PHOSPHORYLATION, SUBCELLULAR-LOCALIZATION, AND MEMBRANE ORIENTATION OF THE ALZHEIMERS DISEASE-ASSOCIATED PRESENILINS

Presenilins 1 and 2 are unglycosylated proteins with apparent molecula r mass of 45 and 50 kDa, respectively, in transfected COS-1 and Chines e hamster ovary cells. They colocalize with proteins from the endoplas mic reticulum and the Gels apparatus in transfected and untransfected cells. In COS-1 cells low amounts of intact endogeneous presenilin 1 m igrating at 45 kDa are detected together with relative larger amounts of presenilin 1 fragments migrating between 18 and 30 kDa. The preseni lins have a strong tendency to form aggregates (mass of 100-250 kDa) i n SDS-polyacrylamide gel electrophoresis, which can be partially resol ved when denatured by SDS at 37 degrees C instead of 95 degrees C. Sul fation, glycosaminoglycan modification, or acylation of the presenilin s was not observed, but both proteins are posttranslationally phosphor ylated on serine residues. The mutations Ala-246 --> Glu or Cys-410 -- > Tyr that cause Alzheimer's disease do not interfere with the biosynt hesis or phosphorylation of presenilin 1. Finally, using low concentra tions of digitonin to selectively permeabilize the cell membrane but n ot the endoplasmic reticulum membrane, it is demonstrated that the two major hydrophilic domains of presenilin 1 are oriented to the cytopla sm. The current investigation documents the posttranslational modifica tions and subcellular localization of the presenilins and indicates th at postulated interactions with amyloid precursor protein metabolism s hould occur in the early compartments of the biosynthetic pathway.