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Developmental studies of the 43kD (ν1 ) proteins in Torpedo marmorata electrocyte
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MOLECULAR GENETICS OF CELL ADHESION DURING NEURONAL DEVELOPMENT IN DROSOPHILA Corey S. Goodman, Howard Hughes Medical Institute, Department of Biochemistry, University of California; Berkeley, 94720, USA
B-50/GAP43 IN NEURONAL DIFFERENTIATION AND PLASTICITY Verhaagen, J), Boonstra, A. 1, De Graan, P.N.E. x, Oestreicher, A.B#, Schotman, P?, Schrarna, L.H?, Margolis, F) and Gispen, W.H. 1' Division of Molecular Neurobiology, University of Utrecht ~ and Dept. of Neurosciences, Roche Inst. Mol. Biol., Nufley0 USA 2.
We are using molecular genetic and classical genetic approaches to study the mechanisms that underline how neuronal growth cones find and recognize their correct targets during development. To this end, we have been studying the expression and function of cell and substrate adhesion molecules in Drosophila. We cloned the genes encoding the three subunits of Drosophila laminin, and the genes encoding two different Drosophila cadherins, and have identified and characterized mutations in several of these genes. Second, we cloned the genes encoding four different surface glycoproteins, called fasciclin I, fasciclin II, fasciclin III, and neuroglian, which are dynamically expressed on different overlapping subsets of axon fascicles and glia during embryonic development. Two of these molecules (fascielin II and neuroglian) are part of the immunoglobulin superfamily and are related to vertebrate neural cell adhesion molecules. The other two proteins (Fasciclin I and III) are unrelated thus far to anything else in the data bank. Using transfection of cell lines and aggregation assays, we have shown that both faseiclin III and fasciclin I are homoDhilic adhesion molecules which appear to define new classes of adhesion molecules. We have identified mutations in the fasciclin I, fasciclin III, and neuroglian genes, and are presently characterizing their phenotypes to determine their function during neuronal development.
S173
The growth associated protein (GAP43)/B50, a presynaptic substrate of protein kinase C, is expressed at high levels during neuronal differentiation, development of the nervous system and axonal regeneration. The expression of this neuron-specific protein decreases during synaptogenesis. Differentiation of PC12 cells by NGF results in transtocation of the protein from cytosolic stores to the membrane of newly formed neurite extensions. The protein is associated with the inner membrane leaflet of growth cones. B-50 was used as a marker to study the development of the olfactory system examining both the expression of the B-50 mRNA and the protein in embryonal and neonatal rodents. Bulbectomy (central) or TX-100 lesioning (peripheral) of the olfactory epithelium results in a differential expression of B-50 and the olfactory marker protein characterizing two stages in the regeneration of this epithelium. Evidence that the degree of phosphorylation may codetermine the role of B-50 in neurite outgrowth will be discussed.
572 Developmental studies of the 43kD (~i) proteins In Torpedo marmorata electrocyte. H.O. NGHIEM, J. HILL, and J.P. CHANGEUX. Neuroblologle Mol~culaire, Instltut Pasteur, 75015 Paris. The 43kD (Vl) protein is a peripheral protein associated with the AchR (i:I) on the cytoplasmic face of the postsynaptlc membrane of adult Torpedo marmorata electrocyte. At an early developmental stage, the 43kD protein is detected malnly in cytosolie fractions, the AchR being already clustered on the ventral face of the cell. At a later stage, the 43kD protein is enriched in both cytosollc and particulate fractions and is mainly distributed on the ventral pole of the cell on its cytoplasmic face. In the mature electrocyte, the 43kD protein detected mainly in the partlcul~te fraction is associated with the AchR, on a codlstrlbutlon basis at the cytoplasmic face of the postsynaptlc membrane. The data have been interpreted in terms of a "43kD-AchR association" after the clustering of the AChR (J Cell Biol. 1989, 108, 127-139). The cytosollc and particulate 43kD proteins have been isolated and show an apparent M.W. of 44kD and 43kD respectively. They do not correspond to the two mRNA detected by Frail et al. (1988) and likely possess the C terminal structure derived from the longer messenger.
571
MOLECULAR GENETICS OF CELL ADHESION DURING NEURONAL DEVELOPMENT IN DROSOPHILA Corey S. Goodman, Howard Hughes Medical Institute, Department of Biochemistry, University of California; Berkeley, 94720, USA
B-50/GAP43 IN NEURONAL DIFFERENTIATION AND PLASTICITY Verhaagen, J), Boonstra, A. 1, De Graan, P.N.E. x, Oestreicher, A.B#, Schotman, P?, Schrarna, L.H?, Margolis, F) and Gispen, W.H. 1' Division of Molecular Neurobiology, University of Utrecht ~ and Dept. of Neurosciences, Roche Inst. Mol. Biol., Nufley0 USA 2.
We are using molecular genetic and classical genetic approaches to study the mechanisms that underline how neuronal growth cones find and recognize their correct targets during development. To this end, we have been studying the expression and function of cell and substrate adhesion molecules in Drosophila. We cloned the genes encoding the three subunits of Drosophila laminin, and the genes encoding two different Drosophila cadherins, and have identified and characterized mutations in several of these genes. Second, we cloned the genes encoding four different surface glycoproteins, called fasciclin I, fasciclin II, fasciclin III, and neuroglian, which are dynamically expressed on different overlapping subsets of axon fascicles and glia during embryonic development. Two of these molecules (fascielin II and neuroglian) are part of the immunoglobulin superfamily and are related to vertebrate neural cell adhesion molecules. The other two proteins (Fasciclin I and III) are unrelated thus far to anything else in the data bank. Using transfection of cell lines and aggregation assays, we have shown that both faseiclin III and fasciclin I are homoDhilic adhesion molecules which appear to define new classes of adhesion molecules. We have identified mutations in the fasciclin I, fasciclin III, and neuroglian genes, and are presently characterizing their phenotypes to determine their function during neuronal development.
S173
The growth associated protein (GAP43)/B50, a presynaptic substrate of protein kinase C, is expressed at high levels during neuronal differentiation, development of the nervous system and axonal regeneration. The expression of this neuron-specific protein decreases during synaptogenesis. Differentiation of PC12 cells by NGF results in transtocation of the protein from cytosolic stores to the membrane of newly formed neurite extensions. The protein is associated with the inner membrane leaflet of growth cones. B-50 was used as a marker to study the development of the olfactory system examining both the expression of the B-50 mRNA and the protein in embryonal and neonatal rodents. Bulbectomy (central) or TX-100 lesioning (peripheral) of the olfactory epithelium results in a differential expression of B-50 and the olfactory marker protein characterizing two stages in the regeneration of this epithelium. Evidence that the degree of phosphorylation may codetermine the role of B-50 in neurite outgrowth will be discussed.
572 Developmental studies of the 43kD (~i) proteins In Torpedo marmorata electrocyte. H.O. NGHIEM, J. HILL, and J.P. CHANGEUX. Neuroblologle Mol~culaire, Instltut Pasteur, 75015 Paris. The 43kD (Vl) protein is a peripheral protein associated with the AchR (i:I) on the cytoplasmic face of the postsynaptlc membrane of adult Torpedo marmorata electrocyte. At an early developmental stage, the 43kD protein is detected malnly in cytosolie fractions, the AchR being already clustered on the ventral face of the cell. At a later stage, the 43kD protein is enriched in both cytosollc and particulate fractions and is mainly distributed on the ventral pole of the cell on its cytoplasmic face. In the mature electrocyte, the 43kD protein detected mainly in the partlcul~te fraction is associated with the AchR, on a codlstrlbutlon basis at the cytoplasmic face of the postsynaptlc membrane. The data have been interpreted in terms of a "43kD-AchR association" after the clustering of the AChR (J Cell Biol. 1989, 108, 127-139). The cytosollc and particulate 43kD proteins have been isolated and show an apparent M.W. of 44kD and 43kD respectively. They do not correspond to the two mRNA detected by Frail et al. (1988) and likely possess the C terminal structure derived from the longer messenger.