Abstract
HIV-1 Env protein functions in the entry process and is the target of neutralizing antibodies. Its intrinsically high mutation rate is certainly one of driving forces for persistence/survival in hosts. For optimal replication in various environments, HIV-1 Env must continue to adapt and evolve through balancing sometimes incompatible function, replication fitness, and neutralization sensitivity. We have previously reported that adapted viruses emerge in repeated and prolonged cultures of cells originally infected with a macaque-tropic HIV-1NL4-3 derivative. We have also shown that the adapted viral clones exhibit enhanced growth potentials both in macaque PBMCs and individuals, and that three single-amino acid mutations are present in their Env V1/C2/C4 domains. In this study, we investigated how lab-adapted and highly neutralization-sensitive HIV-1NL4-3 adapts its Env to macaque cells with strongly replication-restrictive nature for HIV-1. While a single and two mutations gave a significantly enhanced replication phenotype in a macaque cell line and also in human cell lines that stably express either human CD4 or macaque CD4, the virus simultaneously carrying the three adaptive mutations always grew best. Entry kinetics of parental and triple mutant viruses were similar, whereas the mutant was significantly more readily inhibited for its infectivity by soluble CD4 than parental virus. Furthermore, molecular dynamics simulations of the Env ectodomain (gp120 and gp41 ectodomain) bound with CD4 suggest that the three mutations increase binding affinity of Env for CD4 in solution. Thus, it is quite likely that the affinity for CD4 of the mutant Env is enhanced relative to the parental Env. Neutralization sensitivity of the triple mutant to CD4 binding site antibodies was not significantly different from that of parental virus, whereas the mutant exhibited a considerably higher resistance against neutralization by a CD4-induced epitope antibody and Env trimer-targeting V1/V2 antibodies. These results suggest that the three adaptive mutations cooperatively promote viral growth via increased CD4 affinity, and also that they enhance viral resistance to several neutralization antibodies by changing the Env-trimer conformation. In total, we have verified here an HIV-1 adaptation pathway in host cells and individuals involving Env derived from a lab-adapted and highly neutralization-sensitive clone.
Highlights
HIV-1 Env protein consists of gp120 and gp41, which are cleaved and matured products of the gp160 precursor protein (Freed and Martin, 1995, 2013; Clapham and McKnight, 2002; Wilen et al, 2012)
Sequence analysis showed that 5R Env-gp120 contains two amino acid alterations, T138I in V1 domain and F275L in C2 domain (HIV-1NL4−3 amino acid numbering from start site M), compared to ScaVR (Figure 1) (Kamada et al, 2006)
These mutations spontaneously arose in adaptation processes in macaque cells of HIV-1mt clones derived from HIV-1NL4−3
Summary
HIV-1 Env protein consists of gp120 and gp, which are cleaved and matured products of the gp160 precursor protein (Freed and Martin, 1995, 2013; Clapham and McKnight, 2002; Wilen et al, 2012). Binding of gp120 to the co-receptor triggers large conformational changes of gp, and thereby induces virus-cell membrane fusion. In addition to functions in the entry process, Env is targeted by neutralizing antibodies (NAbs), because it is the only viral protein expressed on virion surface. NAbs are categorized by defining an epitope or epitope cluster on Env: CD4 binding site (CD4bs), CD4-induced epitope (CD4i), V1/V2, GlycanV3, silent face center, fusion peptide, subunit interface, and membrane-proximal external region (Benjelloun et al, 2012; Kwong and Mascola, 2018). In the adaptation process, it is biologically important for HIV-1 Env to appropriately balance the efficient viral entry via interaction with receptor/coreceptors and the escape from recognition of NAbs by masking epitopes
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