Abstract
Rhinoviruses (RVs) cause recurrent infections of the nasal and pulmonary tracts, life-threatening conditions in chronic respiratory illness patients, predisposition of children to asthmatic exacerbation, and large economic cost. RVs are difficult to treat. They rapidly evolve resistance, and are genetically diverse. Here, we provide insight into RV drug resistance mechanisms against chemical compounds neutralizing low pH in endo-lysosomes. Serial passaging of RV-A16 in presence of the vacuolar proton ATPase inhibitor bafilomycin A1 (BafA1) or the endo-lysosomotropic agent ammonium chloride (NH4Cl) promoted the emergence of resistant virus populations. We found two reproducible point mutations in the viral proteins 1 and 3 (VP1, VP3), A2526G (serine 66 to asparagine; S66N), and G2274U (cysteine 220 to phenylalanine; C220F), respectively. Both mutations conferred cross-resistance to BafA1, NH4Cl, and the protonophore niclosamide, as identified by massive parallel sequencing and reverse genetics, but not the double mutation, which we could not rescue. Both VP1-S66 and VP3-C220 locate at the interprotomeric face, and their mutations increase the sensitivity of virions to low pH, elevated temperature and soluble intercellular adhesion molecule-1 receptor. These results indicate that the ability of RV to uncoat at low endosomal pH confers virion resistance to extracellular stress. The data endorse endosomal acidification inhibitors as a viable strategy against RVs, especially if inhibitors are directly applied to the airways.
Murer, Luca