NEURONAL CHONDROITIN SULFATE PROTEOGLYCAN NEUROCAN BINDS TO THE NEURAL CELL-ADHESION MOLECULES NG-CAM L1/NILE AND N-CAM, AND INHIBITS NEURONAL ADHESION AND NEURITE OUTGROWTH/

We have previously shown that aggregation of microbeads coated with N- CAM and Ng-CAM is inhibited by incubation with soluble neurocan, a cho ndroitin sulfate proteoglycan of brain, suggesting that neurocan binds to these cell adhesion molecules (Grumet, M., A. Flaccus, and R. U. M argolis. 1993. J. Cell Biol. 120:815). To investigate these interactio ns more directly, we have tested binding of soluble I-125- neurocan to microwells coated with different glycoproteins. Neurocan bound at hig h levels to Ng-CAM and N-CAM, but little or no binding was detected to myelin-associated glycoprotein, EGF receptor, fibronectin, laminin, a nd collagen IV. The binding to Ng-CAM and N-CAM was saturable and in e ach case Scatchard plots indicated a high affinity binding site with a dissociation constant of similar to 1 nM. Binding was significantly r educed after treatment of neurocan with chondroitinase, and free chond roitin sulfate inhibited binding of neurocan to Ng-CAM and N-CAM. Thes e results indicate a role for chondroitin sulfate in this process, alt hough the core glycoprotein also has binding activity. The COOH-termin al half of neurocan was shown to have binding properties essentially i dentical to those of the full-length proteoglycan. To study the potent ial biological functions of neurocan, its effects on neuronal adhesion and neurite growth were analyzed. When neurons were incubated on dish es coated with different combinations of neurocan and Ng-CAM, neuronal adhesion and neurite extension were inhibited. Experiments using anti -Ng-CAM antibodies as a substrate also indicate that neurocan has a di rect inhibitory effect on neuronal adhesion and neurite growth. Immuno peroxidase staining of tissue sections showed that neurocan, Ng-CAM, a nd N-CAM are all present at highest concentration in the molecular lay er and fiber tracts of developing cerebellum. The overlapping localiza tion in vivo, the molecular binding studies, and the striking effects on neuronal adhesion and neurite growth support the view that neurocan may modulate neuronal adhesion and neurite growth during development by binding to neural cell adhesion molecules.