Hepatitis C virus core protein uses triacylglycerols to fold onto the endoplasmic reticulum membrane

Abstract

Lipid droplets (LDs) are involved in viral infections, but exactly how remains unclear.Here, we study the hepatitis C virus (HCV) whose core capsid protein binds to LDs but is also involved in the assembly of virions at the endoplasmic reticulum (ER) bilayer. We found that the amphipathic helix-containing domain of core, D2, senses triglycerides (TGs) rather than LDs per se. In the absence of LDs, D2 can bind to the ER membrane but only if TG molecules are present in the bilayer. Accordingly, the pharmacological inhibition of the diacylglycerol O-acyltransferase enzymes, mediating TG synthesis in the ER, inhibits D2 association with the bilayer. We found that TG molecules enable D2 to fold into alpha helices. Sequence analysis reveals that D2 resembles the apoE lipid-binding region. Our data support that TG in LDs promotes the folding of core, which subsequently relocalizes to contiguous ER regions. During this motion, core may carry TG molecules to these regions where HCV lipoviroparticles likely assemble. Consistent with this model, the inhibition of Arf1/ COPI, which decreases LD surface accessibility to proteins and ER-LD material exchange, severely impedes the assembly of virions. Altogether, our data uncover a critical function of TG in the folding of core and HCV replication and reveals, more broadly, how TG accumulation in the ER may provoke the binding of soluble amphipathic helix-containing proteins to the ER bilayer.

Structural insights into perilipin 3 membrane association in response to diacylglycerol accumulation

“…To best visualize PLIN3 recruitment to ER membranes, we generated intracellular giant ER vesicles (GERVs) by submitting cells to hypotonic medium [66, 71, 72] (Fig. 6A) .…”

“…The cells were then incubated at 37°C, 5% CO2 for 5 minutes, to induce GERVs, and then they were visualized. Neutral lipids’ synthesis was triggered by the addition of OA and Z-stacks imaging of different entire cells was performed before and 15 min after OA administration, as described in [72]. For the DGAT1 (Sigma, PF-04620110) and DGAT2 (Sigma, PF-06424439) inhibitors used in Cos7, the dilution applied was 1/1000 for a final concentration of 3ug/mL.…”

Nup98 Is Subverted from Annulate Lamellae by Hepatitis C Virus Core Protein to Foster Viral Assembly

“…The infection of Huh7.5 cells by HCV (top right panel) induces the redistribution of AL to ER-derived remodeled membranes called membranous web (MW) ( 6 ), which is induced by HCV to achieve the viral replication, translation, and assembly within a protected environment. HCV core, probably through its high mobility on ER membranes ( 56 ), interacts with Nup98 from AL and translocates it within the MW and in close proximity of assembly sites, where it interacts with HCV RNA. In the absence of Nup98 (bottom left panel), the viral genome is excluded from the replication, assembly, and translation areas, leading to a decrease of infectious particle assembly.…”

The interplay between lipid droplets and virus infection

“…The interaction between the D2 domain of core protein and LDs is critical for HCV assembly efficiency 93,94 . Ajjaji et al showed that the D2 domain of core protein resembles the ApoE lipid‐binding region and senses TAG in the bilayer membranes 95 . TAG in LDs facilitates the folding of core protein, which relocalizes to the contiguous ER regions, and blockage of TAG synthesis impedes HCV assembly 95 .…”

“…Ajjaji et al showed that the D2 domain of core protein resembles the ApoE lipid‐binding region and senses TAG in the bilayer membranes 95 . TAG in LDs facilitates the folding of core protein, which relocalizes to the contiguous ER regions, and blockage of TAG synthesis impedes HCV assembly 95 . Two proline residues (P138 and P143) of core protein contribute to core‐LD interaction and core protein carrying P138A and P143A has no effect on LD morphology 96 .…”