Identification and characterization of novel developmentally regulated proteins in rat spinal cord.

We previously used 2-dimensional (2-D) gel electrophoresis to identify novel proteins that may be involved in the genesis of the mammalian nervous system [1]. Several novel proteins that were up- or down-regulated coincident with neurogenesis and neuronal migration in rat neocortex were identified. To further investigate the expression of some of these developmentally regulated proteins during a comparable period in spinal cord development, 2-D electrophoresis is used to study their regulation in the crude membrane and soluble fractions of spinal cord at embryonic day 12 (E12) and embryonic day 21 (E21). This analysis indicates that 7 of the proteins that exhibited large changes in their synthesis in cerebral cortex between embryonic day 14 (E14) and embryonic day 21 (E21) demonstrate similar up- or down-regulation during spinal cord neurogenesis. However, two proteins are restricted in their expression or developmental regulation. One of these, 667-800, appears cortex-specific, while the up-regulation of protein SC.1 appears to be spinal cord specific. Several of these proteins also appear to be enriched in both the cortex and spinal cord relative to non-neural tissues (117, 162, 182, 310 [TOAD-64], 667-800) and may be neural specific. To further characterize its expression, one of these neural-specific, up-regulated proteins, TOAD-64 (protein 310) [2-4], is studied throughout embryonic and postnatal spinal cord development using peptide-specific polyclonal antibodies. As suggested by the 2-D gel analysis and the previously reported expression pattern in cerebral cortex [3], TOAD-64 is transiently expressed in postmitotic spinal cord neurons early in their development and sharply down-regulated after the second postnatal week. In the adult spinal cord, TOAD-64 expression is remarkably restricted to a subset of primary afferents to the spinal cord. This expression pattern, coupled with its recently discovered homology to two proteins implicated in axon pathfinding in the chick and nematode [5,3], suggests that TOAD-64 may have a fundamental role in axon pathfinding.