Intracellular generation and accumulation of amyloid beta-peptide terminating at amino acid 42.
Amyloid beta-peptide (Abeta) is known to accumulate in senile plaques of Alzheimer's disease (AD) patients and is now widely believed to play a major role in the disease. Two populations of peptides occur terminating either at amino acid 40 or at amino acid 42 (Abeta1-40 and Abeta1-42). Alternative N-terminal cleavages produce additional heterogeneity (Abetax-40 and Abetax-42). Peptides terminating at amino acid 42 are believed to be the major player in sporadic AD as well as familial AD (FAD). Whereas the cellular mechanism for the generation of Abeta terminating at amino acid 40 is well understood, very little is known about the cleavage of Abeta after amino acid 42. By using two independent methods we demonstrate intracellular Abeta1-42 as well as Abetax-42 but less Abetax-40 and Abeta1-40 in kidney 293 cells stably transfected with wild type beta-amyloid precursor protein (betaAPP) or the FAD-associated Val/Gly mutation. Moreover, retention of betaAPP within the endoplasmic reticulum (ER) by treatment with brefeldin A does not block the cleavage at amino acid 42 but results in an increased production of all species of Abeta terminating at amino acid 42. This indicates that the cleavage after amino acid 42 can occur within the ER. Treatment of cells with monensin, which blocks transport of (betaAPP) within the Golgi causes a marked accumulation of intracellular Abetax-42 and Abetax-40. Therefore these experiments indicate that the gamma-secretase cleavage of Abeta after amino acid 42 can occur within the ER and later within the secretory pathway within the Golgi. Moreover inhibition of reinternalization by cytoplasmic deletions of betaAPP as well as inhibition of intracellular acidification by NH4Cl does not block intracellular Abeta1-42 or Abetax-42 production.
Wild-Bode
C
C
Central Institute of Mental Health, Department of Molecular Biology, J5, 68159 Mannheim, Germany.
Yamazaki
T
T
Capell
A
A
Leimer
U
U
Steiner
H
H
Ihara
Y
Y
Haass
C
C
D016229:Amyloid beta-Peptides / Q000378:metabolism*; D020126:Brefeldin A; D002460:Cell Line; D003517:Cyclopentanes / Q000494:pharmacology; D004721:Endoplasmic Reticulum / Q000378:metabolism; D006801:Humans; D007668:Kidney / Q000378:metabolism / Q000648:ultrastructure
D016229:Amyloid beta-Peptides; D003517:Cyclopentanes; D020126:Brefeldin A