Our aim is to define the role of cell surface glycoproteins (GP) in nervous system development and function. Work will continue on the purification and characterization of the principal antigenic cell surface and released GP of PC12 cells and sympathetic neurons, two homogeneous, well characterized model neuronal systems. A number of antigenic cells surface glycoproteins also present in brain have been identified on PC12 cells and sympathetic neurons including those at apparent Mr = 230, 180, 160, 140, 118 and 25 kd. We have studied the 230 kd component extensively and named it after its properties, the NGF-inducible large external (NILE) GP. Specific antibodies (Ab) have been prepared against the NILE and the 180 kd GP and monoclonal and monospecific polyclonal Ab will be produced against the others. The Ab will be coupled to insoluble matrices and for the immunoaffinity purification of each of these macromolecules. The Ab will also be used to identify cross-reactive proteins in brain which will be purified and compared to their membrane bound and released PC12 and sympathetic neuron counterparts. The polyclonal sera will be used in conjunction with trypsin-treatment of the intact cells and tunicamycin-inhibition of glycosylation to determine the structure of each of these components with respect to the cell membrane. The Ab will also be used in anatomical localization studies in cultured and in fixed CNS, PNS and adrenals of different developmental ages. in an attempt understand the role of these GP, the effects of the antisera and the purified GP will be assessed on a variety of neuronal behaviors. Immunological methods will be used to select mutants of PC12 cells which lack or are altered in specific cell surface GP and these will also be screened for functional changes. Transplacental exposure of embryos to Ab against the individual GP will be studied to understand the developmental role of these molecules. Finally a radioimmunoassay for the released GP will be developed for immunodiagnosis of neural crest tumors in animals.
