Asymmetric HIV-1 envelope trimers bound to one and two CD4 molecules are intermediates during membrane binding

Abstract

Human immunodeficiency virus 1 (HIV-1) infection is initiated by binding of the viral envelope glycoprotein (Env) to the cell-surface receptor CD4. Although high resolution structures of Env complexed with soluble domains of CD4 have been determined, the binding process is less understood on native membranes. Here, we apply cryo-electron tomography (cryo-ET) to monitor Env-CD4 interactions at membrane-membrane interfaces formed between HIV-1 and CD4-presenting virus-like particles. Env-CD4 complexes organized into clusters and rings, bringing opposing membranes closer together. Additionally, Env-CD4 clustering was dependent on capsid maturation. Subtomogram averaging and classification revealed that Env bound one, two, and finally three CD4 molecules, upon which Env adopted a partially open state. Our data indicate that asymmetric HIV-1 Env trimers bound to one and two CD4 molecules are detectable intermediates during virus binding to host cell membranes, which likely has consequences for antibody-mediated immune responses and vaccine immunogen design.

Structural characterization of HIV-1 Env heterotrimers bound to one or two CD4 receptors reveals intermediate Env conformations

“…These results provide further evidence of native-like soluble SOSIP Env trimers resembling their virion-bound counterparts 4,5 , both in the closed, prefusion conformation and in various CD4-bound conformations that adopt different conformations compared with unliganded Env trimers. Thus, this and the accompanying cryo-ET study 41 , together with previous Env structures, complete a description of the conformations of HIV-1 Env trimers at each stage of engaging CD4, starting with no bound receptors to the final conformation with three bound receptors (Fig, 5; Supplementary Movie 1).…”

“…We found that binding of one CD4 resulted in minor structural changes to a native-like soluble Env trimer in the closed, prefusion state; for example, we did not observe opening of any of the gp120 subunits of the trimer or the accompanying changes in the CD4-bound gp120 that result from CD4 associating with gp120 in CD4-bound open trimers [6][7][8][9][10] (in particular, changes resulting from insertion of Phe43CD4 into a gp120 hydrophobic cavity, which facilitates induced changes such as V1V2 displacement in CD4-bound gp120 subunits of fully-saturated open Env trimers [6][7][8]23,24 , were minor). By contrast, the one CD4-bound conformation of membrane-bound Env trimer revealed by cryo-ET/sub-tomogram averaging showed a partially open conformation in which the CD4-bound protomer appeared to undergo CD4-induced conformational changes 41 .…”

“…The ability to confirm single-particle soluble Env heterotrimer conformations that include residue-level details using lower-resolution Env trimer structures derived by cryo-ET under more physiological conditions 41 lends confidence to the proposed order of structural transitions induced by CD4 binding (Fig. 5; Supplementary Movie 1).…”

“…Cryo-electron tomography (cryo-ET) and sub-tomogram averaging was used to determine the conformations of membrane-bound Envs complexed with sub-stoichiometric numbers of membrane-bound CD4s 41 . We can therefore compare our higher resolution soluble CD4–soluble heterotrimer Env structures with structures of CD4-Env complexes investigated under more physiological conditions.…”

“…In addition, neither of these types of experiments included structural characterizations to examine the conformational effects of sub-stoichiometric CD4 interactions with individual Env trimers. Thus, our single-particle cryo-EM investigation of Env heterotrimers binding one or two CD4s, together with the accompanying cryo-ET visualization of the native HIV-1 virions and membrane-bound CD4 41 , adds to our knowledge of Env structures, which was previously limited to closed, prefusion Env conformations with either no bound CD4s or three CD4s bound to fully-saturated open Env trimers [2][3][4][5][6][7][8][9]41 .…”