Fig. 6.

Structural analysis of the mechanism of action of a small molecule that stimulates HN to trigger F. (A) Cells cotransfected with HN and F or transfected with F only were incubated in the presence of zanamivir alone to block HN-receptor binding, or with zanamivir together with CM9 or CSC11, two molecules that interact with HN to induce it to trigger F (Bottom-Tanzer et al., 2019). The cells were stained with monoclonal antibodies specific for the post-activated state of F. The proportions of activated F are shown as percentages of cells expressing post-triggered F on the y axis (±SD). (B) Overview of HPIV3 without and with PAC-3066, a next-generation small molecule that stimulates HN to activate F. Bars, 50nm (Marcink et al., 2020a). Lower panels show the viral surface glycoproteins without and with PAC-3066. (C) Cryo-electron 2D-subtomogram averages from a subset of viral surface glycoproteins in the absence of PAC-3066, where pre-fusion F can be identified (left) and with PAC3066, showing post-fusion F (Marcink et al., 2020a). Structural comparison coordinates of HN (PDB accession number 4MZA) and pre-fusion F (PDB accession number 6MJZ) are shown in pink and cyan, respectively. (D) Subtomogram averages where post-fusion F can be identified after PAC-3066 incubation. Structural comparison coordinates of the post-fusion F (PDB accession number 1ZTM) are shown in light blue. Panel (A): Adapted from Bottom-Tanzer, S.F., Rybkina, K., Bell, J.N., Alabi, C.A., Mathieu, C., Lu. M., et al., 2019. Inhibiting human parainfluenza virus infection by preactivating the cell entry mechanism. MBio 10 (1), with permission. Panels (B and C): From Marcink, T.C., Yariv, E., Rybkina, K., Mas, V., Bovier, F.T., des Georges, A., et al., 2020. Hijacking the fusion complex of human parainfluenza virus as an antiviral strategy, mBio 11 (1), with permission.