LOCALIZATION AND PROCESSING OF GLYCOSYLPHOSPHATIDYLINOSITOL-ANCHORED CATHEPSIN-D

We have investigated the effect of a glycosylphosphatidylinositol anch or on the distribution of the soluble lysosomal enzyme cathepsin D. On ly 10% of the chimeric protein (CD-GPI) could be detected on the plasm a membrane after transfection in CHO cells. Similarly to endogenous ca thepsin D, intracellular CD-GPI was detected in vesicular structures, suggesting that CD-GPI is targeted to lysosomes. CD-GPI is present as three forms with Mr 55, 50 and 37 kD which could correspond to the pre cursor, intermediate and mature forms of cathepsin D, respectively. CD -GPI was shown to be GPI anchored by differential extractability with Triton X-114 before and after phosphatidylinositol phospholipase C hyd rolysis. Intracellular CD-GPI is mainly substituted with oligosacchari des containing uncovered mannose 6-phosphate residues whereas these re sidues are covered in the cell surface precursor form of CD-GPI. Ammon iun chloride treatment reduces the lysosomal delivery of CD-GPI and in creases the cell surface expression of its precursor form. (C) 1995 Ac ademic Press, Inc.