Antisense-induced reduction of presenilin 1 expression selectively increases the production of amyloid beta42 in transfected cells.
Autosomal dominant mutations in the presenilin 1 (PS1) gene are associated with familial, early-onset Alzheimer's disease. Although the pathogenic mechanism of these mutations is unclear, their common feature is that they lead to an increased concentration of amyloid beta-peptide (Abeta) 42 in the plasma of early-onset patients, in the conditioned media of transfected cells, and in the brains of transgenic mice that overexpress mutant PS1. To address the mechanism(s) by which the pathogenic PS1 mutations increase Abeta42, we constructed human cell lines expressing a doxycyclin (dox)-inducible antisense PS1 RNA and measured its effects on the levels of PS1, amyloid precursor protein (APP), and Abeta. In time course experiments, we observed a statistically significant (p = 0.0038) more than twofold elevation in secreted Abeta42 as early as 12 days after addition of dox. This correlated with an 80% decrease in the 46-kDa PS1 holoprotein and a 30% decrease in the 26-kDa N-terminal fragment (NTF). Furthermore, there was a significant fivefold (p = 0.002) increase in Abeta42 after 14-day dox treatment; this correlated with a >90% decrease in PS1 holoprotein and 60% decrease in NTF. At no time point did we observe significant changes in Abeta40, APP holoprotein, presenilin 2, or tubulin. Ten days after the removal of dox, we observed a return to constitutive levels for Abeta42, PS1 holoprotein, and NTF. These results suggest that in human cell lines, the reduction of normal PS1 activity results in the increased production of Abeta42. Furthermore, our results are consistent with a loss of function or dominant negative mechanism for the pathogenic PS1 mutations.
Refolo
L M
LM
Department of Pharmacology, Mayo Clinic Jacksonville, Florida 32224, USA.
Eckman
C
C
Prada
C M
CM
Yager
D
D
Sambamurti
K
K
Mehta
N
N
Hardy
J
J
Younkin
S G
SG
D000544:Alzheimer Disease / Q000235:genetics*; D016229:Amyloid beta-Peptides / Q000096:biosynthesis* / Q000235:genetics; D002460:Cell Line; D004318:Doxycycline / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D007668:Kidney; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000235:genetics; D053764:Presenilin-1; D016372:RNA, Antisense / Q000494:pharmacology*; D014162:Transfection; D014404:Tubulin / Q000096:biosynthesis / Q000235:genetics
D016229:Amyloid beta-Peptides; D008565:Membrane Proteins; C508853:PSEN1 protein, human; D010446:Peptide Fragments; D053764:Presenilin-1; D016372:RNA, Antisense; D014404:Tubulin; C075222:amyloid beta-protein (1-42); D004318:Doxycycline