Trends in Microbiology
Volume 17, Issue 11, November 2009, Pages 514-521
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Review
Dealing with low pH: entry and exit of alphaviruses and flaviviruses

https://doi.org/10.1016/j.tim.2009.08.002Get rights and content

The alphaviruses and flaviviruses include many important human pathogens, such as the dengue, West Nile, and Chikungunya viruses. These enveloped viruses infect cells by a membrane fusion reaction triggered by the low pH in endosomes. Fusion is mediated by viral membrane proteins through their acid-dependent conversion from a dimer on the virus surface to a homotrimer inserted into the host cell membrane. Here we review recent studies on the regulatory mechanisms that silence these fusion proteins during virus exit and that sense low pH and mediate protein refolding during virus entry. We discuss results using truncated proteins to dissect the fusion reaction, and future research directions including the development of antiviral therapies against these medically important viruses.

Section snippets

Virus entry: insights from the alphaviruses and flaviviruses

Enveloped viruses are surrounded by a membrane bilayer that protects the virus genome from the extracellular milieu and fuses with a host cell membrane to release the genome into the cytoplasm. This critical membrane fusion reaction is mediated by specialized transmembrane (TM) proteins known as virus membrane fusion proteins. Recent work has detailed the prefusion and postfusion structures of a number of virus fusion proteins (reviewed in Refs. 1, 2). While the structural features of such

Alphavirus and flavivirus organization and fusion protein structure

The alphavirus particle contains 240 copies of the capsid protein associated with the RNA genome to form a nucleocapsid with icosahedral symmetry (Figure 1a) [3]. This core is surrounded by a membrane containing 240 copies each of the transmembrane E2 and E1 glycoproteins, also organized with icosahedral symmetry. E2 and E1 associate as trimers of heterodimers (E2–E1)3 on the particle surface. E1 is the membrane fusion protein. It lies tangential to the virus membrane and forms an icosahedral

Virus assembly and maturation

The structural proteins of the alphaviruses and flaviviruses are synthesized as polyproteins that are co- and post-translationally processed by viral and cellular proteases. The alphavirus envelope proteins are translocated into the endoplasmic reticulum (ER) and transported through the secretory pathway to the plasma membrane, where virus budding occurs [3]. The E2 protein is synthesized as a precursor, termed p62, that associates with E1 and assists in its folding and transport. Based on the

Inhibition of alphavirus and flavivirus fusion and infection

Understanding the pre- and postfusion conformations of class I viral fusion proteins has led to inhibitors that block protein refolding during virus fusion (reviewed in Refs. 32, 33, 34). Here we will summarize strategies and progress to date on developing inhibitors for the class II fusion proteins.

The hinge region connecting DI and DII (Figure 2) undergoes important changes during flavivirus maturation (reviewed in Ref. [6]), and during class II protein refolding to the postfusion homotrimer

Control of low-pH-dependent fusion

Viral fusion proteins that are triggered by the acidic environment of endosomes have evolved specific mechanisms to respond to low pH. The term ‘pH sensor’ will be used here to denote regions or residues that, upon proton binding, destabilize the prefusion conformation to initiate fusion-protein refolding. Low pH can also act by titrating residues that are involved in forming or stabilizing the postfusion conformation, thus helping to drive the fusion reaction to completion. Important examples

Mechanistic studies of alphavirus E1 trimerization

Trimerization of the E1 or E protein is a crucial step in alphavirus and flavivirus fusion. Soluble E1 or E ectodomains can interact to form trimers, demonstrating that the interactions involved in trimerization are inherent to the proteins rather than requiring the specific geometry found on the virus particle. The structures of the alphavirus and flavivirus homotrimers show inter-chain interactions through contacts in DI and DII, with DIII clamping into the grooves of this central trimer

Concluding remarks and future directions

The alphavirus and flavivirus fusion proteins are assembled with companion proteins that protect them from acid pH during exocytic transport and that are processed by furin to permit fusion in the physiological pH range. During virus entry in the low-pH environment of endosomes, the E1 and E fusion proteins dissociate from their dimeric interactions, insert into the endosome membrane, trimerize, and refold to a hairpin conformation, thus driving membrane fusion (see model in Figure 4). As

Note added in proof

In contrast to the TBE studies discussed above, note that a recent study of West Nile virus did not support a role for any single histidine residue as an initial pH sensor [63].

Acknowledgements

We dedicate this review to the memory of our colleague and friend Dr. Dennis Shields. We thank all of the members of our lab for helpful discussions, and Gwen Taylor, Gleyder Roman-Sosa, Aihua Zheng, Kartik Chandran and Félix Rey for their comments on the manuscript. We also thank Mark Girvin for insightful discussions of amino acid titration. Work in our laboratory was supported by grants to M.K. from the National Institutes of Health (AI075647, AI067931, GM057454, U54AI057158) and by Cancer

Glossary

Ectodomain
the portion of a membrane protein that extends into the extracellular space.
In silico docking
a computational method to characterize the complementarity of a ligand and a protein target. It can be used to identify small molecules that bind to a defined region of a target protein structure.
Icosahedral symmetry
an icosahedron is composed of 20 triangular faces, contains 12 vertices, and has five-fold, three-fold, and two-fold axes of rotational symmetry. The organization of many virus

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