Morphology and ultrastructure of retrovirus particles

Abstract

Retrovirus morphogenesis entails assembly of Gag proteins and the viral genome on the host plasma membrane, acquisition of the viral membrane and envelope proteins through budding, and formation of the core through the maturation process. Although in both immature and mature retroviruses, Gag and capsid proteins are organized as paracrystalline structures, the curvatures of these protein arrays are evidently not uniform within one or among all virus particles. The heterogeneity of retroviruses poses significant challenges to studying the protein contacts within the Gag and capsid lattices. This review focuses on current understanding of the molecular organization of retroviruses derived from the sub-nanometer structures of immature virus particles, helical capsid protein assemblies and soluble envelope protein complexes. These studies provide insight into the molecular elements that maintain the stability, flexibility and infectivity of virus particles. Also reviewed are morphological studies of retrovirus budding, maturation, infection and cell-cell transmission, which inform the structural transformation of the viruses and the cells during infection and viral transmission, and lead to better understanding of the interplay between the functioning viral proteins and the host cell.

Keywords: Gag lattice; capsid protein shell; cryo-electron microscopy; cryo-electron tomography; envelop protein complex; retrovirus structure; sub-tomogram averaging.

Figure 1

Gag and retrovirus assembly. The cartoon depicts three stages in retrovirus morphogenesis: a partially assembled particle on the host cell’s plasma membrane, an immature particle that is composed of a paracrystalline Gag structure, and a mature virus particle that has a distinct core [13,20]. Gag is a polyprotein including matrix (MA), capsid (CA) and nucleocapsid protein (NC) domains. Env represents the trimeric envelope protein complex. The figure is adapted from two review papers [13,20].

Figure 2

Morphology of mature retrovirus virions. The cryo-EM images and cryo-ET cross sections of the viruses display heterogeneous morphology and size of the viral core in six retrovirus genera. RSV: Rous sarcoma virus [25], reprinted with permission from Elsevier; MMTV: mouse mammary tumor virus [51], amended with permission from American Society for Microbiology; MoMLV: Moloney murine leukemia virus [52]; Copyright (2005) National Academy of Sciences, U.S.A. HTLV-1: human T-cell leukemia virus type 1 [40]; HIV-1: human immunodeficiency virus type 1 [53], reprinted with permission from Elsevier; FV: Foamy virus [54]. The FV Gag protein is not processed into the classical orthoretroviral MA, CA and NC subunits during particle morphogenesis. The black arrowheads show regular Gag assemblies in the wild type virus [54]. Reprinted by permission from the journal.

Figure 3

Structure of Gag lattice within immature HIV-1. (A) Computational slice through a Gaussian-filtered tomogram containing immature HIV-1 particles treated with the protease inhibitor amprenavir [67]. The white arrow marks a slice through the CA layer illustrating the hexagonal lattice. Defects of the Gag lattice are evident in these particles. Figure courtesy of John Briggs. (B–D) Surface rendering of the reconstruction of immature HIV-1 particle [31]. Adapted by permission from the journal. (B) A cross section perpendicular to the membrane. RNP represents ribonucleoprotein complex. (C) Surface cut tangential to the membrane at a radius indicated by the black dash line in B, and looking down on the NTDs of the CA lattice. (D) Surface cut through the CTDs of the CA lattice. (E) Surface rendering of the CA organization in the HIV-1 (EMD-2706) and MPMV (EMD-2707) immature particles [67], viewed from the same position shown in C. The CTDs in both viruses are colored in orange, while the NTDs of CA in HIV-1 and MPMV is colored in blue and green respectively. The numbers represent two-fold, three-fold and six-fold symmetry axes. These figures are produced using UCSF Chimera [68].

Figure 4

Interactions stabilizing the immature HIV-1 CA lattice. (A) A diagram of CA molecule, indicating sequence of the helices in the NTD and CTD domains. CypA-BL represents Cyclophilin-A binding loop. The diagram was adapted from Figure 2a by Schur et al. [67]. (B) Cartoon that depicts CTD interactions. L7–8 represents the linker region between helix 7 and helix 8. The two-fold, three-fold and six-fold symmetry axes are indicated by a black parallelogram, triangle and hexagon respectively. (C) Cartoon that depicts NTD interactions. The elements that are at the equivalent positions and have the same color code in B represent the same CA molecule.

Figure 5

Mature HIV-1 capsid structure. (A) Cryo-EM reconstruction of recombinant A92E CA tubular assembly with (−12, 11) symmetry (EMD 5582, [69]). The hexamer rings of NTDs are shown in blue. The CTDs are shown in yellow. Figure courtesy of Peijun Zhang. (B) A schematic diagram showing interactions between the CA proteins in mature HIV-1 capsid. The hexagons represent NTD and the circles represent CTD. Connected elements in the same color are from the same CA molecule. H1, H2, H3, H9 and H10 represent helices 1, 2, 3, 9 and 10 respectively.

Figure 6

Comparison of structures of trimeric Env in the closed, pre-fusion and open, activated conformations. (A) Zoomed-in top views of the closed (left) and open (right) quaternary states derived from structures of trimeric HIV-1 Env in complex with VRC03 and 17b antibodies, respectively. The ribbons representing the central helices are in an identical position in both panels indicating that the location of the central density is approximately the same in closed and open quaternary conformations. B) Molecular models for the two conformations in (A) show how Env activation results in major rearrangements of gp120 location relative to the central gp41 stalk. The outward rotation of each gp120 protomer repositions the V1V2 loop (base of loop shown in red) from the center to the periphery and alters its position relative to the location of the V3 loop (base of loop shown in green). Reprinted by permission from Macmillan Publishers Ltd: Nat. Struct. Mol. Biol. [73], copyright (2013).

Figure 7

Virus-cell interactions and cell-cell transmission [153,154]. (A–B) A contact region between HIV-1-pulsed dendritic cells (DC) and CD4+ T cells (T) illustrating HIV-1 distribution within the virological synapses [153]. Reprinted by permission from the journal. (A) A ~200 nm thick tomographic slice obtained from a FIB-SEM reconstruction map. (B) Segmentation of the region boxed in A to indicate membrane contact and virus (red) location. (C) MLV moves along the outer surface of filopodial bridges toward target cells [154]. Fluorescently labeled viral particles (green, MLV Gag-CFP; white arrows) moving along filopodial bridges (red, mCAT1-YFP) correlated to single approximately 100 nm particles observed by SEM (black arrows). The boxed areas in the upper panels are magnified in the lower panels. Reprinted by permission from Macmillan Publishers Ltd: Nature Cell Biology [154], copyright (2007).