Matrix-mediated changes in the expression of HNF-4 alpha isoforms and in DNA-binding activity of ARP-1 in primary cultures of rat hepatocytes

Recently, we have developed a culture system in which rat hepatocytes dedif ferentiate and proliferate and after the addition of EMS-gel redifferentiat e, During both developmental stages HNF-4 alpha 2 mRNA was more abundant th an HNF-4 alpha 1 mRNA. However, Western blot analysis using COS-7 cell-expr essed MNF-4 alpha 1 and HNF-4a2 proteins as standards revealed that (i) HNF -4 alpha 2 protein was not expressed in dedifferentiated hepatocytes and (i i) either HNF-4 alpha 2 protein or a highly phosphorylated HNF-4 alpha 1 pr otein was the dominating isoform in redifferentiated hepatocytes. The chang es in HNF4-isoform expression could not be mimicked by DMSO, suggesting the m to be matrix specific. Furthermore, DMSO was less efficient than EHS-gel in reinducing liver-specific gene expression. EHS-gel overlay also led to r eduction of ARP-I DNA binding activity, while overall ARP-1 protein levels did not change. These results suggest that EMS-matrix overlay regulates the expression of different HNF-4 alpha isoforms on a posttranscriptional leve l while ARP-1 DNA binding activity is regulated by posttranslational mechan isms. (C) 1999 Academic Press.