Abstract. Lectin affinity chromatography combined with mAb production was used to identify chick neural cell surface molecules related to L1 antigen, a mouse neural glycoprotein implicated in cell-cell adhesion (Rathjen, E G., and M. Schachner, 1984, EMBO (Eur. MoL Biol. Organ.)J., 3:1-10).A glycoprotein, G4 antigen, isolated by mAb G4 from adult chick brain is described which comprises a major 135-kD component, a minor doublet at 190 kD, and diffusely migrating bands at 80 and 65 kD in SDS PAGE. This molecule is structurally related to mouse L1 antigen according to NH2-terminal amino acid sequence (50% identity) as well as the behavior of its components in two-dimensional IEF/SDS PAGE gels. A second chicken glycoprotein, FU antigen, was isolated from adult chick brain using mAb Fll. This protein has also a major 135-kD component and minor components at 170 kD and 120 kD. Both immunotransfer analysis with polyclonal antibodies to mAb G4 and to mAb Fll isolate and the behavior on IEF/SDS PAGE gels indicates that the major 135-kD component of Fll antigen is distinct from G4 antigen components. However, the 135-kD component of FI1 antigen shares with G4 antigen and the neural cell adhesion molecule (NCAM) the HNK-1/L2 carbohydrate epitope.In immunofluorescence studies, I34 and Fll antigenie sites were found to be associated mainly with the surface of process-bearing cells, particularly in fiber-rich regions of embryonic brain.Although Fab fragments of polyclonal antibodies to rnAbs G4 or FU immunoaffinlty isolate only weakly inhibit the Ca2+-independent aggregation of neural cells, they strongly inhibit fasciculation of retinal axons. Together these studies extend the evidence that bundling of axons reflects the combined effects of a group of distinct cell surface glycoproteins.T un formation of specific connections in the nervous system during embryonic development is dependent on a variety of processes, including the adhesion of cells to each other and to extracellular matrices. In the last few years several neural cell surface glycoproteins have been identified which appear to be involved in adhesive interactions between neurons, glial cells, and matrices. Among these are the major Ca2+-independent cell-cell adhesion molecules, of which the known ones can be classified into two groups: the very closely related neural cell adhesion molecules (NCAMs) t (10, 39) and the L1 group of antigens (20,38,40).Mouse L1 antigen is biochemically and immunologically distinct from NCAM (13, 37). However, there is some functional similarity between the two molecules in that both anti-NCAM and anti-L1 Fab fragments inhibit the calciumindependent aggregation of cultured mouse cerebellar cells and neuroblastoma N2A cells (37,38). Moreover, the inhibition of aggregation of N2A cells by a combination of 1. Abbreviations used in this paper: NCAM, neural cell adhesion molecule; NgCAM, neuron-glia cell adhesion molecule; WGA, wheat germ agglutinin.anti-NCAM and anti-L1 is synergistic rather than additive (37). Anti-L1 Fab fragments ha...

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Abstract. The chick axon-associated surface glycoprotein neurofascin is implicated in axonal growth and fasciculation as revealed by antibody perturbation experiments. Here we report the complete cDNA sequence of neurofascin. It is composed of four structural elements: At the NH2 terminus neurofascin contains six Ig-like motifs of the C2 subeategory followed by four fibronectin type III (FNl/I)-related repeats. Between the FNIII-like repeats and the plasma membrane spanning region neurofasein contains a domain 75-amino acid residues-long rich in proline, alanine and threonine which might be the target of extensive O-linked glycosylation. A transmembrane segment is followed by a 113-amino acid residues-long cytoplasmic domain. Sequence comparisons indicate that neurofascin is most closely related to chick Nr-CAM and forms with L1 (Ng-CAM) and Nr-CAM a subgroup within the vertebrate Ig superfamily.Sequencing of several overlapping cDNA probes reveals interesting heterogeneities throughout the neurofascin polypeptide. Genomic Southern blots analyzed with neurofasein cDNA clones suggest that neurofascin is encoded by a single gene and its premRNA might be therefore alternatively spliced. Northern blot analysis with domain specific probes showed that neurofascin mRNAs of about 8.5 kb are expressed throughout development in embryonic brain but not in liver.Isolation of neurofasein by immunoaffinity chromatography results in several molecular mass components. To analyze their origin the amino-terminal sequences of several neurofascin components were determined. The NH2-terminal sequences of the 185, 160, and 110-135 kD components are all the same as the NH2 termini predicted by the eDNA sequence, whereas the other neurofascin components start with a sequence found in a putative alternatively spliced segment between the Ig-and FNIII-like part indicating that they are derived by proteolytic cleavage. A combination of enzymatic and chemical deglycosylation procedures and the analysis of peanut lectin binding reveals O-and N-linked carbohydrates on neurofascin components which might generate additional heterogeneity.

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