HCV particles circulate in association with lipoproteins and contain viral and host components. They enter cells by interacting with receptors involved in lipoprotein metabolism, including SR-BI, LDL-R, and GAGs. Lipid droplets aid primary HCV assembly. HCV hijacks the VLDL assembly and secretion pathway to exit cells and produce new viral particles.

1. HCV Life Cycle

2. HCV circulation in blood

3. HCV Virion Produced in infected patient
• Heterogeneous density (1.006-1.25 g/ml).
• Infectivity is inversely correlated to the density of
particles. (Andre P et al., 2002)
• The majority of viral RNA is associated with
density<1.06g/ml. (Andre P et al., 2002)
• The particles are irregular in shape and contain core,
E1,E2 (HCV) and ApoB, ApoE and ApoC1 (VLDL
derivatives).
Chang, K.S. et al., 2007; Nielsen, S.U. et al. 2006; Meunier, J.C. et al., 2008
HCV+Lipoprotein  LVPs

4. HCV Virion Produced in cell culture
• Higher density compare to HCV derived from patients
(1.1 vs. 1.06 g/ml).
• Transfer of HCVcc to chimpanzee ---> viral particles
with a lower density, similar to that of patient-derived
LVP---> again back to Huh7.5 cells --->Viral particles
with a higher density similar to HCVcc.
(Lindenbach, B.D. et al. (2006)
• The particles contain core, E1,E2 (HCV) and ApoE and
ApoC1 but not ApoB.
(Chang, K.S. et al., 2007; Merz, A. et al., 2010; Meunier, J.C. et al., 2008)

5. LVP and HCVcc
From: Bartenschlager et al., 2011

6. HCV Assembly: Core Protein
J. McLauchlan / Biochimica et Biophysica Acta 1791 (2009) 552–559
Core can be divided to two
domains D1, D2 and signal
peptide.
Core is released from HCV
polyprotein by SP and SPP and
translocates to cytosolic lipid
droplet (cLD)
D1 is hydrophilic positively
charge (attachment to negative
PO4 backbone of HCV RNA)
and faced to cytosol.
D2 is hydrophobic and anchors
D1 to a lipid membrane (cLD) .

7. HCV Assembly: Replication Complex, Core & cLD
From: Bartenschlager et al., 2011

8. Co-translational
lipidation of ApoB100
the main protein
component of VLDL by
MTP and formation of
poorly lipidated pre-
VLDL.
Formation of luminal
lipid droplets from
cytosolic lipid droplets
by possibly MTP. They
may contain ApoE.
Fusion of luminal lipid
droplets with pre-VLDL
and formation of mature
fully lipidated VLDL.
Taken from Ohsaki Y et al.,2009
VLDLAssembly Pathway

9. From: Bartenschlager et al., 2011
VLDL Assembly
and HCV Release
Inhibition of MTP, reduces the
HCV egress.
(Huang et al. 2007)
SiRNA down regulation of ApoB
reduces the secretion of HCV.
(Huang et al. 2007)
Cells were treated with brefeldin
A, produced high-density HCV
particles which were degraded.
(Gastaminza et al. 2008)
Dwon-reglation of ApoE reduces
the production of infectious HCV
particles
(Cheng et al. 2007)

10. Hepatitis C virus cell entry:
cellular receptors and role of lipoproteins.

11. Tetraspanin CD81
• 4 TMD, intracellular N- and C-terminal,
small and large extracellular loops (SEL, LEL).
• An HCV E2 binding region maps to the LEL of CD81, species –
specific.
(Pileriet al.,1998, Flint et al., 2006)
• Inhibition of HCV infection by mAbs directed against CD81 , a
soluble form of E2 (sE2) and down regulation of CD81 expression
using siRNA.
(Bartosch et al., 2003; Wakita et al.2005; Meuleman et al. 2008; Zhang et al., 2004)
• Activation of GTP-ases and the actin-dependent relocalization of the
E2/CD81 complex to cell–cell contact areas with tight junction
proteins occludin and CLDN-1
(Brazzoli et al.,2008)
SEL
LEL

12. Scavenger Receptor Class B Type I (SR-BI)
• 2 TMD, single extracellular loops.
• A receptor for various classes of Lipoproteins; HDL, LDL, VLDL,
LDLox. Important role in reverse cholesterol transport by CE
uptake from HDL.
• HCV E2 binds to SR-BI, species –specific. In this case SR-BI co-
operatively interacts with CD81 in HCV cell entry.
(Scarselli et al., 2002; Zeisel et al., 2007)
• Inhibition of HCV infection by mAbs directed against SR-BI . The
expression levels of SR-BI can modulate HCVcc infectivity.
(Grove et al., 2007; Catanese et al., 2007)
• Enhancement of SR-BI dependent HCV entry by HDL, and
inhibition by VLDL, LDLox..
(Bartosch et al.,2005; Maillard et al.,2006; Von Hahn et al.,2006; Lavie et.al, 2006)

13. Tight Junction Proteins:
Claudin 1 (CLDN-1)
• 4 TMD, intracellular loop N- and C-terminal, 2 extracellular loops (EL1
& EL2).
• EL1 seems to be involve in HCV entry but still no evidence for a direct
interaction between CLDN1 and the HCV virus.
(Evans et al., 2007)
• Inhibition of HCV entry to permissive cells (Huh7.5) by silencing
CLDN-1.
(Evans et al., 2007)
• Enhancing susceptibility of HCV infection in non-permissive cells
(293T) by CLDN-1 expression.
(Evans et al., 2007)
EL1

14. Tight Junction
Proteins: Occludin
• 4 TMD, intracellular loop N- and C-terminal,
2 extracellular loops (EL1 & EL2).
• Critical role for HCV entry. Interacts directly with HCV E2
(colocalization).
(Liu et al., 2009; Benedicto et al., 2008).
• Inhibition of HCV entry to permissive cells by SiRNA
silencing of Occludin.
(Liu et al., 2009)
• Human occludin renders murine cells infectable with
HCVpp.
(Ploss et al., 2009) Possibility for introducing immuno-competent
murine HCV model
EL1 EL2

15. LDL Rceptor (LDL-R)
• Cell surface receptor recognizes apoB100 of LDL and apoE of
LDL/Chylomicron remnant. It has been proposed to be involved in
HCV entry. (Agnello et al., 1999; Monazahian et al., 1999)
• Pros:
1. Association of HCV with lipid particles.
2. Inhibition of HCV (derived from plasma) entry (Cellular viral
RNA) by mAb against ApoB. (Molina et al.,2007)
3. Correlation between accumulation of cellular HCV RNA and
LDL-R mRNA level. (Molina et al.,2007)
• Cons:
1. The role of LDL-R in HCVcc has not yet been demonstrated
2. Lipoprotein lipase inhibits HCV cell entry while it should
enhance it. (Andreo et al., 2007)

16. Other Receptors: Glycosaminoglycans (GAGs)
• Primary, low affinity receptor for HCV entry. Binds to sE2 but not E1-
E2 heterodimer (?!).GAGs also is able to attach to lipoproteins.
(Callens et al., 2005)
• Heparan Sulphate (GAGs) inhibition by heparinase (degradation) or
heparin (competition) reduces HCV entry.
(Barth et al., 2006)
Other Receptors: DC-SIGN, L-SIGN
• DC-SIGN is expressed in Kuppfer cells, DC and lymphocytes.L-SIGN
in sinusoidal cells.
• Capture receptors, tissue tropism acts. (Cormier et al., 2004)

17. Summary
• HCV particles circulates in patient’s sera in
association with lipoproteins in particular VLDL
(LVPs) and contains ApoB, E and C1.
• Several receptors for HCV entry are also involved
in lipoprotein reuptake/metabolism; SR-BI, LDL-
R, GAGs.
• Lipid droplets act as a scaffold for primary HCV
assembly.
• VLDL assembly and secretion pathway is
essential for HCV secretion.