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Journal of Virology, August 2002, p. 7736-7746, Vol. 76, No. 15
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.15.7736-7746.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Cell Clones Selected from the Huh7 Human Hepatoma Cell Line Support Efficient Replication of a Subgenomic GB Virus B Replicon

Amedeo De Tomassi, Maura Pizzuti, Rita Graziani, Andrea Sbardellati, Sergio Altamura, Giacomo Paonessa, and Cinzia Traboni*

Istituto di Ricerche di Biologia Molecolare P. Angeletti (IRBM), 00040 Pomezia (Rome), Italy

Received 30 January 2002/ Accepted 30 April 2002

Tamarins (Saguinus species) infected by GB virus B (GBV-B) have recently been proposed as an acceptable surrogate model for hepatitis C virus (HCV) infection. The availability of infectious genomic molecular clones of both viruses will permit chimeric constructs to be tested for viability in animals. Studies in cells with parental and chimeric constructs would also be very useful for both basic research and drug discovery. For this purpose, a convenient host cell type supporting replication of in vitro-transcribed GBV-B RNA should be identified. We constructed a GBV-B subgenomic selectable replicon based on the sequence of a genomic molecular clone proved to sustain infection in tamarins. The corresponding in vitro-transcribed RNA was used to transfect the Huh7 human hepatoma cell line, and intracellular replication of transfected RNA was shown to occur, even though in a small percentage of transfected cells, giving rise to antibiotic-resistant clones. Sequence analysis of GBV-B RNA from some of those clones showed no adaptive mutations with respect to the input sequence, whereas the host cells sustained higher GBV-B RNA replication than the original Huh7 cells. The enhancement of replication depending on host cell was shown to be a feature common to the majority of clones selected. The replication of GBV-B subgenomic RNA was susceptible to inhibition by known inhibitors of HCV to a level similar to that of HCV subgenomic RNA.

* Corresponding author. Mailing address: Istituto di Ricerche di Biologia Molecolare P. Angeletti (IRBM), Via Pontina Km 30.600, 00040 Pomezia (Rome), Italy. Phone: 39 06 91093241. Fax: 39 06 91093654. E-mail: cinzia_traboni{at}merck.com.

Journal of Virology, August 2002, p. 7736-7746, Vol. 76, No. 15
0022-538X/02/$04.00+0     DOI: 10.1128/JVI.76.15.7736-7746.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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