The cloning and characterization of TOAD-64

Jane Elizabeth Minturn, Yale University.

This is an Open Access Thesis


During the development of the mammalian central nervous system, a pool of morphologically homogeneous, mitotically active progenitor cells in the neural tube gives rise to the enormously diverse population of postmitotic cells that assume the properties of neurons or glia. Postmitotic neurons elaborate processes that are involved in the migration of neurons to their adult positions as well as the elaboration of axonal and dendritic arbors that will ultimately form specific synaptic contacts. The generation of postmitotic neurons from precursor cells is a central differentiative event in development, but molecular markers of this event are few.Using 2D gel electrophoresis, proteins that are developmentally regulated over the course of neurogenesis were identified. One of these, TOAD-64 (Turned On After Division, 64kDa) is expressed by neurons immediately after their birth, and is dramatically down-regulated in the adult.The expression of the TOAD-64 protein and gene is coincident with initial neuronal differentiation and down regulated when the majority of axon growth is complete. TOAD-64 is up-regulated following neuronal induction of P19 and PC12 cells; it is re-expressed in adult spinal motor neurons following axotomy of the sciatic nerve; and is present on the lamellipodia and filopodia of growth cones.The gene encoding TOAD-64 shows sequence homology to the unc-33 gene of C. elegans, mutations in which lead to aberrations in axon outgrowth. A chick homolog (CRMP-62) of TOAD-64 has also recently been cloned, and appears to mediate growth cone collapse in response to collapsin, suggesting a similar role for TOAD-64 in the rat.Taken together these results suggest that TOAD-64 may be a central element in the processes underlying axonal outgrowth and pathfinding.