Given these circumstances we wanted to explore if BAs can be used to stimulate replication of wild type HCV genomes that have poor replication efficiency in cell culture. To this end we transfected Lunet G-Luc cells with subgenomic Con1 luciferase replicons carrying either wild type NS3 to NS5B proteins, a single REM in NS4B or the highly adapted replicon with mutations in NS3 and NS4B, designated ET. Interestingly, culturing cells until 72 h post transfection in the presence of 200 mM CDCA increased replication of the wild type luciferase replicon ca. 10-fold, the highly adapted ET genome ca. 50-fold and the K1846T-adapted replicon more than 200-fold. Next we explored if CDCA also stimulates replication of full length Con1 genomes with our without REMs and if this facilitates production of infectious viral progeny and quantification of infection events in cell culture. Similar results were obtained when we transfected these genomes into Lunet cells expressing the MAVS-GFP indicator of cellular HCV infection described by Jones et al. More specifically, these cells express a GFP with nuclear localization signal fused to the C-terminus of MAVS, which includes a mitochondrial localization signal and the cleavage site for the HCV NS3/4A protease. As a consequence, expression of the HCV NS3/4A protease results in cleavage of the MAVSGFP protein and subsequent re-localization of GFP from the mitochondria into the nucleus, which is a simple biomarker to determine if a cell expresses the NS3/4A protease. The impact of CDCA treatment on the number of HCV expressing cells was determined at 10 days after transfection, since the background levels of residual nuclear GFP, relocated after initial translation of NS3/4A from the transfected RNA was negligible at this time point. CDCA treatment increased Atractylodin the number of HCV expressing cells for the K1846T-adapted genome. In contrast, for the wild type genome no significant stimulation was observed, possibly due to the very low replication of this genome. To explore if CDCA facilitates infection of particles released after transfection of these genomes, we co-cultured Lunet cells transfected with these genomes with naive Lunet MAVS-GFP cells. In this setup detection of nuclear localized GFP in the latter cells indicates productive infection by viral progeny produced from the transfected Lunet cells. Interestingly, four days after initiation of the co-culture we observed a few Lunet-MAVS-GFP cells with nuclear localized GFP when co-cultured with cells transfected the wild type or the K1846T-adapted genome. At least in case of the K1846T mutant supplementation of CDCA conferred a moderate, but statistically significant, increase in the number of cells with nuclear GFP. These results indicate that particles produced in the presence of CDCA are infectious and that the moderate stimulatory effect of this BA on HCV replication likely facilitates detection of infected cells in the MAVS-GFP-based HCV infection bioassay. The interplay between Atractylenolide-III has attracted considerable scientific attention. This was primarily due to the clinical observation that high serum levels of these compounds correlated with poor response rates to IFN-based therapies. Previous reports have highlighted that at least replication of GT1 subgenomic replicons was increased by high doses of BAs. However, efforts to show a broader cross-genotype regulation of HCV replication by bile acids failed since except for GT2a no alternative and robust cell based HCV replication systems are available. As highly efficient JFH1-based GT2a replicons apparently did not respond to treatment with BAs it was unclear if the findings for GT1 replicons are more generally applicable for other viral strains. Likewise, it was unclear at what stage or stages of the viral replication cycle these molecules may influence HCV.