[Signal peptid peptidase and hepatitis C virus]. / Signal peptide peptidase et virus de l'hépatite C.

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[Hepatitis B virus morphogenesis]. / Morphogenèse du virus de l'hépatite B.

The human hepatitis B virus (HBV) is a small hepatotropic enveloped virus associated with chronic infection that can lead to cirrhosis and hepatocellular carcinoma. The HBV genome is a DNA molecule contained in an icosahedral capsid. Although HBV is not a retrovirus, the replication of its genome involves reverse transcription. Another distinctive feature of HBV is the production, in great excess over virions, of non-infectious subviral particles (SVP) consisting of membrane phospholipids and the three envelope proteins (small [S], medium [M] and large [L]). These empty non-infectious particles are highly immunogenic, and their in vitro production is at the basis of the current vaccine against hepatitis B. Despite numerous studies that lead to a better understanding of the HBV replication, little is known about the morphogenesis of the virion and its associated SVP. Recent approaches suggest that the mechanisms responsable for assembly of the virions and the SVP could be distinct.

Hepatitis C virus core protein, lipid droplets and steatosis.

Lipid droplets are intracellular organelles involved not only in lipid storage but also in cell signalling and the regulation of intracellular vesicular trafficking. Recent basic studies have suggested that interactions between hepatitis C virus (HCV) core protein and lipid droplets are required for the HCV infection cycle. In infected cells, the HCV core protein is associated with the surface of lipid droplets and the endoplasmic reticulum membranes closely surrounding these droplets, and its self-assembly drives virion budding. This interaction also seems to be directly linked to a virus-induced steatosis, which involves the deposition of triglycerides in the liver and contributes to the progression of fibrosis in patients with chronic hepatitis C. Many clinical studies have reported that virus-induced steatosis is significantly more severe with HCV genotype 3 than with other genotypes, and this phenomenon has been modelled in recent basic studies based on the production of HCV core proteins of various genotypes in vitro. The association of HCV core protein with lipid droplets seems to play a central role in HCV pathogenesis and morphogenesis, suggesting that virus-induced steatosis may be essential for the viral life cycle.

The genotype 3-specific hepatitis C virus core protein residue phenylalanine 164 increases steatosis in an in vitro cellular model.

BACKGROUND AND AIMS: The prevalence and severity of liver steatosis are higher in patients infected with genotype 3 hepatitis C virus (HCV) than in patients infected with other genotypes. HCV core protein is known to affect lipid metabolism, inducing lipid droplet accumulation both in vitro and in vivo. An in vitro cellular model was used to investigate whether an HCV core protein with residues specific to genotype 3 increased this phenomenon. METHODS: Sequence comparisons for HCV core protein domain II, which is known to interact with lipid droplets, identified the phenylalanine (F) residue at position 164 as the only residue specific to genotype 3. The area covered by lipid droplets in sections of cells producing a wild-type genotype 1a HCV core protein was compared with that in cells producing a Y164F mutant protein. RESULTS: Cumulative lipid droplet area was significantly greater in sections of cells producing the Y164F mutant HCV core protein than in cells producing the wild-type protein (p<0.001). The frequency of cell sections containing more than 3 mum(2) of lipid droplets, in particular, was higher for the mutant than for the wild-type protein. CONCLUSION: The data provide a molecular explanation for HCV genotype 3-specific lipid accumulation. This difference between genotypes may be due to phenylalanine having a higher affinity for lipids than tyrosine (Y). These observations provide useful information for further studies of the mechanisms involved in HCV-induced steatosis.

Identification of the glycoprotein 41(TM) cytoplasmic tail domains of human immunodeficiency virus type 1 that interact with Pr55Gag particles.

We investigated the protein/protein interactions that occur during human immunodeficiency virus (HIV-1) budding. We evaluated the binding to Pr55Gag particles of peptides mapping to the cytoplasmic tail of gp41TM and of host-cell proteins, in a cell-free, in vitro assay. Host-cell proteins and irrelevant viral envelope peptides did not bind. Peptides corresponding to a large central domain of the gp41TM cytoplasmic tail (93 residues) bound to Pr55Gag particles. This demonstrates that a Gag/Env interaction is responsible for the specific incorporation of the Env glycoprotein into nascent HIV-1 virions, and defines more accurately the gp41TM domain involved in this interaction.

DNA-containing and empty hepatitis B virus core particles bind similarly to envelope protein domains.

DNA synthesis within the hepatitis B virus (HBV) nucleocapsid appears to be coupled to nucleocapsid envelopment. The nature of the envelopment signal is unknown, but is thought to involve a conformational change at the surface of the capsid that facilitates interaction with HBV envelope proteins. In binding assays in vitro, it was found that empty HBV core particles bound synthetic peptides corresponding to HBV envelope protein domains with the same affinity as did HBV DNA-containing core particles. This suggests that the selection of replication-competent nucleocapsids for envelopment is not related to the capacity of DNA-containing core particles to bind specifically to HBV envelope proteins, and that there must be an alternative mechanism.

Interaction between hepatitis delta virus-encoded proteins and hepatitis B virus envelope protein domains.

Hepatitis delta virus (HDV) packaging requires prenylation of the HDV large protein (p27), as well as a direct protein-protein interaction between HDV proteins and hepatitis B virus (HBV) envelope protein domains. To investigate this interaction, we have analysed the binding capacity of baculovirus-expressed delta p24 and p27 proteins to synthetic peptides specific for the HBV envelope. Although a higher degree of binding was observed with p27, both p24 and p27 could bind HBV envelope peptides. One such peptide corresponded to residues 56-80 located in the cytosolic loop of the small HBV envelope protein, and another corresponded to 23 carboxy-terminal residues of the pre-S1 specific to the large HBV envelope protein. This indicates that in addition to p27, p24 may contribute to packaging of HDV through a protein-protein interaction with HBV envelope domains, and that an interaction between the pre-S1 polypeptide and delta proteins may play a role in infectivity.

Both pre-S1 and S domains of hepatitis B virus envelope proteins interact with the core particle.

The three envelope proteins of the hepatitis B virus (HBV) are encoded by a single open reading frame in the genome containing three separate in-phase AUG codons. This organization defines three protein domains (pre-S1, pre-S2, S) which form the small (S), middle (M, pre-S2/S), and large (L, pre-S1 /pre-S2/S) proteins. Mature virions are generated by the budding of preformed nucleocapsids through endoplasmic reticulum (ER) membranes containing S and L proteins, whereas the M protein is not necessary. This suggests an important function for the pre-S1 domain. To investigate the protein-protein interactions involved during the maturation process of the HBV virion, we studied in vitro the binding affinity to purified HBV core particles of various synthetic peptides identical to regions of the envelope proteins. Data previously obtained with deletion mutants were confirmed and refined. The 13 C-terminal amino acids of pre-S1 bound efficiently to core particles, whereas other pre-S domains did not. Moreover, the amino acid sequence 56-80 in the cytosolic loop of S bound efficiently to the HBV core. This double interaction between the HBV capside and both S and pre-S1 domains may be required for virion morphogenesis.