Abstract
It has been proposed that gamma-secretase-mediated release of the amyloid precursor protein (APP) intracellular domain (AICD) results in nuclear translocation and signaling through a complex with the adaptor protein Fe65 and the histone acetyltransferase Tip60. Here, we show that APP and Fe65 activate transcription through a Gal4-Tip60 reporter in presenilin-1/2-deficient cells lacking generation of AICD. APP and Fe65 also activated transcription in the presence of gamma-secretase inhibitors that prevent amyloid beta-peptide production in human embryonic kidney 293 and SH-SY5Y cells. In contrast to the transcriptionally active Notch intracellular domain, expression of AICD did not activate transcription. An alternative mechanism for APP signal transduction is suggested by the identification of essential cyclin-dependent kinase (CDK) phosphorylation sites in Tip60. Mutation of these Tip60 phosphorylation sites or treatment with the CDK inhibitor roscovitine blocked the ability of APP to signal through Tip60. Moreover, APP stabilized Tip60 through CDK-dependent phosphorylation. Subcellular fractionation and confocal immunofluorescence showed that APP recruited Tip60 to membrane compartments. Thus, APP may signal to the nucleus by a gamma-secretase-independent mechanism that involves membrane sequestration and phosphorylation of Tip60.
Highlights
Main shedding followed by intramembrane cleavage is reminiscent of the processing of the Notch receptor following binding to its ligands
amyloid precursor protein (APP) Signaling in Presenilin-deficient Cells—Activation of the Notch signaling pathway requires ␥-secretase cleavage by the presenilins to release the Notch intracellular domain (NICD), which translocates to the nucleus and activates target genes
To determine whether APP could signal to the nucleus by a similar mechanism, we utilized embryonic stem (ES) cells deficient in both presenilin-1 and presenilin-2 (presenilin-1/2 knockout (PS-KO) cells)
Summary
Main shedding followed by intramembrane cleavage is reminiscent of the processing of the Notch receptor following binding to its ligands (reviewed in Ref. 3). It has been suggested that APP may signal by a mechanism similar to that of the Notch receptor, in which the intracellular tail released by presenilin/␥-secretase translocates to the nucleus and activates transcription. This idea was supported by the observation that APP, together with the adaptor protein Fe65, could activate the transcriptional activity of the histone acetyltransferase Tip60 [18]. We provide evidence that, in contrast to Notch, APP is able to activate transcription through Tip independently of ␥-secretase cleavage by recruiting Tip to the membrane, leading to Tip activation through cyclin-dependent kinase-mediated phosphorylation This is followed by nuclear translocation of Tip and Fe65 and activation of transcription
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