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RNA interference and single particle tracking analysis of hepatitis C virus endocytosis. | LitMetric

AI Article Synopsis

  • The study investigates the entry process of Hepatitis C virus (HCV) into liver cells (hepatocytes) and identifies essential cellular proteins involved in this infection.
  • Researchers utilized a targeted siRNA library to pinpoint 16 host genes related to clathrin-mediated endocytosis and other trafficking processes crucial for HCV entry and production.
  • The findings suggest that HCV interacts with cellular components during entry and highlights potential antiviral targets based on new cofactor identifications and imaging techniques for studying HCV infection.

Article Abstract

Hepatitis C virus (HCV) enters hepatocytes following a complex set of receptor interactions, culminating in internalization via clathrin-mediated endocytosis. However, aside from receptors, little is known about the cellular molecular requirements for infectious HCV entry. Therefore, we analyzed a siRNA library that targets 140 cellular membrane trafficking genes to identify host genes required for infectious HCV production and HCV pseudoparticle entry. This approach identified 16 host cofactors of HCV entry that function primarily in clathrin-mediated endocytosis, including components of the clathrin endocytosis machinery, actin polymerization, receptor internalization and sorting, and endosomal acidification. We next developed single particle tracking analysis of highly infectious fluorescent HCV particles to examine the co-trafficking of HCV virions with cellular cofactors of endocytosis. We observe multiple, sequential interactions of HCV virions with the actin cytoskeleton, including retraction along filopodia, actin nucleation during internalization, and migration of internalized particles along actin stress fibers. HCV co-localizes with clathrin and the ubiquitin ligase c-Cbl prior to internalization. Entering HCV particles are associated with the receptor molecules CD81 and the tight junction protein, claudin-1; however, HCV-claudin-1 interactions were not restricted to Huh-7.5 cell-cell junctions. Surprisingly, HCV internalization generally occurred outside of Huh-7.5 cell-cell junctions, which may reflect the poorly polarized nature of current HCV cell culture models. Following internalization, HCV particles transport with GFP-Rab5a positive endosomes, which is consistent with trafficking to the early endosome. This study presents technical advances for imaging HCV entry, in addition to identifying new host cofactors of HCV infection, some of which may be antiviral targets.

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Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2790617PMC
http://dx.doi.org/10.1371/journal.ppat.1000702DOI Listing

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