Dataset Information

EWI-2wint promotes CD81 clustering that abrogates Hepatitis C Virus entry.

ABSTRACT: CD81 is a major receptor for Hepatitis C Virus (HCV). It belongs to the tetraspanin family whose members form dynamic clusters with numerous partner proteins and with one another, forming tetraspanin-enriched areas in the plasma membrane. In our study, we combined single-molecule microscopy and biochemistry experiments to investigate the clustering and membrane behaviour of CD81 in the context of cells expressing EWI-2wint, a natural inhibitor of HCV entry. Interestingly, we found that EWI-2wint reduces the global diffusion of CD81 molecules due to a decrease of the diffusion rate of mobile CD81 molecules and an increase in the proportion of confined molecules. Indeed, we demonstrated that EWI-2wint promotes CD81 clustering and confinement in CD81-enriched areas. In addition, we showed that EWI-2wint influences the colocalization of CD81 with Claudin-1 - a co-receptor required for HCV entry. Together, our results indicate that a change in membrane partitioning of CD81 occurs in the presence of EWI-2wint. This study gives new insights on the mechanism by which HCV enters into its target cells, namely by exploiting the dynamic properties of CD81.

SUBMITTER: Potel J

PROVIDER: S-EPMC7162402 | biostudies-literature | 2013 Jul

REPOSITORIES: biostudies-literature

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EWI-2wint promotes CD81 clustering that abrogates Hepatitis C Virus entry.

Potel Julie J Rassam Patrice P Montpellier Claire C Kaestner Laura L Werkmeister Elisabeth E Tews Birke A BA Couturier Cyril C Popescu Costin-Ioan CI Baumert Thomas F TF Rubinstein Eric E Dubuisson Jean J Milhiet Pierre-Emmanuel PE Cocquerel Laurence L

Cellular microbiology 20130216 7

CD81 is a major receptor for Hepatitis C Virus (HCV). It belongs to the tetraspanin family whose members form dynamic clusters with numerous partner proteins and with one another, forming tetraspanin-enriched areas in the plasma membrane. In our study, we combined single-molecule microscopy and biochemistry experiments to investigate the clustering and membrane behaviour of CD81 in the context of cells expressing EWI-2wint, a natural inhibitor of HCV entry. Interestingly, we found that EWI-2wint ...[more]

PMID: 23351194

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Project description:UnlabelledIn spite of the high variability of its sequence, hepatitis C virus (HCV) envelope glycoprotein E2 contains several conserved regions. In this study, we explored the structural and functional features of the highly conserved E2 segment from amino acid (aa) 502 to 520, which had been proposed as a fusion peptide and shown to strongly overlap a potential conserved neutralizing epitope. For this purpose, we used reverse genetics to introduce point mutations within this region, and we characterized the phenotypes of these mutants in the light of the recently published structure of E2. The functional analyses showed that their phenotypes are in agreement with the positions of the corresponding residues in the E2 crystal structure. In contrast, our data ruled out the involvement of this region in membrane fusion, and they indicate that alternative conformations would be necessary to expose the potential neutralizing epitope present in this segment. Of particular interest, we identified three specific mutations (Y507L, V514A, and V515A) located within this neutralizing epitope which only mildly reduced infectivity and showed no assembly defect. These mutations modulated HCV dependence on the viral receptor SRB1, and/or they also modulated virion sensitivity to neutralizing antibodies. Importantly, their characterization also showed that amino acids Y507, V514, and V515 contribute to E2 interaction with HCV receptor CD81. In conclusion, our data show that the highly conserved E2 segment from aa 502 to 520 plays a key role in cell entry by influencing the association of the viral particle with coreceptors and neutralizing antibodies.ImportanceHepatitis C virus (HCV) envelope proteins E1 and E2 exhibit sequence variability. However, some segments of the envelope proteins are highly conserved, suggesting that these sequences play a key role at some steps of the HCV life cycle. In this work, we characterized the function and structure of a highly conserved E2 region that is targeted by neutralizing antibodies and had been proposed as a fusion peptide. Our data ruled out the involvement of this region in membrane fusion but allowed for the identification of new residues modulating the interaction of the virus with entry factors and its sensitivity to neutralizing antibodies. Moreover, structural data suggest that alternative conformations could exist for E2, which would explain the presence of a partially masked neutralizing epitope in this segment in the currently available E2 structure. Overall, our findings highlight the importance of conserved regions in the sequences of HCV envelope proteins.

Dataset Information

EWI-2wint promotes CD81 clustering that abrogates Hepatitis C Virus entry.

Publications

EWI-2wint promotes CD81 clustering that abrogates Hepatitis C Virus entry.

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