Aph-1 functions as a substrate docking site within the Alzheimer's disease relevant gamma -secretase complex

2009 2009

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Abstract (summary)

γ-secretase is an intramembrane protease complex composed of presenilin, nicastrin, Aph-1, and Pen-2 that together cleave an ever increasing list of type I transmembrane proteins. Cleavage of one substrate, the Amyloid Precursor Protein (APP), generates the AP peptide which plays a critical role in the pathogenesis of Alzheimer's disease (AD). In this regard, γ-secretase has emerged as a promising therapeutic target for the treatment of AD; however, the exact mechanism of how γ-secretase cleaves and the function of APP processing remain unclear. My thesis was focused on these two important issues.

In the first part of my thesis, I wanted to determine which component within γ-secretase functioned as a substrate docking site. I identified Aph-1 as a potential candidate as it associated with both full-length and ectodomain-shed substrates. This was observed in overexpressed systems and validated under endogenous expression levels in cell lines and animal tissues with two unrelated γ-secretase substrates, APP and Jagged-1.

Next, I performed mutagenesis on Aph-1 to identify the regions within the protein that may play a critical role in the binding of substrates and association with the other γ-components. I performed both truncation and domain-swapped mutagenesis, where the transmembrane domains of Aph-1 were replaced with that of an unrelated protein. While these techniques have worked successfully with Presenilin, all Aph-1 mutants generated were unstable or failed to incorporate into γ-secretase.

In the final part of my dissertation, I examined the putative function of the APP intracellular domain (AICD) in transcriptional regulation. A Notch-AICD chimera was generated to allow for inducible generation of AICD via addition of the Notch ligand, Delta, or calcium depletion. As a control for AICD regulated transcription, I examined the Aβ degrading enzyme, neprilysin. Using the same cell lines and techniques employed by the original authors that identified this target, I was unable to reproduce their findings. In light of my negative data, multiple independent reports have also been unable to find a role of AICD in transcriptional regulation shedding considerable doubt on the role of AICD in gene regulation.

Indexing (details)

0317: Neurosciences
Identifier / keyword
Biological sciences; APP; Alzheimer's disease; Amyloid precursor protein; Amyloid-beta; Aph-1; Gamma-secretase; Presenilin
Aph-1 functions as a substrate docking site within the Alzheimer's disease relevant gamma -secretase complex
Chen, Allen Ching-Way
Number of pages
Publication year
Degree date
School code
DAI-B 70/07, Dissertation Abstracts International
Place of publication
Ann Arbor
Country of publication
United States
Selkoe, Dennis J.
Harvard University
University location
United States -- Massachusetts
Source type
Dissertations & Theses
Document type
Dissertation/thesis number
ProQuest document ID
Database copyright ProQuest LLC; ProQuest does not claim copyright in the individual underlying works.
Document URL
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