THE MODE OF ANCHORAGE TO THE CELL-SURFACE DETERMINES BOTH THE FUNCTION AND THE MEMBRANE LOCATION OF THY-1 GLYCOPROTEIN

The surface glycoprotein, Thy-1, when expressed by transfection in NG1 15/401L neural cells, inhibits their neurite outgrowth over astrocytes . We have investigated the role of the glycosylphosphatidylinositol an chor of Thy-1 in this inhibition. Hybrid molecules, in which the lipid anchor was replaced by polypeptide transmembrane domains, were expres sed by transfection. Lines expressing Thy-1 with the transmembrane and full cytoplasmic domains of NCAM-140, or with the transmembrane and t runcated cytoplasmic domain of CD8, were not inhibited in their abilit y to extend neurites. over astrocytes. Truncation of the cytoplasmic d omain of NCAM-140 to just two amino acids, however, produced a transme mbrane form of Thy-1 that, when expressed at high levels, inhibited ne urite outgrowth. All forms of Thy-1 were concentrated in clusters that occurred primarily on fine filopodia. In double transfectants express ing normal Thy-1 and Thy-1 with the full NCAM cytoplasmic tail, the cl usters of each form were separate, with no instances of the transmembr ane form being found within the clusters of lipid-anchored Thy-1. Thy- 1 with the two-amino-acid cytoplasmic domain of NCAM also occurred in clusters separate from those occupied by lipid-anchored Thy-1, but sub stantial 'invasion' of the clusters of normal Thy-1 by this transmembr ane construct occurred. We suggest that the ability of this hybrid pro tein to enter the lipid-anchored clusters enables it to activate the s ignalling pathways that normal Thy-1 uses. Thus the membrane anchor, i n targetting Thy-1 to different microdomains on the cell surface, dete rmines its ability to inhibit neurite outgrowth on astrocytes.