Activin a-induced HepG2 liver cell apoptosis: Involvement of activin receptors and Smad proteins

A balance between cell proliferation and apoptosis is important for regulat ing normal liver function. Proteins of the transforming growth factor-beta superfamily are known to be important mediators of apoptosis in the liver. In this study we demonstrate that activin A potently induces apoptotic cell death in a hepatoma cell line, HepG2 cells. To determine the roles of acti vin receptors and downstream signaling proteins in activin A-induced apopto sis in these cells, the activin signaling pathway was analyzed using the tr anscription of an activin-responsive reporter gene, p3TP-Lux, as an assay. Although individual activin receptors had little effect on transcriptional activity, coexpression of an activin type I receptor and a type II receptor significantly increased both basal and activin-induced transcriptional act ivation, with the combination of receptors IB and IIB being the most potent . Similarly, expression of individual Smad proteins had only a modest effec t on reporter gene activity, but the combination of Smad2 and Smad4 strongl y stimulated transcription. Activin signaling induced a rapid relocation of Smad2 to the nucleus, as determined using a green fluorescence protein-Sma d2 fusion protein. In contrast, green fluorescence protein-Smad4 remained l ocalized to the cytoplasm unless it was coexpressed with Smad2. In agreemen t with the transcriptional response assays, overexpression or suppression o f activin signaling components in HepG2 cells altered apoptosis. Overexpres sion of receptors IB and IIB or Smad proteins 2 and 4 stimulated apoptosis, whereas dominant negative mutant forms of the activin type IIB receptor or Smad2 blocked activin-stimulated apoptosis. These studies suggest that sig naling from the cell surface to the nucleus through Smad proteins is a requ ired component of the activin A-induced cell death process in liver cells.