Organization of cellular receptors into a nanoscale junction during HIV-1 adhesion

Abstract

The fusion of the human immunodeficiency virus type 1 (HIV-1) with its host cell is the target for new antiretroviral therapies. Viral particles interact with the flexible plasma membrane via viral surface protein gp120 which binds its primary cellular receptor CD4 and subsequently the coreceptor CCR5. However, whether and how these receptors become organized at the adhesive junction between cell and virion are unknown. Here, stochastic modeling predicts that, regarding binding to gp120, cellular receptors CD4 and CCR5 form an organized, ring-like, nanoscale structure beneath the virion, which locally deforms the plasma membrane. This organized adhesive junction between cell and virion, which we name the viral junction, is reminiscent of the well-characterized immunological synapse, albeit at much smaller length scales. The formation of an organized viral junction under multiple physiopathologically relevant conditions may represent a novel intermediate step in productive infection.

Figure 1. System detailing interaction of a viral particle with a flexible plasma membrane.

The plasma membrane was modeled using Cartesian coordinates to evaluate the positions of discrete points. The plasma membrane was also populated with primary, CD4 (red), and secondary, CCR5 (blue), cellular receptors for HIV-1. The viral particle was modeled as a rigid sphere with radius,  = 50nm, and was populated by gp120 trimers (green) capable of binding up to 3 CD4 and 3 CCR5 molecules. All entities were prohibited from diffusing through one another. The viral particle was allowed to move in 3 dimensions as well as rotate about its center, proteins were allowed to move in 2 dimensions along their respective surfaces and discrete plasma membrane points were allowed to move in 1 dimension either up or down. Movement directions within the plane here are shown with grey arrows.

Figure 2. Organization of proteins within the viral junction for fixed, evenly distributed viral trimers.

(A) Schematic describing the three main phases through which the organization of intermolecular bonds between a virion and cellular receptors on the plasma membrane evolves. (B) Probability distributions of cellular receptors bound to the virion for the three organizational phases as a function of the radial distance from the viral center. The steady state (phase III) bond probability distributions for bound CD4 and CCR5 receptors were time-averaged over the last 2.5×10⁸ iterations (Δt∼0.5s). Phase I and II probability distributions were time-averaged over 2×10⁷ iterations (Δt∼0.02s). (C and D) Three-dimensional projections of the steady state (phase III) probability distributions for bound CD4 (C) and CCR5 (D) receptors (origin is the viral center). (E) The number of bound CD4 (red) and CCR5 (blue) bonds in the viral junction increase with time as the system progresses through the three distinct phases (I, II, and III indicated by arrows) toward steady state. Error bars are depicted every 5×10⁷ iterations. (F) As the plasma membrane deforms and bonds begin to form, the height of the virion (, the change in position of viral center from initial cell contact) decreases, approaching a steady state. (G) A typical realization of the viral junction including a rigid sphere above a flexible plasma membrane with CD4/CCR5 bond locations projected beneath. (H) The plasma membrane of the system deforms and approaches the curvature of the partially engulfed virion at steady state. Shown here is a radial profile of the plasma membrane i.e., its vertical displacement as a function of the radial distance from the viral center. Simulations were performed with 15 gp120 trimers per virion and eight such simulations were averaged to produce the results illustrated here.

Figure 3. Example organization of the gp120 trimers facing the plasma membrane and resulting CD4 bond organization.

(A and B) Initial organization of bound and unbound gp120 trimers on the viral surface facing the plasma membrane (i) and the resulting CD4 bond organization on the cell surface (ii). Final organization of bound and unbound gp120 trimers on the viral surface facing the plasma membrane (iii) and the resulting CD4 bond organization on the cell surface (iv). Simulations were conducted using either 15 (A) or 20 gp120 trimers (B) per virion. The orientation used here is that of looking down along the y-axis through the virion (represented by the dashed circle) onto the plasma membrane.

Figure 4. Bond profiles of bound cellular receptors for different numbers of evenly distributed viral trimers.

(A–E) Example 2-D CD4/CCR5 bond probability distributions produced by viral particles with different numbers of gp120 trimers evenly distributed on the viral surface adhering to the cell membrane. As the number of gp120 trimers on the viral particles increases, the complexity of the bond probability distribution also increases.

Figure 5. The organization of the viral junction depends on the rigidity of the plasma membrane and the stability of the gp120-CD4 bond – the case of gp120 trimers diffusing on the viral surface.

(A–D) Two- and three-dimensional bond probability distributions of bound receptors when gp120 trimers are allowed to diffuse on the viral surface. The following illustrative cases for the organization of the viral junction are shown: (A) stable CD4/CCR5 molecules (Stable CD4) with plasma membrane rigidity of κ = 20 k_bT/nm, (B) stable CD4/CCR5 molecules on a completely flat surface (Hyper rigid PM), (C) CD4 molecules with an induced instability (Unstable CD4; see text for details) and plasma membrane rigidity of κ = 20 k_bT/nm and (D) stable CD4/CCR5 molecules on a rigid plasma membrane (Rigid PM, κ = 100 k_bT/nm). Bond probability distributions were time-averaged over the last 2×10⁷ iterations (Δt∼0.015s, n = 8). (E and F) The number of bound CD4 (E) and CCR5 (F) bonds increase with time and approach steady states. Conditions correspond to panels A–D, as indicated. (G) The engulfment of the virion can be measured by its change in height where is the change in position of viral center from initial cell contact. Conditions correspond to panels A–D, as indicated. (H) The steady state profile of the plasma membrane (the x-axis is measured in nm from the viral center, while the y-axis is the average variation of the membrane height from initialization of the system). Conditions correspond to panels A–D, as indicated. Simulations were performed with 15 gp120 trimers per virion and eight such simulations were averaged to produce the results displayed here.

Figure 6. Cellular membrane rigidity will determine the characteristic deformation of the membrane and directly affect the adhesion competent area on the cell.

(A–C) Schematic of area allowing for bond formation between a viral particle and cellular receptors located on a completely rigid plasma membrane (A), a flexible plasma membrane (B), or a comparatively more flexible cellular membrane (C). The shaded areas here illustrate the adhesion competent area (i.e. at an optimal distance for bonds to form) between the virion and the cell.

Figure 7. The organization of the viral junction depends on the rigidity of the plasma membrane, the stability of the gp120-CD4 bond, and the concentrations of cellular receptors – the case of fixed gp120 trimers.

(A–D) Two- and three-dimensional bond probability distributions of bound receptors CD4 and CCR5 for a fixed (evenly spaced) configuration of gp120 trimers. The following illustrative cases are shown: (A) stable CD4/CCR5 molecules (Stable CD4) with plasma membrane rigidity of, κ = 20 k_bT/nm, (B) stable CD4/CCR5 molecules on a completely flat surface (Hyper rigid PM), (C) CD4 molecules with an induced instability (Unstable CD4; see text for details) and plasma membrane rigidity of, κ = 20 k_bT/nm,, and (D) stable CD4/CCR5 molecules on a rigid plasma membrane (Rigid PM, κ = 100 k_bT/nm). Bond probability distributions were time-averaged over the last 2×10⁷ iterations (Δt∼0.015s, n = 8). (E and F) The number of CD4 (E) and CCR5 bonds (F) increase with time and approach a steady state. Conditions correspond to panels A–D, as indicated. (G) The engulfment of the virion can be measured by its change in height where is the change in position of viral center from initial cell contact. Conditions correspond to panels A–D, as indicated. (H) The steady state profile of the plasma membrane (the x-axis is measured in nm from the viral center, while the y-axis is the average variation of the membrane height from initialization of the system). Conditions correspond to panels A–D, as indicated. Simulations were performed with 15 gp120 trimers per virion and eight such simulations were averaged to produce the results displayed here.