ER-LOCALIZATION AND FUNCTIONAL EXPRESSION OF THE HBV TRANSACTIVATOR MHBS(T)

In two recently reported cases, integrated hepatitis B virus (HBV) DNA s cloned from hepatocellular carcinoma were found to express a transcr iptional transactivator from 3'-terminally truncated HBV surface (preS /S) genes. In this study, we characterized the transactivator at the p rotein level. Expression of a 3'-truncated preS2/S gene in Spodoptera frugiperda (Sf9) insect cells resulted in a C-terminally truncated mid dle surface protein of 76 amino acids (MHBs(t76)), which was found to be associated with membranes of the endoplasmic reticulum and retained from Golgi processing and secretion, Accordingly, the microsome fract ion of MHBs(t76)-expressing Sf9 cells displayed transactivator activit y after electric field-mediated transfer into Chang liver cells. In co ntrast to full-length MHBs, MHBs(t76) is unglycosylated, and glycosyla tion is not required for transactivation as shown by mutation of the g lycosylation site at asparagine-4. Since highly purified MHBs(t76) der ived from an E. coli expression system also showed transactivator acti vity, it is concluded that unglycosylated MHBs(t76) protein is the aut hentic transactivating factor. As the transactivator protein derives f rom inactive MHbs by rearrangements of integrated HBV DNA, it may be i mportant for HBV-associated liver carcinogenesis.