Interestingly, infection of Huh-7 cells with such particles led us to isolate cellular clones exhibiting different levels of permissivity to HCVcc and HCVpp. For most of them, reduced HCV infection levels were directly Proteasome assay related to their reduced expression level of CD81, while

other entry molecules such as SR-BI and CLDN-1 were not modified. Our observation is in accordance with previously published data [29, 48–50]. Ectopic expression of CD81 in Huh-7w7 cells, one of the resistant cell clones, restored HCV permissivity indicating that CD81 deficiency alone was responsible for the resistance to HCV infection in these cells. In agreement with previous studies [29, 48, 51], we did not observe any variation in HCV genome replication in Huh-7w7 selleck chemical cells in comparison to Huh-7 cells (data not shown), suggesting that CD81 is not involved in this step of the viral cycle. Masciopinto et al. showed that CD81 and HCV envelope glycoproteins could be detected in exosomes of mammalian cells,

suggesting that HCV may intracellularly interact with CD81 allowing its export [52]. They pointed out a possible role of CD81 Adavosertib ic50 in assembly and release of HCV particles. However, our results indicate that CD81 does not participate to HCV assembly or release of new viral particles, since the supernatant of Huh-7w7 cells transfected with full-length HCV RNA infected naïve Huh-7 cells to a level comparable to that of the supernatant from transfected Huh-7 cells. Thus, Huh-7w7 cells constitute a new tool allowing to investigate the involvement of CD81 in HCV entry and offering a new single-cycle replication system, as already used by others [29]. The molecular determinants of HCV-CD81

interaction have been analyzed by several groups by using biochemical assays (reviewed in [53]). However, Flint et al have highlighted the limitation of these approaches new since various mutated CD81 sequences previously reported to abrogate E2-CD81 interaction, were able to restore permissivity in HepG2 cells [15]. In our study, we show that ectopic expression of human and mouse CD81 proteins in human hepatoma cells devoid of CD81 conferred susceptibility to infection by HCVcc and HCVpp at various levels. Interestingly, mCD81 protein supports infection by HCVcc and HCVpp bearing glycoproteins from genotypes 2a and 4 suggesting that, in accordance with other studies [15, 17], CD81 is not the sole determinant of species susceptibility to HCV. Other additional cellular factors likely modulate HCV entry. In addition, interaction/organization levels and stoichiometry between entry factors and plasma membrane lipids may regulate species susceptibility to HCV. CD81 belongs to the tetraspanin family of which members have the distinctive feature of clustering dynamically with numerous partner proteins and with one another in membrane microdomains.