The trans-homophilic interaction of TAG-1 directs the position of YFP-Kv1.2 clusters along axons. A, light microscopic image of coculture of hippocampal neurons and HEK293 cells. Transfected neurons (green arrows) at 8 DIV were cocultured with transfected HEK293 cells (blue arrow) for 1 day. B, the Kv1.2, Kvβ2, TAG-1, and Caspr2 constructs used in the neuron/HEK293 coculture. C, coculture of neurons transfected with YFP-Kv1.2 (green), Kvβ2, and TAG-1 (red) and HEK293 cells transfected with TAG-1 and mCherry (blue). The coculture was fixed, permeabilized, and stained with an anti-TAG-1 antibody (red in merge). D, YFP-Kv1.2 and TAG-1 are highly clustered in the region of contact between an axon expressing YFP-Kv1.2, Kvβ2, and TAG-1 and a HEK293 cell expressing TAG-1 and mCherry. E, TAG-1 from HEK293 cells failed to cluster YFP-Kv1.2 along the contacting axons expressing YFP-Kv1.2, Kvβ2, and Caspr2. White arrowheads, axonal segments contacting with transfected HEK293 cells. White arrows, coclusters of YFP-Kv1.2 and TAG-1 along axons not contacting with any HEK293 cells. Scale bars, 100 μm in A and C, 20 μm in D and E.