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 GTR1 embryonic stem (ES) cells expressed cell surface opioid receptors, as well as protein kinase C (PKC)-α, -ß₁, -ß₂, -δ, -ε, and -ζ. Cardiac differentiation was associated with a marked increase in the B_max value for a selective opioid receptor ligand and complex subcellular redistribution of selected PKC isozymes. PKC-α, -ß₁, -ß₂, -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, -ß₁, and -ß₂ 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-ß₁, and PKC-ß₂ 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.

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