Protein kinase C signaling transduces endorphin-primed cardiogenesis in GTR1 embryonic stem cells

Abstract

The prodynorphin gene and its product, dynorphin B, have been found to promote cardiogenesis in embryonic cells by inducing the expression of GATA-4 and Nkx-2.5, two transcription factor-encoding genes essential for cardiogenesis. The molecular mechanism(s) underlying endorphin-induced cardiogenesis remain unknown. In the present study, we found that GTRI embryonic stem (ES) cells expressed cell surface κ opioid receptors, as well as protein kinase C (PKC)-α, -β1, -β2, -δ, -ε, and -ζ. Cardiac differentiation was associated with a marked increase in the Bmax value for a selective opioid receptor ligand and complex subcellular redistribution of selected PKC isozymes. PKC-α, -β1, -β2, -δ, and -ε all increased in the nucleus of ES-derived cardiac myocytes, compared with nuclei from undifferentiated cells. In both groups of cells, PKC-δ and -ε were mainly expressed at the nuclear level. The nuclear increase of PKC-α, -β1, and -β2 was due to a translocation from the cytosolic compartment. In contrast, the increase of both PKC-δ and PKC-ε in the nucleus of ES-derived cardiomyocytes occurred independently of enzyme translocation, suggesting changes in isozyme turnover and/or gene expression during cardiogenesis. No change in PKC-ζ expression was observed during cardiac differentiation. Opioid receptor antagonists prevented the nuclear increase of PKC-α, PKC-β1, and PKC-β2 and reduced cardiomyocyte yield but failed to affect the nuclear increase in PKC-δ and -ε. PKC inhibitors prevented the expression of cardiogenic genes and dynorphin B in ES cells and abolished their development into beating cardiomyocytes.