The hepatitis B virus X protein activates nuclear factor of activated T cells (NF-AT) by a cyclosporin A-sensitive pathway

The X gene product of the human hepatitis B virus (HBx) is a transcriptiona l activator of various viral and cellular genes. We recently have determine d that the production of tumor necrosis factor-alpha (TNF-alpha) by HBV-inf ected hepatocytes is transcriptionally upregulated by HBx, involving nuclea r factor of activated T cells (NF-AT)-dependent activation of the TNF-alpha gene promoter. Here we show that HBx activates NF-AT by a cyclosporin A-se nsitive mechanism involving dephosphorylation and nuclear translocation of the transcription factor. Luciferase gene expression assays demonstrated th at HBx transactivates transcription through NF-AT-binding sites and activat es a Gal4-NF-AT chimeric protein. DNA-protein interaction assays revealed t hat HBx induces the formation of NF-AT-containing DNA-binding complexes. Im munofluorescence analysis demonstrated that HBx induces the nuclear translo cation of NF-AT, which can be blocked by the immunosuppressive drug cyclosp orin A. Furthermore, immunoblot analysis showed that the HBx-induced activa tion and translocation of NF-AT are associated with its dephosphorylation. Thus, HBx may play a relevant role in the intrahepatic inflammatory process es by inducing locally the expression of cytokines that are regulated by NF -AT.