USE OF CHIMERIC F3-NCAM MOLECULES TO EXPLORE THE PROPERTIES OF VASE EXON IN MODULATING POLYSIALYLATION AND NEURITE OUTGROWTH

Differential splicing of VASE exon in the fourth immunoglobulin (Ig) d omain and attachment to the fifth Ig domain of alpha 2-8 linked sialic acid (PSA) both dramatically change, in opposite manner, Neural Cell Adhesion Molecule (NCAM) functional properties. Reciprocal patterns of VASE and PSA expression suggest that they might be mutually exclusive . Here, we tested whether informations conferring polysialylation resi de in NCAM-Ig domains 4 and 5 and the influence of the VASE exon encod ed sequence on this process. We also examined if the VASE sequence was still able to inhibit neurite outgrowth when presented out of its nor mal NCAM context. Constructs have been prepared encoding NCAM-Ig domai ns 4 (with or without the VASE exon) and 5 fused to the F3 molecule. S table clones expressing the chimeric molecules or wild type F3 were th en obtained in the AtT-20 cell line. Although the chimeric molecules w ere expressed on the cell surface none of them was bearing PSA. Thus, polysialylation cannot be conferred to proteins by addition of the NCA M-Ig domains 4 and 5 modular motif and in this molecular context, the VASE sequence is not influencing the process. These chimeric molecules , either expressed at the surface of RIN or COS cells or presented as soluble forms, were examined for their effect on neurite outgrowth. In all cases, the length of neurites of sensory neurons was significantl y reduced when grown in presence of the VASE containing chimera by com parison with the chimera without VASE or wild type F3. When neurons fr om NCAM knock-out mice were used for the assay, the VASE inhibition co uld not be detected. Thus VASE is able to act as a modular motif and N CAM expressed on neurons participates in transducing its effect.