Interaction of hepatitis C virus-like particles and cells: a model system for studying viral binding and entry.
Hepatitis C virus-like particles (HCV-LPs) containing the structural proteins of HCV H77 strain (1a genotype) was used as a model for HCV virion to study virus-cell interaction. HCV-LPs showed a buoyant density of 1.17 to 1.22 g/cm(3) in a sucrose gradient and formed double-shelled particles 35 to 49 nm in diameter. Flow cytometry analysis by an indirect method (detection with anti-E2 antibody) and a direct method (use of dye-labeled HCV-LPs) showed that HCV-LPs binds to several human hepatic (primary hepatocytes, HepG2, HuH7, and NKNT-3) and T-cell (Molt-4) lines. HCV-LPs binding to cells occurred in a dose- and calcium-dependent manner and was not mediated by CD81. Scatchard plot analysis suggests the presence of two binding sites for HCV-LPs with high (K(d) approximately 1 microg/ml) and low (K(d) approximately 50 to 60 microg/ml) affinities of binding. Anti-E1 and -E2 antibodies inhibited HCV-LPs binding to cells. While preincubation of HCV-LPs with very-low-density lipoprotein (VLDL), low-density lipoprotein (LDL), or high-density lipoprotein (HDL) blocked its binding to cells, preincubation of cells with VLDL, LDL, HDL, or anti-LDL-R antibody did not. Confocal microscopy analysis showed that, after binding to cells, dye-labeled HCV-LPs were internalized into the cytoplasm. This process could be inhibited with anti-E1 or anti-E2 antibodies, suggesting that E1 and E2 proteins mediate HCV-LPs binding and, subsequently, their entry into cells. Altogether, our results indicate that HCV-LPs can be used to further characterize the mechanisms involved in the early steps of HCV infection.
Triyatni
Miriam
M
Liver Diseases Section, National Institute of Diabetes and DigestiveKidney Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA.
Saunier
Bertrand
B
Maruvada
Padma
P
Davis
Anthony R
AR
Ulianich
Luca
L
Heller
Theo
T
Patel
Arvind
A
Kohn
Leonard D
LD
Liang
T Jake
TJ
D015703:Antigens, CD / Q000378:metabolism; D006528:Carcinoma, Hepatocellular; D002460:Cell Line; D016174:Hepacivirus / Q000502:physiology*; D022781:Hepatocytes / Q000821:virology*; D006801:Humans; D008074:Lipoproteins / Q000494:pharmacology; D008561:Membrane Fusion; D008565:Membrane Proteins; D018613:Microscopy, Confocal; D013601:T-Lymphocytes / Q000821:virology*; D060225:Tetraspanin 28; D014407:Tumor Cells, Cultured; D014759:Viral Envelope Proteins / Q000378:metabolism; D014771:Virion / Q000502:physiology*
D015703:Antigens, CD; C554798:CD81 protein, human; C087035:E1 protein, Hepatitis C virus; D008074:Lipoproteins; D008565:Membrane Proteins; D060225:Tetraspanin 28; D014759:Viral Envelope Proteins; C091015:glycoprotein E2, Hepatitis C virus