Fig 1: Pk2 colocalizes with AnkG at the onset of neuronal polarity and AIS formation.(A to A?) Illustration of a stage 2 neuron at DIV1 with Pk2 (magenta) and AnkG (green) labeling. Pk2 is detected in several immature neurites (asterisks) together with AnkG. (B to B?) Illustration of a stage 2/3 neuron at DIV1 with Pk2 (magenta) and AnkG (green) labeling. (B to B?) High magnification from (B). Pk2 and AnkG are enriched in clusters in the nascent axon. Arrows point to the protein clusters. Note one neurite in (B) with some Pk2 and AnkG staining (asterisk). (C to C?) Illustration of a stage 3 neuron at DIV1 with Pk2 (magenta) and AnkG (green) labeling after treating the neurons with detergent extraction [Triton (Tx)] during fixation. Both Pk2 and AnkG labelings are maintained. (D) Illustration of a stage 3 neuron at DIV2 with Pk2 (magenta) and AnkG (green) labeling imaged by STED microscopy. Pk2 and AnkG colocalize in clusters or vesicles in the proximal (a) (bracket) and distal (b) (curly bracket) region of the axon. Linescans of (a) and (b) are shown illustrating the parallel profile of the proteins. (E to E?) Illustration of a stage 3 neuron at DIV2 with Pk2 (magenta), AnkG (green), and F-actin (phalloidin, cyan) labeling. Pk2 and AnkG define a cohesive and overlapping profile along the forming axon. Star in (E?) marks the growth cone. (F and G) Illustration of a stage 3 neuron at DIV2 with Pk2 (magenta) and TRIM46 (green) in the nascent axons. The linescans (G) as illustrated in (F) and (F') (asterisk) show only a partial overlap of Pk2 and TRIM46. All labeling done at least in triplicates. Scale bars, 8 µm (A and B), 4 µm (B', B?, and F to F?), 10 µm (C to C'), 8 µm (D), 2 µm (D, insets), and 5 µm (E to E?).