In polarized cells, such as neurons, ankyrin-type proteins are the major molecules that link the actin-spectrin-based membrane cytoskeleton to the plasma membrane. In Drosophila the second ankyrin gene, Dank2, is exclusively expressed in neuronal cells. Similar to ankyrin genes in other organisms, the Dank2 gene generates several ankyrin protein isoforms by differential splicing. Here we report that in Drosophila, the short Dank2 protein isoform is restricted to neuronal cell bodies and is excluded from axons, whereas the long Dank2 isoforms are localized specifically to axons. Thus the long and short Dank2 protein isoforms are localized to complementary neuronal subdomains, demonstrating that in vivo the composition of the neuronal cortical cytoskeleton is highly polarized. We show that once polarization is established, it persists during later stages of Drosophila development. We also present genetic evidence that the absence of axonal Dank2 protein is lethal. (c) 2002 Elsevier Science (USA).