Cloning, expression, and alternative splicing of neogenin1 in zebrafish

Abstract

Caenorhabditis elegans UNC-40, Drosophila Frazzled, and vertebrate Neogenin and DCC constitute a subgroup of the immunoglobulin superfamily (IgSF). They possess four immunoglobulin-like domains and six fibronectin-type III repeats at the extracellular region, a single transmembrane region, and a ,300 amino-acid intracellular region. UNC-40, Frazzled and DCC can function in axon guidance as the receptor of Netrin (Cell Mol. Life Sci. 56 (1999) 62; Curr. Opin. Cell Biol. 10 (1998) 609). Neogenin binds to Netrin-1 with the same affinity as DCC in vitro (Cell 87 (1996) 175), and is expressed by neurons as they project axons (J. Cell Biol. 127 (1994) 2009), suggesting that it is also a DCC-like Netrin receptor. A zebrafish homologue of DCC (zDCC) is reported recently (Mech. Dev. 109 (2001) 105), but so far there is no report of zebrafish Neogenin. To elucidate a possible neural function of vertebrate Neogenin, we cloned and characterized a zebrafish homologue of neogenin, zneo1, and identified four alternative splice sites within it. In the adult, despite broad tissue distribution, our reverse transcription polymerase chain reaction and Northern analyses demonstrated the dominant expression of zneo1 mRNA in brain. We detected zneo1 mRNA in the embryos from 10 hpf onward and revealed its spatiotemporally regulated expression pattern in both neuronal and nonneuronal tissues by in situ hybridization. Our data showed that during early brain development, zneo1 mRNA was not only present in the proliferative ventricular zones but also in the domains of several first postmitotic neuron clusters when they extended axons. Alternative splicing generates several isoforms of zneo1. Most of them are developmentally regulated, showing distinct distribution in brain and other tissues.