Specific Roles for Lipids in Virus Fusion and Exit Examples from the Alphaviruses

Kielian, Margaret; Chatterjee, Prodyot K.; Gibbons, Don L.; Lu, Yanping

doi:10.1007/0-306-46824-7_11

Cited by 72 publications

(79 citation statements)

References 163 publications

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“…SFV and Sindbis virus fusion depends strictly on the presence of cholesterol and sphingolipids in the target membrane (55,61). By contrast the overall fusion of TBEV with liposomes is only facilitated by the presence of cholesterol in target membranes (29), is minimally affected by cholesterol depletion from cell membranes, and sphingolipids have only a weak fusion-facilitating effect (33).…”

Section: Discussionmentioning

confidence: 99%

Hepatitis C Virus Glycoproteins Mediate Low pH-dependent Membrane Fusion with Liposomes

Lavillette

Bartosch

Nourrisson

et al. 2006

Journal of Biological Chemistry

126

176

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It has been suggested that the hepatitis C virus (HCV) infects host cells through a pH-dependent internalization mechanism, but the steps leading from virus attachment to the fusion of viral and cellular membranes remain uncharacterized. Here we studied the mechanism underlying the HCV fusion process in vitro using liposomes and our recently described HCV pseudoparticles (pp) bearing functional E1E2 envelope glycoproteins. The fusion of HCVpp with liposomes was monitored with fluorescent probes incorporated into either the HCVpp or the liposomes. To validate these assays, pseudoparticles bearing either the hemagglutinin of the influenza virus or the amphotropic glycoprotein of murine leukemia virus were used as models for pH-dependent and pH-independent entry, respectively. The use of assays based either on fusion-induced dequenching of fluorescent probes or on reporter systems, which produce fluorescence when the virus and liposome contents are mixed, allowed us to demonstrate that HCVpp mediated a complete fusion process, leading to the merging of both membrane leaflets and to the mixing of the internal contents of pseudoparticle and liposome. This HCVpp-mediated fusion was dependent on low pH, with a threshold of 6.3 and an optimum at about 5.5. Fusion was temperature-dependent and did not require any protein or receptor at the surface of the target liposomes. Most interestingly, fusion was facilitated by the presence of cholesterol in the target membrane. These findings clearly indicate that HCV infection is mediated by a pH-dependent membrane fusion process. This paves the way for future studies of the mechanisms underlying HCV membrane fusion. Hepatitis C virus (HCV)2 belongs to the genus Hepacivirus of the Flaviviridae family (1) and has infected some 170 million people worldwide (2). In the majority of HCV-infected patients, the progression to chronic disease is associated with an increased risk of liver diseases and hepatocellular carcinoma. No vaccine is presently available, and the current treatment for chronic hepatitis C (a combination of pegylated interferon and ribavirin) has a limited efficacy (3). A more detailed understanding of the molecular mechanisms of virus entry would be beneficial to the development of novel therapeutic strategies.The HCV genome is a positive-stranded RNA encoding a precursor polyprotein of about 3,000 amino acids. This polyprotein is cleaved coand post-translationally to generate 10 viral proteins, classified into structural (capsid protein and the envelope glycoproteins E1 and E2) and nonstructural proteins (NS2 to NS5B), separated by the membrane ion channel polypeptide p7 (1, 4). The type I transmembrane envelope glycoproteins E1 and E2 form noncovalently linked heterodimers in the endoplasmic reticulum and are highly glycosylated, containing up to 6 and 11 potential glycosylation sites, respectively (5-8). Studies of the HCV life cycle have been hampered by the lack of an efficient and reliable cell culture system to produce and isolate the functional virus. Most re...

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Section: Discussionmentioning

confidence: 99%

Hepatitis C Virus Glycoproteins Mediate Low pH-dependent Membrane Fusion with Liposomes

Lavillette

Bartosch

Nourrisson

et al. 2006

Journal of Biological Chemistry

126

176

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“…hemolysin | intermedilysin | sterol | toxin | pore M embrane cholesterol is important to a variety of pathogenic processes that include virus fusion and budding (1) and the mechanisms of eukaryotic (2,3) and prokaryotic toxins (4)(5)(6)(7). Whether cholesterol is bound directly by these proteins as a receptor or it indirectly influences the binding or activity of the protein at the membrane surface remains unknown.…”

mentioning

confidence: 99%

Only two amino acids are essential for cytolytic toxin recognition of cholesterol at the membrane surface

Farrand

LaChapelle

Hotze

et al. 2010

Proc. Natl. Acad. Sci. U.S.A.

195

279

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The recognition and binding of cholesterol is an important feature of many eukaryotic, viral, and prokaryotic proteins, but the molecular details of such interactions are understood only for a few proteins. The pore-forming cholesterol-dependent cytolysins (CDCs) contribute to the pathogenic mechanisms of a large number of Grampositive bacteria. Cholesterol dependence of the CDC mechanism is a hallmark of these toxins, yet the identity of the CDC cholesterol recognition motif has remained elusive. A detailed analysis of membrane interactive structures at the tip of perfringolysin O (PFO) domain 4 reveals that a threonine-leucine pair mediates CDC recognition of and binding to membrane cholesterol. This motif is conserved in all known CDCs and conservative changes in its sequence or order are not well tolerated. Thus, the Thr-Leu pair constitutes a common structural basis for mediating CDC-cholesterol recognition and binding, and defines a unique paradigm for membrane cholesterol recognition by surface-binding proteins.M embrane cholesterol is important to a variety of pathogenic processes that include virus fusion and budding (1) and the mechanisms of eukaryotic (2, 3) and prokaryotic toxins (4-7). Whether cholesterol is bound directly by these proteins as a receptor or it indirectly influences the binding or activity of the protein at the membrane surface remains unknown. The cholesterol-dependent cytolysins (CDCs) use cholesterol as their receptor at the membrane surface (7) and contribute to the pathogenesis of a large number of Gram-positive bacterial pathogens (8). The CDC-sterol interaction initiates a cascade of secondary and tertiary structural changes that lead to the formation of a large oligomeric complex, and ultimately a pore in the membrane of eukaryotic cells (9-13). Although significant progress has been made in understanding the assembly of the CDC pore complex, the structural basis for recognition and binding to cholesterol-rich membranes remains elusive.Early studies with the Clostridium perfringens perfringolysin O (PFO) suggested that the highly conserved tryptophan-rich undecapeptide sequence at the base of domain 4 (14, 15) (Fig. S1) mediated the PFO-cholesterol interaction. However, recent studies by Soltani et al. (16) uncoupled cholesterol binding from the undecapeptide and showed that the membrane insertion of loops L1-L3 at the base of domain 4 was cholesterol dependent (Fig. S1). These observations are also consistent with a lack of conservation of the 3D structures of the undecapeptide in the closely related CDCs PFO (17) and Bacillus anthracis anthrolysin O (ALO) (18) (Fig. S1). These studies suggest the residues that comprise the cholesterol recognition motif are located within L1-L3 because these loops and the undecapeptide are the only structures at the tip of domain 4 exposed to the nonpolar bilayer core; the rest of the domain 4 surface is surrounded by water (19).Cholesterol was thought to function as the sole CDC receptor until the discovery of intermedilysin (ILY), a CDC fr...

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“…This feature is very likely to be involved in the conformational change that is necessary to trigger membrane fusion. Binding of a hydrophobic ligand is also known to be necessary for alphavirus membrane fusion, which specifically requires cholesterol and sphingolipids in the target membrane (12,13). No particular lipid requirement has been identified so far for flaviviruses, but a very recent report on West Nile flavivirus suggests that the conformational change can be triggered by interactions with liposomes even at neutral pH, depending on the particular lipid composition (14).…”

mentioning

confidence: 99%

Dengue virus envelope glycoprotein structure: New insight into its interactions during viral entry

Rey

2003

Proc. Natl. Acad. Sci. U.S.A.

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Specific Roles for Lipids in Virus Fusion and Exit Examples from the Alphaviruses

Cited by 72 publications

References 163 publications

Hepatitis C Virus Glycoproteins Mediate Low pH-dependent Membrane Fusion with Liposomes

Hepatitis C Virus Glycoproteins Mediate Low pH-dependent Membrane Fusion with Liposomes

Only two amino acids are essential for cytolytic toxin recognition of cholesterol at the membrane surface

Dengue virus envelope glycoprotein structure: New insight into its interactions during viral entry

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