Visualizing knob structures at the external and internal surfaces.

(A) SEM of an intact CS2-infected RBC (~36 h post-invasion). Zoom shows a representative 1 x 1 μm section. (B) Schematic representation of KAHRP. Protein domains and features are as follows: black–signal sequence; orange–PEXEL; yellow–histidine-rich (His) repeats (amino acids 60–123), blue—5’ repeats (amino acids 329–420), green—3’ repeats (amino acids 506–555), ankyrin-binding domain (amino acids 282–361), spectrin-binding domain (amino acids 370–441) and the binding regions of anti-KAHRP (mAb89: amino acids 424–539 and mAb18.2: amino acids 282–362). Full length CS2 KAHRP has 634 amino acids. (C) dSTORM imaging of a sheared membrane preparation from CS2-infected RBCs (~30 h post-invasion) labelled with anti-KAHRP (mAb89) and anti-mouse Alexa-647 secondary antibody. Zoom shows a representative 1 x 1 μm section. Blue arrows: puncta. White arrows: ring structures. (D) The number of fluorescence events detected by dSTORM within 200 x 200 nm regions drawn around individual anti-KAHRP labelled puncta and ring structures. Counts are presented from 23 (color-coded) sheared membranes. (E) Sheared membranes prepared from CS2-infected RBC membranes at 30 h post-invasion imaged by SEM. Zoom shows a representative 1 x 1 μm section. White arrow: Dimpled disk-structure. (F) STORM-SEM showing overlay of physical dimpled disks and anti-KAHRP labelled ring structures (white arrow). Scale bars: (A,C, E, F) 500 nm.