The proapoptotic effect of hepatitis B virus HBx protein correlates with its transactivation activity in stably transfected cell lines

UVDDB p127-binding modulates activities and intracellular distribution of Hepatitis B virus X protein

“…In agreement with the latter observations, transfection into CCL-13 cells of an episomic vector (pDR2) carrying the wt WHx gene, reduced e ciency of hygromycin-resistant colony formation, in a dosedependent way ( Figure 4A). We previously reported that in the WT1 cell line, constitutive expression of low level of active HBx results in increased susceptibility to apoptosis (Bergametti et al, 1999). Consistent with this ®nding, WHx-mediated toxic e ect was enhanced in WT1, as compared to CCL-13 ( Figure 4A).…”

“…Thus, a signi®cant alteration in UVDDBbinding e ciency (either markedly increased or markedly decreased interaction) was associated with loss of transactivation capacity. Since intracellular concentration of interaction partners is predicted to modulate formation of the corresponding protein complex, we next examined the e ect of UVDDB overexpression on the constitutive X transactivation activity shown by the WT1 cell line, which expresses low levels of active HBx (Bergametti et al, 1999). Transient overexpression of UVDDB stimulated expression of an X-responsive reporter, speci®cally in cells expressing active HBx ( Figure 3B).…”

“…H: histidine, 3-AT: 3-aminotriazole. Results obtained using the lacZ reporter gene (right) are expressed as in (B) (Bergametti et al, 1999;Chirillo et al, 1997;Kim et al, 1998;Su and Schneider, 1997) although both the underlying mechanism and the biological signi®cance of these e ects are still elusive. Moreover, in several studies, HBx overexpression has been shown to directly trigger apoptosis, resulting in strong suppression of colony formation (Chirillo et al, 1997;Kim et al, 1998).…”

“…This recipient cell line was then used to establish new stable cell lines expressing chimeric proteins consisting of a protein A IgG-binding domain (ZZ) fused to either WX1 (ZZ-WX1) or WX3 (ZZ-WX3). Crude protein extracts (lanes 1 and 2) or proteins speci®cally retained on rabbit IgG sepharose beads (lanes 3 and 4) were analysed by Western blotting using rabbit -polyclonal anti-FLAG, which simultaneously revealed UVDDB-FLAG and ZZ-WHx fusions via the FLAG antibody binding site and the IgG Fc moiety respectively (Bergametti et al, 1999), were cotransfected with a luciferase reporter gene driven by CRE regulatory elements (CRE-luc), and either empty pcDNA3 vector or pcDNA3-UVDDB. Shown are the results of two independent experiments expressed as normalized luciferase activity in WT1 relative to that in CCL-13 cells dicate that, in the case of wt WHx and WX7 cell survival requires strong counter-selection of viral protein expression.…”

“…In others, X appears to act far more upstream, by modulating the signal transduction cascade in the cytoplasm (Benn et al, 1996;Chirillo et al, 1996;Henkler et al, 1998;Kekule et al, 1993;Klein and Schneider, 1997;Lee and Yun, 1998;Su and Schneider, 1996). In addition to its widely documented transactivation activity, X has been reported to interfere with DNA repair (Becker et al, 1998;Wang et al, 1994), cell cycle control (Benn and Schneider, 1995) and apoptosis (Bergametti et al, 1999;Chirillo et al, 1997;Kim et al, 1998;Su et al, 1998). However, the functional connections between these activities and the cellular Xbinding proteins are still missing.…”