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A novel cleavage of ␤-amyloid precursor protein (APP), referred to as ⑀-cleavage, occurs downstream of the ␥-cleavage and generates predominantly a C-terminal fragment (CTF␥) that begins at Val-50, according to amyloid ␤-protein (A␤) numbering. Whether this cleavage occurs independently of, or is coordinated with, ␥-cleavage is unknown. Using a cell-free system, we show here that, although A␤40 and CTF␥ 50 -99 were the predominant species produced by membranes prepared from cells overexpressing wild-type (wt) APP and wt presenilin (PS) 1 or 2, the production of CTF␥ 49 -99, which begins at Leu-49, was remarkably enhanced in membranes from cells overexpressing mutant (mt) APP or mtPS1/2 that increases the production of A␤42. Furthermore, a ␥-secretase inhibitor, which suppresses A␤40 production and paradoxically enhances A␤42 production at low concentrations, caused the proportion of CTF␥ 50 -99 to decrease and that of CTF␥ 49 -99 to increase significantly. These results strongly suggest a link between the production of A␤42 and CTF␥ 49 -99 and provide an important insight into the mechanisms of altered ␥-cleavage caused by mtAPP and mtPS1/2.Senile plaques, one of the neuropathological hallmarks of Alzheimer's disease (AD), 1 are composed primarily of amyloid ␤-protein (A␤) (1). Two major A␤ species consisting of 40 and 42 residues are generated mainly in neurons and constitutively secreted. A shorter one, A␤40, is predominant, and a longer one, A␤42, is a minor species (Ͻ10%) among secreted A␤ species. A␤ is produced from ␤-amyloid precursor protein (APP), through sequential cleavage by proteases referred to as ␤-and ␥-secretases. ␤-Secretase was identified as a type I membrane aspartic protease ␤-site APP-cleaving enzyme (BACE) (2), but the identity of ␥-secretase has remained unknown. ␥-Secretase cleaves APP in the middle of the transmembrane domain, releasing A␤ and its counterpart, C-terminal fragment ␥ of APP (CTF␥). Most recent studies have shown that ␥-secretase forms a large complex composed of presenilin (PS) 1 or 2, nicastrin, PEN-2, and APH-1, and the activity of ␥-secretase is now known to depend on these proteins (3-7).One of the A␤ species, A␤42, has a much higher aggregation potential (8, 9) and is believed to be initially deposited in senile plaques (10). It is reasonable to postulate that A␤42 accumulation in the brain is the very initial event in the development of AD including sporadic AD. Indeed, all mutations of PS1/2 and some mutations of APP that cause familial AD result in increased A␤42 production (11).Recently, we and other groups found that APP is cleaved by PS-dependent ␥-secretase, not only in the middle of the transmembrane domain (␥-cleavage) but also near the cytoplasmic membrane boundary (⑀-cleavage) (12-15). The major product of the latter process is a CTF␥ of APP that begins at Val-50. This cleavage site is a few residues inside the membrane from the cytoplasmic/membrane boundary and is similar to site 3 cleavage of Notch (16). Since production of CTF␥ is inhibited by a dominan...

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The amyloid precursor protein (APP) plays a critical role in Alzheimer’s disease (AD) pathogenesis. APP is proteolytically cleaved by β- and γ-secretases to generate the amyloid β-protein (Aβ), the core protein component of senile plaques in AD. It is also cleaved by α-secretase to release the large soluble APP (sAPP) luminal domain that has been shown to exhibit trophic properties. Increasing evidence points to the development of synaptic deficits and dendritic spine loss prior to deposition of amyloid in transgenic mouse models that overexpress APP and Aβ peptides. The consequence of loss of APP, however, is unsettled. In this study, we investigated whether APP itself plays a role in regulating synaptic structure and function using an APP knock-out (APP−/−) mouse model. We examined dendritic spines in primary cultures of hippocampal neurons and CA1 neurons of hippocampus from APP−/− mice. In the cultured neurons, there was a significant decrease (~35%) in spine density in neurons derived from APP−/− mice compared to littermate control neurons that were partially restored with sAPPα-conditioned medium. In APP−/− mice in vivo, spine numbers were also significantly reduced but by a smaller magnitude (~15%). Furthermore, apical dendritic length and dendritic arborization were markedly diminished in hippocampal neurons. These abnormalities in neuronal morphology were accompanied by reduction in long-term potentiation. Strikingly, all these changes in vivo were only seen in mice that were 12-15 months in age but not in younger animals. We propose that APP, specifically sAPP, is necessary for the maintenance of dendritic integrity in the hippocampus in an age-associated manner. Finally, these age-related changes may contribute to Alzheimer’s changes independent of Aβ-mediated synaptic toxicity.

Leucine-rich repeat kinase 2 (LRRK2), a large protein kinase containing multi-functional domains, has been identified as the causal molecule for autosomal-dominant Parkinson's disease (PD). In the present study, we demonstrated for the first time that (i) LRRK2 interacts with tau in a tubulin-dependent manner; (ii) LRRK2 directly phosphorylates tubulin-associated tau, but not free tau; (iii) LRRK2 phosphorylates tau at Thr181 as one of the target sites; and (iv) The PD-associated LRRK2 mutations, G2019S and I2020T, elevated the degree of tau-phosphorylation. These results provide direct proof that tau is a physiological substrate for LRRK2. Furthermore, we revealed that LRRK2-mediated phosphorylation of tau reduces its tubulin-binding ability. Our results suggest that LRRK2 plays an important role as a physiological regulator for phosphorylation-mediated dissociation of tau from microtubules, which is an integral aspect of microtubule dynamics essential for neurite outgrowth and axonal transport.