Nuclear hormone receptors activate HBV replication in a nonhepatoma
cell line. (A) Structure of the HBV DNA (4.1 kbp)
construct used in transient transfection analysis. The 4.1-kbp
greater-than-genome length HBV DNA sequence in this construct spans
coordinates 1072–3182/1–1990 of the HBV genome (subtype
ayw). The locations of the HBV 3.5-, 2.4-, 2.1-, and
0.7-kb transcripts are indicated. EnhI/Xp, enhancer I/X-gene
promoter region; Cp, nucleocapsid or core promoter; pA, polyadenylation
site; PS1p, presurface antigen promoter; Sp, major surface antigen
promoter; X, X-gene; S, surface antigen gene; C, core gene; P,
polymerase gene. (B–E) Cells were transiently
transfected with the HBV DNA (4.1 kbp) construct and liver-enriched
transcription factors. Mouse NIH 3T3 fibroblasts (3T3), human
differentiated hepatoma cells (Huh7 and HepG2), and human
dedifferentiated hepatoma cells (HepG2.1) were used for this analysis.
(B and D) RNA (Northern) filter
hybridization analysis of HBV transcripts. The glyceraldehyde
3-phosphate dehydrogenase (GAPDH) transcript was used as an internal
control for RNA loading per lane. (C and
E) DNA (Southern) filter hybridization analysis of HBV
replication intermediates. HBV RC DNA, HBV relaxed circular DNA; HBV SS
DNA, HBV single-stranded DNA. All-trans retinoic acid and clofibric
acid at 1 μM and 1 mM, respectively, were used to activate the
nuclear hormone receptors RXRα and PPARα (+lig).