Human hepatitis B virus X protein is detectable in nuclei of transfected cells, and is active for transactivation

Subcellular localization and transactivation of human hepatitis B virus X p rotein (HBx, a plausible causative factor for hepatocellular carcinogenesis , were studied in transiently transfected cells. The transactivation was de tected not only by the cis-element driven chloramphenicol acetyltransferase (CAT) assay but also by immunostaining of CAT protein cotransfected into h uman hepatoma cell line HepG2. Scanning fluorescence microscopy showed the majority of immunological signals of HBx to be at the perinuclear region of transfected cytoplasm, HBx was also clearly detectable in the nucleus. tho ugh less intensely expressed. This was confirmed by Western analysis and co immunoprecipitation of HBx with transcription factor IIB (TFIIB) in subcell ular fractionations. The percentage of HBx-positive cells coincided with th at of CAT-positive cells, and confocal laser microscopy revealed the coexis tence of CAT signals in GFP-HBx positive cells. The SV40 large T antigen nu clear localization signal (NLS) appended HBx, regardless of whether NLS was added to the N- or C-terminus, transactivated all the examined X-responsiv e elements (XRE) similarly as did wild-type HBx. Similar results were obtai ned in p53 negative Saos-2 cells. The detected nuclear HBx may be involved in modulating the transcription at the promoter level whereas the HBx in cy toplasm may be working through signal transduction pathways. (C) 1999 Elsev ier Science B.V. All rights reserved.