Project: Research project

Project Details


The 4 kD (39-43 residue) amyloid beta protein (AB), which is deposited
as amyloid in Alzheimer's disease (AD), is encoded as an internal peptide
that begins 99 residues from the carboxyl terminus of a set of 695-770
residue glycoproteins referred to as the amyloid Beta protein precursor
(BAPP). Recent data from our laboratory and others has established that
normal processing of the BAPP involves (i) a constitutive secretory
pathway in which the BAPP is cleaved within AB to produce a large,
secreted, NH2-terminal derivative and an 8.7 kD COOH-terminal fragment,
neither of which can produce amyloid because they do not contain the
entire AB, (ii) endosomal/lysosomal processing which produces a complex
set of COOH-terminal derivatives that includes potentially amyloidogenic
forms with the entire AB a or near their NH2 terminus, and (iii) the
production of 4 kD AB(essentially identical to the AB deposited as
amyloid in AD) that is released from cultured cells and readily detected
in CSF. Strong evidence that amyloid deposition plays a critical role
in the development of AD has come from the identification of familial AD
(FAD) kindreds in which the AD phenotype cosegregates with mutations in
the BAPP gene. Three of these mutations alter the valine mutation (NL)
alters the lysine-methionine located immediately amino to AB1
(lys670=met671 in BAPP) to isoleucine, phenylalanine, or glycine. A
fourth double mutation (NL) alters the lysine-methionine located
immediately amino to A1 (lys670-met671 in BAPP770) to asparagine-leucine.
The location of these mutations in close proximity to AB immediately
suggests that they may cause AD by altering BAPP processing in a way that
is amyloidogenic. To evaluate this possibility, we have recently compare
human neuroblastoma (M17) cells expressing normal BAPP695 or FAD-linked
mutant BAPP 695. Cells expressing the BAPP NL mutant showed a 5-fold
increase in the relative amount of an 11.4 kD AB-bearing carboxyl-
terminal BAPP derivative, and they released 6-fold more 5 kD AB into the
medium. These observations provide strong evidence (i) that this mutant
BPP causes AD because it undergoes altered processing that releases
increased amounts of AB, and (ii) that the pathway producing AB in
cultured cells is highly relevant to AD. In this application, we propose
to expand our analysis of AB production by undertaking experimentation
designed t o (1) test the hypothesis that the 4kD AB produced and
released by normal cellular processing consists of several peptides with
variable COOH-termini (AB39-43) similar to the AB peptides that have been
identified in AD amyloid, (2) test the hypothesis that the BAPP717
mutants favor production of the longer AB1-42 or 43 forms, which
selectivity deposit as plaque core amyloid, without increasing the
production of AB as expected from the observation by our group and others
that phorbol esters substantially increase the production of the large
secreted BAPP derivative. In addition, we propose (4) to compare BAPP
processing and the AB released by transfected mouse or human cells
expressing wild type BAPP or the FAD-linked mutant BAPPs in order to (i)
determine if the disappointing results obtained to date in transgenic
mouse model of AD are related to low rates of AB production in mouse
cells expressing human BAPPs, and (ii) identify constructs likely to
significantly increase CNS AB production when they are introduced into
transgenic mice. Finally, we plan (5) to carry out a systematic
comparison of the amount of soluble AB in various regions of AD and
control brains in an effort to determine if the amount of soluble AB is
correlated with AD and/or amyloid deposition.
StatusNot started


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