EXCHANGING THE EXTRACELLULAR DOMAIN OF AMYLOID PRECURSOR PROTEIN FOR HORSERADISH-PEROXIDASE DOES NOT INTERFERE WITH ALPHA-SECRETASE CLEAVAGE OF THE BETA-AMYLOID REGION, BUT RANDOMIZES SECRETION IN MADIN-DARBY CANINE KIDNEY-CELLS

Secretory processing and polarized sorting of horseradish peroxidase f used to the amyloid precursor protein transmembrane domain were compar ed with those of wild-type amyloid precursor protein in COS and polari zed Madin-Darby canine kidney (MDCK) cells. The cellular and secreted forms of the chimeric protein were enzymatically active in colorimetri c and cytochemical assays after reconstitution with hemin and Ca2+. Th e peroxidase enzyme was secreted by a proteolytic process, similar to the parent amyloid precursor protein. In polarized MDCK cells, amyloid precursor protein was secreted exclusively in the basolateral compart ment, while the peroxidase chimeric protein was secreted in both compa rtments. The basolateral sorting determinant for secretion must theref ore be located in the extracellular domain of amyloid precursor protei n, On the other hand, cell surface-associated peroxidase chimeric prot ein was similar to cell surface associated wild-type amyloid precursor protein, mainly expressed at the basolateral side. The basolateral ce ll-surface expression, in contrast to the basolateral secretion, is th erefore controlled by determinants in the cytoplasmic domain. Methylam ine inhibited and bafilomycin slightly increased the basolateral secre tion of both proteins, but both drugs strongly increased apical secret ion. The default secretory pathway of COS cells and the basolateral (b ut not the apical) secretory pathway of MDCK cells are therefore compa rably sensitive to methylamine and not to bafilomycin.