TOAD-64, A GENE EXPRESSED EARLY IN NEURONAL DIFFERENTIATION IN THE RAT, IS RELATED TO UNC-33, A C-ELEGANS GENE INVOLVED IN AXON OUTGROWTH

Using two-dimensional gel electrophoresis we previously identified mem brane-associated proteins that are upregulated over the course of neur ogenesis. One of these, TOAD-64 (Turned On After Division, 64 kDa), is expressed immediately after neuronal birth and is dramatically downre gulated in the adult. The gene encoding TOAD-64 has now been cloned, a nd its sequence shows homology to the unc-33 gene from C. elegans, mut ations in which lead to aberrations in axon outgrowth. Northern and in situ hybridization show that TOAD-64 mRNA is enriched in the nervous system and is developmentally regulated in parallel with the protein. The expression of the TOAD-64 protein and gene coincident with initial neuronal differentiation and the downregulation when the majority of axon growth is complete suggests a role in axon elaboration. Three add itional lines of evidence support this possibility: TOAD-64 is upregul ated following neuronal induction of pig and PC12 cells; the protein i s found in lamellipodia and filopodia of growth cones; and axotomy of the sciatic nerve induces reexpression. While the sequence of TOAD-64 lacks a signal sequence and therefore is likely to encode a cytoplasmi c protein, biochemical experiments demonstrate that the protein is tig htly, but noncovalently, associated with membranes. The data presented here suggest that TOAD-64 could be a central element in the machinery underlying axonal outgrowth and pathfinding, perhaps playing a role i n the signal transduction processes that permit growing axons to choos e correct routes and targets.