siRNA-knockdown screening for ESCRT factors required for HBV replication in HepG2 cells.

<p>(A) A cartoon illustrates the ESCRT cascade and the interactions between each component members of ESCRT-0, I, II, III, and the VPS4 ATPase complex. Of note, the exact stoichiometry of certain component members remains tentative [<a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005123#ppat.1005123.ref005" target="_blank">5</a>]. (B) An HBV replicon plasmid containing a genomic dimer and individual siRNA against each ESCRT factor were co-transfected into HepG2 cells. Viral DNA replication, viral RNA expression and intracellular capsid particles were analyzed by Southern, Northern and Western blot analysis at day 5 post-transfection. The 28S rRNA was included as an RNA loading control. RC: relaxed circular DNA, DL: double-strand linear DNA, SS: single-strand DNA. We detected significant effect on HBV replication from 18 out of 30 tested ESCRT factors (as summarized in <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1005123#ppat.1005123.t001" target="_blank">Table 1</a>). Data shown here are representative of at least three independent experiments.