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. 2015 May 21;6(5):e1770. doi: 10.1038/cddis.2015.136

Figure 1.

Figure 1

HBV is able to induce autophagosome formation, which is required for replication of itself. (a) The effect of HBV transfection on LC3 accumulation in HepG2 cells. HepG2 cells were transfected with empty vector pUC19 or pHBV1.3. Forty-eight hours posttransfection, cells were subjected to western blotting using antibodies against LC3 or HBcAg. The expression of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as a loading control. (b) The effect of HBV transfection on autophagosome formation by FACS analysis in HepG2 cells. Cells were transfected with pUC19 or pHBV1.3. Forty-eight hours posttransfection, cells were first washed with phosphate-buffered saline containing 0.05% saponin and then incubated subsequently with anti-LC3 and FITC-labeled second antibody, followed by the FACS analysis. (c) Comparison of autophagosome formation in HepG2 with that in HepG2.2.15 cells by western blotting. HepG2 or HepG2.2.15 cells were subjected to western blotting using antibodies against LC3, HBc, and GAPDH as in panel (a). (d) Comparison of autophagy formation in HepG2 with that in HepG2.2.15 cells by FACS analysis. Cells were subjected to FACS analysis as in panel (b). (e and f) The effect of ATG5 or ATG7 siRNA on autophagome formation in HepG2 cells. HepG2 cells were electro-transfected with ATG5- or ATG7-specific siRNA using Amaxa Nucleofector technology. Forty-eight hours posttransfection, cells were further transfected with pHBV1.3; 48 h later, cells were ubjected to western blotting using antibodies against ATG5, ATG7 or LC3 (e), and HBV quantification was determined by quantitative real-time PCR, bars represent means±S.E.M (n=3); *P<0.05 (f). (g and h) The effect of ATG5 or ATG7 siRNA on autophagome formation and HBV replication in HepG2.2.15 cells. HepG2.2.15 cells were electro-transfected with ATG5- or ATG7-specific siRNA; 96 h posttransfection, cells were harvested. The expression of ATG5, ATG7 LC3 or GAPDH was determined by western blotting (g) and HBV DNA was quantified by real-time PCR, bars represent means±S.E.M (n=3); *P<0.05 (h)

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