TRANSACTIVATION BY HEPATITIS-B VIRUS X-PROTEIN IS PROMISCUOUS AND DEPENDENT ON MITOGEN-ACTIVATED CELLULAR SERINE THREONINE KINASES

The X protein of hepatitis B virus (HBV-X) can act as a transactivator of transcription but its mechanism of action remains obscure. We have analyzed HBV-X transactivation in several cell types using 13 unrelat ed viral and cellular promoters and found that transactivation is more or less apparent in most cell types and is promiscuous and unrelated to specific sequence motifs within the target promoters. In general, t hough, HBV-X appears to act on enhancer elements since HBV-X had no ef fect on a minimal promoter, whereas HBV-X was able to transactivate af ter insertion of an AP-1 minienhancer. Several lines of evidence exclu de the possibility that HBV-X interacts directly with the AP-1 enhance r or its binding proteins and suggest that the proximal target of HBV- X is peripheral to the transcription complex. This hypothesis is suppo rted by the observation that inhibition of serine/threonine kinases, w hich regulate AP-1 activity (phorbol ester down-regulation or staurosp orine inhibition of protein kinase C and a dominant negative mutant of Raf-1), blocked the ability of HBV-X to transactivate without affecti ng basal promoter activity. Furthermore, basal transcription from the AP-1-dependent promoter was increased by overexpression of protein kin ase C and Raf-1 but HBV-X was unable to further stimulate, indicating that these kinases act subsequently to HBV-X. These data suggest that transactivation by HBV-X is an indirect result of the activation of ce llular serine/threonine kinases including protein kinase C and Raf-1. This mode of action implies that HBV-X may affect other cellular proce sses, besides transcription, that are regulated by these kinases.