Fig. 6: Detailed arrangement of the viral replication complex on nanorings revealed by super-resolution imaging.

a Schematic of expansion microscopy (ExM) for higher-resolution imaging. b Post-ExM images of a CHIKV-infected cell cultured on fluorescent gelatin-coated nanorings (cyan). The cell was stained for nsP1 (yellow) and dsRNA (magenta). A representative replication site is indicated by a white arrow. n = 3 repeats. c Zoomed-in x-y image (left) and resliced z-axis image along the white dashed line (right) show one nanoring with uniform gelatin coating (cyan). The CHIKV-infected cell exhibits dsRNA signals concentrated in the center of the inner ring (magenta), while nsP1 is located at the periphery of the dsRNA puncta near the nanoring rim (yellow). n = 3 repeats. d Schematic of a nanoring showing that the incomplete PM adhesion to the nanoring sidewall can create space at the bottom of the outer ring, allowing CHIKV replication in this region. e Post-ExM images of a CHIKV-infected cell cultured on fluorescent gelatin-coated nanorings (cyan). Incomplete PM adhesion, indicated by dark voids in the gelatin coating around the nanoring (example highlighted by a yellow ring), allows for viral replication within these spaces. The cell was stained for nsP1 (yellow) and dsRNA (magenta). n = 3 repeats. f Zoomed-in x-y image (left) and resliced z-axis image along the white dashed line (right) show replication occurring at the bottom of the outer ring. n = 3 repeats. g Illustration showing two distinct curvatures generated during spherule formation: 1) a saddle curvature at the neck of the spherule with increasing positive curvature along the x-z plane (pink), sensed by nsP1 via hydrophobic insertion of the MA loops, and a fixed negative curvature along the x-y plane (red) fixed by the size of the nsP1 dodecamer ring; and 2) a progressively increasing positive curvature along the crown of the spherule (blue). Scale bars: 2 μm. Source data are provided as a Source Data file.