Replication and IRES-dependent translation are both affected by core coding sequences in subgenomic GB virus B replicons

The yield of G418-resistant Huh7 cell clones bearing subgenomic dicistronic GB virus B (GBV-B) is significantly affected by the insertion of a portion of the viral core gene between the GBV-B 5' untranslated region and the exogenous neomycin phosphotransferase selector gene (A. De Tomassi, M. Pizzuti, R. Graziani, A. Sbardellati, S. Altamura, G. Paonessa, and C. Traboni, J. Virol. 76:7736-7746, 2002). In this report, we have dissected this phenomenon, examining the effects of the insertion of core sequences of different lengths on GBV-B IRES-dependent translation and RNA replication by using experimental approaches aimed at analyzing these two aspects independently. The results achieved indicate that an enhancement of translation efficiency does occur and that it correlates with the length of the inserted core sequences. Interestingly, the insertion of these sequences also has a direct similar effect on the efficiency of replication of the GBV-B replicon. These results suggest that in GBV-B replicon RNA and potentially in the complete viral genome, the core coding sequences not only are part of the IRES but also take part in the replication process, independently of the presence of the corresponding whole protein.