association of PKC-α/Cx-43 with that of PKC-ε, a PKC isozyme that does not significantly increase in failing hearts. The possibility of a PKC-α or PKC-ε/Cx-43 association in non-failing hearts was also investigated. Co-immunoprecipitation of PKC-α or PKC-ε and Cx-43 in non-failing and failing left ventricle was achieved using antibodies to PKC-α or Cx-43. Confocal microscopy confirmed that PKC-α distribution within the cardiomyocyte included co-localization with connexin-43 in both falling and non-failing myocardium. In a similar manner, confocal imaging of PKC-ε showed cardiomyocyte distribution in both cytosol and membrane, and colocalization of PKC-ε with Cx-43. Recombinant PKC-α or -ε increased PKC activity significantly above endogenous levels in the co-immunoprecipitated Cx-43 complexes (P<0.05). However, phosphorylation of purified human Cx-43 (isolated from failing human left ventricle) by recombinant PKC-α or PKC-ε resulted in only PKC-ε mediated Cx-43 phosphorylation. Thus, in the human heart PKC-α, PKC-ε, and Cx-43 appear to form a closely associated complex. Whereas only PKC-ε directly phosphorylates Cx-43, both PKC isoforms result in increased phosphorylation within the Cx-43 co-immunoprecipitated complex.

Original languageEnglish (US)
Pages (from-to)789-798
Number of pages10
JournalJournal of Molecular and Cellular Cardiology
Volume33
Issue number4
DOIs
StatePublished - Jan 1 2001

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Keywords

  • Cardiomyopathy
  • Gap junctions
  • Phosphoprotein
  • Protein kinase
  • Signal transduction

ASJC Scopus subject areas

  • Molecular Biology
  • Cardiology and Cardiovascular Medicine

Cite this

Bowling, N., Huang, X. D., Sandusky, G. E., Fouts, R. L., Mintze, K., Esterman, M., Allen, P. D., Maddi, R., McCall, E., & Vlahos, C. J. (2001). Protein kinase C-α and -ε modulate connexin-43 phosphorylation in human heart. Journal of Molecular and Cellular Cardiology, 33(4), 789-798. https://doi.org/10.1006/jmcc.2000.1349