Flavivirus structural heterogeneity: implications for cell entry

Abstract

The explosive spread of Zika virus is the most recent example of the threat imposed to human health by flaviviruses. High-resolution structures are available for several of these arthropod-borne viruses, revealing alternative icosahedral organizations of immature and mature virions. Incomplete proteolytic maturation, however, results in a cloud of highly heterogeneous mosaic particles. This heterogeneity is further expanded by a dynamic behavior of the viral envelope glycoproteins. The ensemble of heterogeneous and dynamic infectious particles circulating in infected hosts offers a range of alternative possible receptor interaction sites at their surfaces, potentially contributing to the broad flavivirus host-range and variation in tissue tropism. The potential synergy between heterogeneous particles in the circulating cloud thus provides an additional dimension to understand the unanticipated properties of Zika virus in its recent outbreaks.

Figure 1. Schematics of flavivirus particles, with protein E in pastel colors (sand, brown and pink) and prM in bright colors (purple, blue and red).

A. Immature virion after budding in the ER (neutral pH) B. Immature virion after exposure to low pH in the TGN and rearrangement of envelope proteins C. Mature virion after secretion from infected cells

Figure 2. Flavivirus particle maturation and dynamics.

View down a 5-fold icosahedral axis of the particle, with the underlying viral membrane in green, and the prM/E protomers colored as in Fig. 1. There are three independent prM/E heterodimers in the icosahedral asymmetric unit, colored red/pink (by the 5-fold axes), blue/brown (2-fold axes) and purple/sand (3-fold axes). prM has a globular N-terminal half, which binds to the tip of domain II of E, and an extended C-terminal half. A) Immature flavivirus particle at neutral pH, displaying 60 (prM/E)₃ “tipi” shaped trimeric projections. This panel used the immature structure of DENV serotype 1 [5] (PDB code 4B03). B) Rearrangement at acidic pH into a lattice of 90 (prM/E)₂ dimers, completely hiding the viral membrane. The double arrow between panels A and B indicates that this change is reversible with pH. This panel was made with the immature structure of DENV serotype 2 at low pH [22] (PDB code 3C6R). C) The mature virion. Upon furin clavage, exposure to neutral pH leads to shedding of pr, leaving the virus particle in the same “herringbone” arrangement of E dimers as in the low pH immature particles. The arrow indicates that the change from B to C is irreversible. This panel used the structure of mature DENV serotype 2 [4] (PDB code 3j27). D. Partially mature mosaic particle resulting from incomplete prM cleavage in the TGN E. Illustration of breathing of a mature virion, using a “bumpy” particle as example F. Potential aspect of a partially mature virion after breathing in the mature side, further exposing the viral membrane.

Fig. 3. Changes in ZIKV prM in the strains circulating in the Americas compared to African strains.

A) Graphical representation of mutations in prM related to time and location of isolation. Changes in the ZIKV prM amino acid sequence over time was observed at seven positions, as indicated at the top of the figure. The results displayed are deduced from a comparison based on a ClustalO alignment (http://www.ebi.ac.uk/Tools/msa/clustalo/) of 210 complete prM sequences downloaded from the Virus Pathogen Resource (ViPR) database (www.viprbrc.org) on 4 January, 2017. Row 1: With a single exception, all 29 African strain sequences in the ViPR data base have identical amino acids at these positions. The exception is strain IbH-30656 with a valine at position 26. Row 2: The first Asian isolate from Malaysia 1966 (strain P6-740) differs from the African strains at 4 positions, indicated in blue. Position 21 was K in two deposits and E in two further deposits of the same strain, which may reflect different passaging histories. The same sequence with E at position 21 was recently submitted for an Indian strain. Row 3: Strain FSM from the outbreak in Yap Island 2007 is identical to 7 strains isolated from SE Asia since 2010, all of which differ from the Malaysia 1966 strain at position 31 (V31M). Row 4: All strains from the Pacific Islands since 2013 and the Americas (a total of 145 sequences) had the same sequence and differed from the Yap and Asian strains in Row 3 at position 17 (S17N). B) Structure of a trimer of prM-E heterodimers (PDB code 4B03). Color code is the same as in Fig. 1 and 2. The mutations observed in the ZIKV prM protein over time since its discovery in Uganda 1947 (see panel A) are displayed in yellow and labelled with arrows by their amino acid numbers.