Protein kinase C-alpha and -epsilon modulate connexin-43 phosphorylation in human heart

Area of Science:

• Cardiovascular Biology
• Molecular Cardiology
• Cell Signaling

Background:

• Protein kinase C (PKC)-alpha is elevated in failing human left ventricles, localizing to cardiomyocyte intercalated disks.
• The interaction and functional consequences of PKC-alpha and PKC-epsilon with connexin-43 (Cx-43) in failing hearts remain unclear.

Purpose of the Study:

• To investigate the spatial and functional interaction between PKC-alpha and Cx-43 in failing human hearts.
• To compare the PKC-alpha/Cx-43 association with that of PKC-epsilon/Cx-43.
• To explore these associations in non-failing hearts.

Main Methods:

• Co-immunoprecipitation of PKC-alpha, PKC-epsilon, and Cx-43 from non-failing and failing human left ventricles.
• Confocal microscopy to determine co-localization of PKC isoforms and Cx-43 within cardiomyocytes.
• In vitro phosphorylation assays using purified Cx-43 and recombinant PKC isoforms.

Main Results:

• PKC-alpha and Cx-43 co-localized in cardiomyocytes of both failing and non-failing human hearts.
• PKC-epsilon also co-localized with Cx-43 in cardiomyocytes.
• Both PKC-alpha and PKC-epsilon increased PKC activity in Cx-43 immunoprecipitated complexes, but only PKC-epsilon directly phosphorylated Cx-43.

Conclusions:

• PKC-alpha, PKC-epsilon, and Cx-43 form a closely associated complex in the human heart.
• PKC-epsilon directly phosphorylates Cx-43, while both isoforms contribute to increased phosphorylation within the complex.
• These findings elucidate novel signaling pathways in cardiac function and dysfunction.