Tumor Suppressor P53 Inhibits Replication Of Hepatitis C Virus Subgenomic Replicon in Human Hepatoma Cells
[from browsing Digestive Week abstracts, if this can be accomplished in the clinic..diez casas en Barcelona<g>]
Authors: N. Dharel, N. Kato, H. Taniguchi, M. Otsuka, M. Moriyama, R. Muroyama, Y. Wang, R. Shao, T. Kawabe, M. Omata
Abstract ID: 524
Background & Aims: Chronic hepatitis C virus (HCV) infection is a major cause of liver cirrhosis and hepatocellular carcinoma worldwide. Host cellular factors regulating HCV replication are still to be elucidated.
Methods: 1) Naive Huh7 cells and Huh7-168 (high replication permissive Huh7 cells), were transfected with subgenomic HCV replicon and their replication permissiveness was compared. 2) p53 expression among high or low-replication permissive cells was examined by Western blot. 3) p53 expression in Huh7 cells non-permissive to HCV replicon was silenced by siRNA, and the p53 knocked down cells were transfected with HCV replicon. 4) p53 overexpressing plasmid or p53 silencing plasmid were co-transfected with HCV replicon into Huh7-168, and colony formation assay was performed. 5) p53 overexpressing plasmid was transfected into replicon harboring cells, and immunofluorescence staining of p53 and HCV NS5A was performed. 6) p53 mutant with point mutation at cell cycle regulating or apoptosis regulating residue was constructed, and their effect on HCV replication was compared to that of the wild type p53.
Results: 1) Numerous neomycin resistant colonies were obtained from Huh7-168 but no colony was obtained from naïve Huh7 cells. 2) Higher-replication supporting cells (confirmed by Western blotting for NS5B and real time RT-PCR) had a tendency of lower p53 expression. 3) Following p53 silencing, replication non-permissive naïve Huh7 cells supported a high level replication of HCV replicon. 4) p53 overexpression reduced the number of colonies, whereas p53 silencing increased. 5) HCV NS5A expression was low in cells overexpressing p53, whereas high expression of HCV NS5A was seen in p53 knocked down cells. 6) Mutant p53 with point mutation at cell cycle regulating or apoptosis regulating residue suppressed HCV replication as well as the wild type.
Conclusions: p53 inhibits HCV replication independent of cell cycle arrest and apoptotic function. Exploration of the mechanism of this hitherto unrecognized link between p53 and HCV could enhance the understanding of HCV replication, and could lead to the development of better therapeutic options. |
