Shear-reversible clusters of HIV-1 in solution: stabilized by antibodies, dispersed by mucin

Abstract

ABSTRACT HIV-1 has eluded vaccine therapy for the past 40 years. The virus mutates rapidly and is protected by a shifting glycan shield of mannose sugars, which has hindered the broad neutralization of the virus by antibodies (Abs). Studies have shown that mannose residues are self-adhesive, but it is not known if these adhesions drive HIV-1 to aggregate in solution, further complicating Ab neutralization. The behavior of HIV-1 in culture media was monitored using Dynamic Light Scattering and complementary atomic force microscopy (AFM) imaging in the presence of anti-gp120 Abs, lectins, mannosidase, and mucin. After accounting for the serum contribution from the culture media, HIV-1 was found to be diffusing in solution in 400–700 nm clusters. These clusters could be sheared into single virus particles by filtration, but the dispersed particles clustered back within a short time frame. Sample preparation prior to AFM and transmission electron microscopy (TEM) imaging appears to disperse clusters, but the clusters become visible in AFM when they are stabilized by Abs in solution. The clustered form of the virus appears to restrict access of Abs, lectins, and glycosidases to surfaces within the cluster. Mannosidase treatment following virus dispersion by filtration prevented clustering, suggesting that the mannose glycan shield is involved in cluster formation. Dispersed HIV-1 particles that were bound by Abs did not re-cluster back. Free mucin molecules (porcine gastric mucin) effectively dispersed HIV-1 clusters, even those stabilized by Abs. HIV-1-loaded mucin dried on the AFM surface with a fern-like fractal pattern, similar to that seen clinically in cervical mucin during the more penetrable ovulation stage. IMPORTANCE The phenomenon of reversible clustering is expected to further nuance HIV immune stealth because virus surfaces can escape interaction with antibodies (Abs) by hiding temporarily within clusters. It is well known that mucin reduces HIV virulence, and the current perspective is that mucin aggregates HIV-1 to reduce infections. Our findings, however, suggest that mucin is dispersing HIV clusters. The study proposes a new paradigm for how HIV-1 may broadly evade Ab recognition with reversible clustering and why mucin effectively neutralizes HIV-1.