NicoleBoyer,†,1ToshitakaKawarai,2Nade `geGirardot,3PeterSt.George-Hyslop,2andFre ´de ´ricChecler1
289,Pitie ´-Salpe ˆtrie `reHospital,Paris,France
death. We demonstrate that PS deficiency, catalytically inactive PS mutants, ?-secretase inhibitors, and ?APP or amyloid precursor
Several studies have reported on an increased p53-like immuno-
reactivity in sporadic Alzheimer’s disease (AD), especially in a
subpopulation of cortical neurons undergoing degeneration (de
la Monte et al., 1997; Kitamura et al., 1997; Garcia-Ospina et al.,
2003; Ohyagi et al., 2005), but the cellular mechanisms by which
p53 expression could be altered in AD remained puzzling. Prese-
nilin 1 (PS1) and PS2, together with nicastrin, anterior pharynx
defective-1 (Aph-1), and presenilin enhancer-2 (Pen-2), form
high-molecular-weight complexes that are necessary for the in-
tramembranous proteolysis of several type 1 transmembrane
proteins (termed ?-secretase cleavage) (for review, see De
Strooper, 2003). Many of these ?-secretase substrates (e.g.,
Notch, p75) are involved in biologically important signal trans-
duction pathways. Mutations in PS1 or PS2 in humans cause AD
disease characterized by progressive dementia, cerebral amyloid
brain. These mutations alter the processing of one of the
?-secretase substrates [?-amyloid precursor protein (?APP)],
generating increased amounts of a neurotoxic proteolytic frag-
ment (termed A?) (Checler, 1995). Furthermore, primary cul-
tured hippocampal neurons prepared from mice harboring mu-
tant PS1 exhibit increased vulnerability to apoptosis and
activated caspase (Guo et al., 1999). Several studies have shown
zin et al., 1996; Alves da Costa et al., 2002), likely via a p53-
dependent mechanism (Alves da Costa et al., 2002). However, it
consequence of the accumulation of neurotoxic A? or whether
cell death also occurs because other signaling pathways are per-
turbed as well. We therefore investigated a possible direct link
between PS-associated ?-secretase activity and the control of cell
Biochemical analysis of p53 in human and murine brain tissues. Biochem-
ical analysis of p53 immunoreactivity was performed on frontal cortices
from normal and AD-affected human brains. Samples from Rouen
(France) correspond to a control brain (female, 74 years old) and one
This work was supported by the National Center of Scientific Research, by the United for Alzheimer’s Disease
Research Foundation, by the Foundation for Medical Research, and by European Union contract LSHM-CT-2003-
dation for Medical Research. We are grateful to Drs. B. de Strooper (Leuven, Belgium), P. Saftig (Gottingen, Ger-
also thank Drs. C. Dumanchin and D. Campion (Rouen, France) for providing human brain samples. This work is
Correspondenceshouldbeaddressedtoeitherofthefollowing:Dr.Fre ´de ´ricChecler,InstituteofMolecularand
of Autonomous Syndicates, 660 Route des Lucioles, Sophia Antipolis, 06560 Valbonne, France, E-mail:
TheJournalofNeuroscience,June7,2006 • 26(23):6377–6385 • 6377
familial AD (FAD) brain (female, 51 years old, Leu392Val-PS1 muta-
tion).SamplesfromlaPitie ´-Salpe ˆtrie `re(Paris,France)correspondtotwo
control brains (males, 74 and 55 years old) without history of dementia
or other neurological disease (Braak stage 0), and two FAD brains [fe-
male, 37 years old, Leu235Pro-PS1 mutation; male, 44 years old,
www.jneurosci.org as supplemental material )]. The mean postmortem
delay was 27.8 ? 7.1 h. PS1/PS2 double knock-out murine brains have
been described recently (Saura et al., 2004).
p53 expression, activity, and promoter transactivation. Cellular p53 im-
munoreactivity was analyzed by Western blot, using a 1:10,000 dilution
of an anti-p53 mouse monoclonal antibody (Santa Cruz Biotechnology,
Santa Cruz, CA) in nuclear extracts prepared as previously described for
cytochrome c translocation experiments (Alves da Costa et al., 2002)
while murine brain p53 was analyzed in total homogenates, using the
sient transfection of the PG13-luciferase (PG13) cDNA kindly provided
by Dr. B. Vogelstein (Baltimore, MD). This construction is based on the
genomic DNA consensus sequence recognized by p53 on its target genes
promoter (mPP) (Ginsberg et al., 1990) or human p53 promoter (hPP)
sequences in-frame with luciferase (provided by Dr. M. Oren, Rehovot,
Israel). All activities were measured after cotransfection of 0.5–1 ?g of
the above cDNAs and 0.25–0.5 ?g of ?-galactosidase (?-gal) cDNA to
normalize transfection efficiencies. Importantly, because p53 could be
at a 60–70% cell density.
Real-time quantitative PCR. Total RNA from cells was extracted at the
the manufacturer (Qiagen, Hilden, Germany). After treatment with
DNase I, 2 ?g of total RNA was reverse transcribed, using oligo-dT
priming and avian myeloblastosis virus reverse transcriptase (Promega,
Madison, WI). Real-time PCR was performed in an ABI PRISM 5700
Sequence Detector System (Applied Biosystems, Foster City, CA), using
the SYBR Green detection protocol as outlined by the manufacturer.
Gene-specific primers were designed by using the Primer Express soft-
ware (Applied Biosystems). Relative expression level of target genes was
normalized for RNA concentrations with two different house keeping
genes [human glyceraldehyde phosphate dehydrogenase (GAPDH),
mouse ?-actin] according to the cell specificity. mRNA values are ex-
representative of three to five independent experiments.
In the experiments dedicated to the analysis of p53 mRNA stability,
human embryonic kidney 293 (HEK 293) cells were transiently trans-
fected with empty or [amyloid intracellular C-terminal domain C59-
encoding (AICDC59-encoding)] pcDNA3 vector. At 33 h after transfec-
tion the cells were treated with 10 ?g/ml of actinomycin D for various
time periods; then total RNA was extracted by using the RNeasy kit,
reverse transcribed, and submitted to real-time PCR as described above.
AICD-induced caspase 3 activation. Wild-type fibroblasts were trans-
fected with empty or AICDC59 coding vectors, alone or in combination
with Tat interactive protein 60 (Tip60) and family B, member 1 (Fe65)
cDNA, using Lipofectamine 2000 (Invitrogen, Cergy-Pontoise, France)
according to the manufacturer’s recommendation. At 36 h after trans-
inhibitor of p53 transcriptional activity (Komarov et al., 1999) or an
inactive analog (kindly provided by M. P. Mattson, Johns Hopkins Hos-
pital and University School of Medicine, Baltimore, MD). Cells subse-
staurosporine (Sigma, St. Quentin-Fallavier, France). Cell were lysed in
previously described (Alves da Costa et al., 2003). To analyze AICDC59-
like immunoreactivity, we resolved 40 ?g of proteins of the same cell
lysate on 16% Tris-tricine gel, transferred them onto Hybond C mem-
the extreme C terminus of ?APP (kind gift from M. Goedert, Medical
Research Council, Cambridge, UK). Fe65- and Tip60-like immunoreac-
tivities were analyzed as described previously (Pardossi-Piquard et al.,
Human brain preparation and immunohistochemical analysis of p53.
Immunohistochemical analysis of p53 was performed on the temporal
month. Samples from isocortex (Brodmann area 22) were embedded in
pH 6, for two cycles of 10 min. Endogenous peroxidases then were
quenched by a solution of H2O2(3%) for 10 min. Slices were left for 60
min in a solution of Tris-buffered saline (TBS) containing Tween 20
(0.5%) and bovine serum albumin (2%) to limit nonspecific fixation of
p53 antibody (clone sc-98, Santa Cruz) at room temperature. The bio-
tinylated secondary antibody and the solution of streptavidin–peroxi-
dase (ChemMate kit; Dako, High Wycombe, UK) were applied for 25
min. Immunological complexes were revealed by diaminobenzidine
(DAB). Between each step the sections were rinsed in TBS containing
0.5% of Tween 20. Slides were counterstained with Harris hematoxylin,
dehydrated, and mounted in DPX medium. Three slides of each sample
were immunolabeled. A total of 200 neuronal profiles per slide were
identified with the use of a 40? objective. The proportion of neuronal
and controls, using ANOVA (model I, fixed, and Fisher’s protected least
significant difference for post hoc comparisons between groups).
Presenilin deficiency lowers p53 transcription in vitro and
p53 is a tumor suppressor protein involved in the direct tran-
caspase 1 (Gupta et al., 2001) and the oncogene bax (Miyashita
and a consensus binding site sequence had been delineated (El-
Deiry et al., 1992). We have used this sequence in-frame with a
itor p53 functional activity (El-Deiry et al., 1992). In PS1- and
PS2-deficient (PS-deficient) fibroblasts and in PS-deficient blas-
tocysts there was a reduction in the amount of p53 protein in
decrease in p53 activity (Fig. 1D). In agreement with these data,
(Saura et al., 2004) by conditional knock-out (46.7 ? 2.9% of
control brains; n ? 5; p ? 0.05) (Fig. 1E).
To assess whether the observed reductions in p53 protein ex-
pression resulted from decreased p53 mRNA transcription, we
harbors the mPP region in-frame with a luciferase reporter gene.
These experiments confirmed that the reductions of p53 immu-
noreactivity and p53 activity were accompanied by a decrease in
the transactivation of the p53 promoter in both PS-deficient fi-
endogenous p53 mRNA levels were significantly lower in PS-
deficient fibroblasts than in wild-type fibroblasts (Fig. 1F).
(Araki et al., 2001; Alves da Costa et al., 2002) via a p53-
dependent pathway (Alves da Costa et al., 2002). We therefore
examined the weight of the contribution of PS2 in the alteration
of p53 observed after depletion of both PSs. Overall, PS2 deple-
6378 • J.Neurosci.,June7,2006 • 26(23):6377–6385AlvesdaCostaetal.•Presenilin-Dependent?-SecretaseModulatesp53
tion fully mimicked the phenotype observed after both PS1 and
Fig. 1A,B, available at www.jneurosci.org as supplemental mate-
rial). Conversely, PS2 depletion led to reduced p53 activity and
expression and lowered the transactivation of p53 promoter
(supplemental Fig. 1C,D, available at www.jneurosci.org as sup-
plemental material). The same effects on p53 activity, promoter
able at www.jneurosci.org as supplemental material) were ob-
served in cells expressing PS2 harboring the N141I FAD
Unlike PS2, PS1 appeared either to be inert in nonstimulated
conditions or to decrease neuronal susceptibility to various apo-
ptotic stimuli (Bursztajn et al., 1998). Interestingly, PS1 overex-
pression decreased p53 activity, expression, and promoter
transactivation (supplemental Fig. 3A,B, available at www.
jneurosci.org as supplemental material), whereas PS1 deletion
led to an opposite phenotype (supplemental Fig. 3C,D, available
at www.jneurosci.org as supplemental material). The distinct
control of p53 by these proteins led us to examine whether there
could be a functional cross talk between the two parent preseni-
lins for this paradigm. In support of such an hypothesis, we pre-
viously demonstrated that PS2 overexpression drastically re-
duced the levels of both overexpressed and endogenous PS1
(Alves da Costa et al., 2002). Here we show that PS1 depletion
drastically increased PS2-like immunoreactivity (supplemental
rial); conversely, PS1 overexpression significantly decreased PS2
expression (supplemental Fig. 4B,C, available at www.jneuro-
sci.org as supplemental material).
In agreement with analyses in PS-deficient cells, the administration
5-phenylpentyl}carbamic acid tert-butyl ester (L685458), two
(Fig. 2B), and transactivation of the p53 promoter (Fig. 2C).
Significantly, these inhibitors were totally ineffective in PS-
deficiency and ?-secretase inhibition suggested that the endoge-
nous product controlling p53 transcription in wild-type fibro-
blasts was likely to be generated via a PS-dependent ?-secretase
activity. To support this hypothesis further, we established stable
transfectants expressing mutated PS1 in which the residues of
aspartyls 257 or 385 had been replaced by aliphatic amino acids
(Fig. 2D). It had been shown previously that these mutated pro-
teins, when overexpressed, substituted for endogenous PSs and
inhibited the ?-secretase-mediated cleavages of ?APP and
Notch. In agreement with the above data, both mutated PS1s
drastically diminished the staurosporine-induced caspase 3 acti-
vation in HEK 293 cells (Fig. 2E). This was accompanied by a
nuclear extracts of wild-type (PS?/?, BD6) or PS-deficient (PS?/?, BD8) fibroblasts (A) or
sient cotransfection of ?-galactosidase and PG13-luciferase cDNA in the indicated fibroblast
determinations. E, p53 immunoreactivity measured in whole proteic extracts prepared from
measured after transfection of the mPP-luciferase construct in the indicated fibroblast and
blastocyst cell lines. Bars correspond to the ratios of luciferase/?-galactosidase activities ex-
the indicated ?-secretase inhibitor (DFK167, 50 ?M; L685, 2 ?M); then p53 expression in
sured as described in Materials and Methods. Bars correspond to the ratios of luciferase/?-
are the means ? SEM of three independent determinations. D–F, Stably transfected TSM1
in E and F are the means ? SEM of nine and three independent experiments, respectively,
Effect of ?-secretase inhibitors on p53. PS?/?and PS?/?fibroblasts were
AlvesdaCostaetal.•Presenilin-Dependent?-SecretaseModulatesp53J.Neurosci.,June7,2006 • 26(23):6377–6385 • 6379
drastic reduction of p53 mRNA levels
measured by real-time PCR (Fig. 2F).
A?40and a 59-amino-acid-long fragment
been demonstrated recently that ?APP
also undergoes an additional cleavage (?-
cleavage) slightly downstream of the
“?-site,” liberating a shorter 50-amino-
acid-long peptide (AICDC50) (Gu et al.,
2001; Sastre et al., 2001). We therefore ex-
amined whether AICDC50 and AICDC59
overexpression could modulate p53 in
wild-type and PS-deficient murine blasto-
cysts as well as in HEK 293 cells. AICD
and transactivation of murine and human
p53 promoters in wild-type blastocysts
(Fig. 3A,B, BD6), PS-deficient blastocysts
(Fig. 3A,B, BD8), and HEK 293 cells (Fig.
3C,D). Accordingly, real-time PCR analy-
sis showed that both AICDC50 and
AICDC59 significantly induced p53
rule out a possible influence of AICD on
AICDC59-transfected HEK 293 cells with
actinomycin D to block neo transcription;
al-time PCR. Figure 3G confirms that
AICDC59-expressing cells display higher
p53 mRNA levels decrease with similar
slopes (29.85 vs 29.69) in both cell lines,
effect on p53 mRNA by affecting its cellu-
lar stability. Indeed, AICDC59 clearly
transactivates p53 promoter and increases
p53 mRNA levels by physically interacting
with the p53 promoter, as demonstrated
by gel shift assays (Fig. 3F).
p53 is a tumor suppressor gene that exerts
its pro-apoptotic signaling via complex
cellular pathways (Fisher, 2001). It has
been established previously that p53-
sociated with the activation of caspase 3
(Cregan et al., 1999). We therefore hy-
pothesized that caspase 3 activation could
be used as a read-out of the function of
AICD in the cellular control of p53. This
hypothesis was confirmed by the observa-
tion that AICDC59 cDNA transfection
(Fig. 4A) led to a statistically significant
increase in staurosporine-stimulated
were not treated (white bars). C, Pifithrin-?-sensitive p53-dependent caspase 3 activation was determined by the difference
AICDC59 increases caspase 3 activity in a p53-dependent manner. Wild-type fibroblasts were transfected with
transfected with PG13-luciferase (A, C), mPP-luciferase (B), or hPP-luciferase (D) vectors together with an empty vector (Ct),
transfected with empty vector (Ct), AICDC50 (C50), or AICDC59 (C59) cDNA; then p53 mRNA levels were analyzed 48 h after
transfection by real-time PCR as described in Materials and Methods. Bars correspond to p53 mRNA density expressed as a
enriched nuclear extracts; lane 2,32P-labeled p53-2 probe in competition with cold p53-2 probe; lane 3, supershift of the
32P-labeled p53-2 probe by anti-myc antibodies; lane 4, supershift of the32P-labeled p53-2 probe with control nonspecific
6380 • J.Neurosci.,June7,2006 • 26(23):6377–6385 AlvesdaCostaetal.•Presenilin-Dependent?-SecretaseModulatesp53
caspase 3 activity in fibroblasts (Fig. 4B). This increase was abol-
ished by the p53 inhibitor, pifithrin-?, but not by one of its inac-
tive analogs (Fig. 4B). To enhance AICDC59-associated pheno-
stabilize AICDs (Kimberly et al., 2001; Kinoshita et al., 2002b).
Furthermore, Tip60, together with Fe65, was reported to favor
AICD translocation into the nucleus where the protein could act
as a transcription factor (Cao and Su ¨dhof, 2001). Clearly,
AICDC59, Fe65, and Tip60 cDNA cotransfection enhanced
AICDC59 immunoreactivity in fibroblasts (Fig. 4A). Interest-
ingly, this increase was associated with a drastic increase in total
(Fig. 4B) and pifithrin-?-sensitive (Fig. 4C) caspase 3 activity.
These data additionally emphasize our observation of an AICD-
associated control of p53 and indicate that AICD-induced p53
We next attempted to examine whether increased caspase 3 acti-
could be abolished by p53 deficiency. We took advantage of re-
cent p19Arf?/?(Kamijo et al., 1997) and p19Arf?/?p53?/?(We-
deficiency. First we showed that p19Arf?/?-deficient cells still
respond to staurosporine by activating caspase 3 and that this
phenotype is reduced by additional p53 depletion (data not
shown). Interestingly, we confirmed that PS2 and N141I-PS2-
both proteins increased caspase 3 activity in p19Arf?/?fibro-
blasts, whereas they were fully inactive in p19Arf?/?p53?/?fi-
broblasts (supplemental Fig. 2D,E, available at www.jneurosci.
org as supplemental material). AICDC59 transfection, together
with Fe65 and Tip60, drastically enhanced pifithrin-?-sensitive
caspase 3 activity, although this phenotype was not observed in
Arf?/?p53?/?fibroblasts (supplemental Fig. 5A,B, available at
www.jneurosci.org as supplemental material). Of most interest,
we showed that AICDC59 transfection increased the number of
terminal deoxynucleotidyl transferase-mediated biotinylated
UTP nick end labeling-positive (TUNEL-positive) cells in
p19Arf?/?cells, which was increased additionally by Fe65 and
supplemental material). However, the lack of p53 fully abolished
the AICDC59 and AICDC59/Tip60/Fe65-induced cell death
(supplemental Fig. 5D,E, available at www.jneurosci.org as sup-
plemental material). These data agree well with previous studies
indicating that the ?-secretase-generated C-terminal fragments
phenotype was linked to Tip60 in human neuroglioma H4 cells
(Passer et al., 2000; Kinoshita et al., 2002a). They additionally
support the fact that this process occurs via the control of p53
deficiencies affect p53 in vitro and in vivo
PS-dependent ?-secretase activity is required for the intramem-
branous cleavage of several proteins (Sisodia and St. George-
derived AICDs, we examined the status of p53 in ?APP?/?
transactivation of the murine p53 promoter construct, p53
tein expression (45 ? 4.3% of reduction; n ? 6; p ? 0.001) (Fig.
5B). Interestingly, p53 protein expression also was affected in a
gene dose-dependent manner in vivo. Thus p53 immunoreactiv-
ity was lower in ?APP?/?mice brain than in control ?APP?/?
mice, a phenotype additionally accentuated in ?APP?/?mice
brain (Fig. 5C,D). These data indicate that at least some of the
endogenous fragments generated by PS-dependent ?-secretase
that modulate p53 were AICDs. Clearly, however, the depletion
of ?APP did not achieve full downregulation of p53 expression
and activity. It should be noted here that fibroblasts do not ex-
press amyloid precursor protein-like protein 1 (APLP1; data not
cleavage (Eggert et al., 2004) leading to biologically active intra-
cellular C-terminal domain (Scheinfeld et al., 2002; Walsh et al.,
trol of p53. Indeed, in fibroblasts the APLP2 knock-out led to
reduced p53 expression (Fig. 6A,B), activity (Fig. 6C), promoter
transactivation (Fig. 6D), and mRNA levels (Fig. 6E). None of
these paradigms appeared to be potentiated additionally by dou-
ble ?APP/APLP2 deficiency (Fig. 6A–E).
ial AD cases, which affect ?-secretase activity, also should modu-
p53 activity in cells. Transient cotransfections of PG13 reporter
sense mutations or deletion (?E9PS1) (Perez-Tur et al., 1995),
blasts. Bars for PG13- and mPP-luciferase activities correspond to the ratios of luciferase/?-
Influence of ?APP depletion on p53 in vitro and in vivo. A, p53 transcriptional
AlvesdaCostaetal.•Presenilin-Dependent?-SecretaseModulatesp53J.Neurosci.,June7,2006 • 26(23):6377–6385 • 6381
led to significantly increased p53 activity (Fig. 7A). These data
were corroborated by immunohistochemical analysis of p53 im-
munoreactivity in the same FAD cases. Figure 7B shows the typ-
ical labeling of endogenous p53 in control (Fig. 7B, top panels)
and FAD-affected (Fig. 7B, bottom panels) temporal cortices. In
nondemented cases the p53 immunoreactivity was diffuse in
neuropil and cell bodies of a few neurons, whereas the p53 stain-
ing of neuronal cell bodies was consistently much more intense
cases (Fig. 7B,C). Cell nuclei, central cores of senile plaques,
immature lesions (diffuse plaques), and neurofibrillary tangles
were not labeled. Close quantification of p53-positive neurons
indicated a higher number of positive neurons in FAD brains
than in unaffected brains (Fig. 7C). In agreement with these in
from three different FAD brains harboring PS1 mutations (Fig.
7D). These data indicated that neuronal p53 expression was
higher in FAD than in nondemented brains at both immunohis-
tochemical and biochemical levels.
However, it should be noted that p53 immunoreactivity and
enhanced ?-secretase activity but rather to post-transcriptional
modifications, we examined the status of insulin-degrading en-
al., 2002). Indeed, we found that in sporadic AD brains, IDE was
reduced significantly (Fig. 7E), thereby suggesting a possible
higher catabolic stability of AICD in spo-
radic brains and thereby contributing to
enhanced p53 expression. It is noteworthy
that IDE levels in FAD brains were not af-
fected (Fig. 7E).
The PS-dependent ?-secretase is borne by
a high-molecular-weight complex com-
PS1 or PS2, nicastrin, Aph-1, and Pen-2
(Herreman et al., 2000; Yu et al., 2000;
Zhang et al., 2000; Francis et al., 2002;
Goutte et al., 2002). Because there are two
different PSs (PS1 and PS2), three murine
Aph-1 homologs (Aph-1a, Aph-1b, and
Aph-1c) (Steiner et al., 2002; Luo et al.,
2003; Ma et al., 2005), and even two splice
isoforms of Aph-1a (Aph-1aL and Aph-
1aS) (Steiner et al., 2002; Gu et al., 2003;
Luo et al., 2003), several types of com-
plexes harboring distinct combinations of
deed, this was demonstrated by Shirotani
and colleagues, who characterized various
complexes with specific composition (He-
ber et al., 2000; Shirotani et al., 2004).
Some lines of evidence indicate that these
physical differences could underlie dis-
tinct biological activities (Hong et al.,
1999; Chen et al., 2003; Gu et al., 2004;
Kang et al., 2005).
It previously had been documented
differently. Thus PS1 decreased neuronal
(C), transactivation of murine p53 promoter (D), and mRNA levels (E) were measured as de-
double ?APP/APLP2-deficient (DKO) fibroblasts. Note that fibroblasts do not express APLP1
Influence of APLP2 depletion on p53. p53 immunoreactivity (A, B), p53 activity
6382 • J.Neurosci.,June7,2006 • 26(23):6377–6385AlvesdaCostaetal.•Presenilin-Dependent?-SecretaseModulatesp53
death appeared to be enhanced in antisense PS1-expressing cells,
a phenotype that could be rescued by overexpression of the anti-
apoptotic oncogene Bcl-2 (Hong et al., 1999). Unlike PS1, PS2
displays a clear pro-apoptotic phenotype (Wolozin et al., 1996;
Costa et al., 2002). Our data here clearly confirm the opposite
functions of the two parent proteins and indicate that both phe-
notypes were associated with a modulation of p53 expression,
p53 activity, and transcriptional activation of the p53 promoter
(supplemental Figs. 1, 3, available at www.jneurosci.org as sup-
plemental material). Our study also establishes a possible cross
talk between the two presenilins. Thus we previously demon-
strated that overexpression of either wild-type or mutated PS2
(Alves da Costa et al., 2002). Here we show that PS1 depletion or
overexpression drastically increases or reduces PS2 expression,
as supplemental material), in agreement with a recent study
showing that mutated PS1 drastically reduced PS2 fragments
(Kang et al., 2005). Because it has been shown that p53 acted as a
repressor of the transcription of the human PS1 gene (Pastorcic
and Das, 2000) in agreement with a previous study demonstrat-
ing the inhibition of PS1 expression promoted by p53 (Roperch
et al., 1998), we can postulate reasonably that PS2-mediated in-
crease in p53 concomitantly leads to reduced PS1 expression.
This intimate cross talk between PS1 and PS2 ultimately could
control the levels of the ?-secretase-mediated AICD formation.
control of p53 by AICD, the PS-dependent ?-secretase cleavage
product of ?APP, and clearly suggests that FAD mutations,
which increase ?-secretase cleavage (Marambaud et al., 1998)
and likely increase susceptibility to subsequent apoptotic cell
death in FAD cases of AD. This agrees well with a study showing
that presenilin 1 mutations increased cell vulnerability to DNA
damage, a cellular traumatism generally associated with p53-
enhanced production (Chan et al., 2002).
The correlation among p53 overexpression, neuronal DNA
damage, and apoptosis (Bar et al., 2004) raises the question as to
whether AICD-regulated changes in p53 expression also could
generally is thought that neuropathological lesions arise from
postproduction changes in ?-secretase products, e.g., the fibrilli-
zation of A? or reduced catabolism of A? and AICD. In this
context it is noteworthy that the activity of IDE, the enzyme that
apparently catabolizes both A? and AICD (Edbauer et al., 2002),
is reduced in brain tissue from late-onset sporadic cases of AD
increase cellular AICD levels and contribute to enhanced AICD-
mediated p53 expression in sporadic AD.
Together, our data demonstrate both a new function for PS-
dependent ?-secretase-associated AICD and an unexpected
implication for AD therapeutics. The strategy aimed at specifi-
cally inhibiting ?-secretase faces major theoretical problems.
of several proteins, the functions of which are linked closely to
sodia and St. George-Hyslop, 2002). Furthermore, we recently
demonstrated that blockade of PS-dependent ?-secretase could
lead to decreased activity of neprilysin, a major A?-degrading
enzyme, thereby enhancing the concentration of A? peptide
(Pardossi-Piquard et al., 2005, 2006). The present report addi-
tionally suggests that chronic treatment with ?-secretase inhibi-
tors could affect p53 drastically and, potentially therefore, ulti-
mately lead to tumorigenicity. In this context it is noteworthy
that an APP-dependent complex including Fe65 and Tip60 also
regulates the transcription of another tumor suppressor, namely
KAI1/CD82 (Baek et al., 2002). This concern is supported addi-
tionally by the observation that PS deficiency leads to malignant
keratocarcinoma in mice (Tournoy et al., 2004) and that the se-
verity of the skin lesions, from benign keratosis to carcinomas,
skin tumors arise specifically via loss of PS-dependent regulation
of p53 per se. Nevertheless, our results emphasize the need for
potential of ?-secretase inhibitors as long-term treatments or
prophylactic therapies in AD.
Alves da Costa C, Paitel E, Mattson MP, Amson R, Telerman A, Ancolio K,
Checler F (2002) Wild-type and mutated presenilin-2 trigger p53-
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Alves da Costa C, Mattson M, Ancolio K, Checler F (2003) The C-terminal
fragment of presenilin 2 triggers p53-mediated staurosporine-induced
apoptosis, a function independent of the presenilinase-derived
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