Recently, we have developed a culture system in which rat hepatocytes dedifferentiate and proliferate and after the addition of EHS-gel redifferentiate. During both developmental stages HNF-4alpha2 mRNA was more abundant than HNF-4alpha1 mRNA. However, Western blot analysis using COS-7 cell-expressed HNF-4alpha1 and HNF-4alpha2 proteins as standards revealed that (i) HNF-4alpha2 protein was not expressed in dedifferentiated hepatocytes and (ii) either HNF-4alpha2 protein or a highly phosphorylated HNF-4alpha1 protein was the dominating isoform in redifferentiated hepatocytes. The changes in HNF4-isoform expression could not be mimicked by DMSO, suggesting them to be matrix specific. Furthermore, DMSO was less efficient than EHS-gel in reinducing liver-specific gene expression. EHS-gel overlay also led to reduction of ARP-1 DNA binding activity, while overall ARP-1 protein levels did not change. These results suggest that EHS-matrix overlay regulates the expression of different HNF-4alpha isoforms on a posttranscriptional level while ARP-1 DNA binding activity is regulated by posttranslational mechanisms.
Copyright 1999 Academic Press.