The Nonconserved Hydrophilic Loop Domain of Presenilin (PS) Is Not Required for PS Endoproteolysis or Enhanced Aβ42 Production Mediated by Familial Early Onset Alzheimer's Disease-linked PS Variants

Abstract

Presenilin 1 (PS1) and presenilin 2 (PS2) are polytopic membrane proteins that are mutated in the majority of early onset familial Alzheimer's disease (FAD) cases. Two lines of evidence establish a critical role for PS in the production of ␤-amyloid peptides (A␤). FAD-linked PS mutations elevate the levels of highly amyloidogenic A␤ ending at residue 42 (A␤42), and cells with ablated PS1 alleles secrete low levels of A␤. Several recent reports have shown that the hydrophilic loop (HL) domain, located between transmembrane domains 6 and 7, contains sites for phosphorylation, caspase cleavage, and sequences that bind several PS-interacting proteins. In the present report, we examined the metabolism of PS polypeptides lacking the HL domain and the influence of these molecules on A␤ production. We report that the deletion of the HL domain does not have a deleterious effect on the regulated endoproteolysis of PS, saturable accumulation of PS fragments, or the selfassociation of PS fragments. A␤ production was not significantly altered in cells expressing HL-deleted PS polypeptides compared with cells expressing full-length PS. Importantly, deletion of the HL domain did not affect FAD mutation-mediated elevation in the production of A␤42. Furthermore, the deletion of the HL domain did not impair the role of PS1 or PS2 in facilitating Notch processing. Thus, our results argue against a biologically or pathologically relevant role for the HL domain phosphorylation and caspase cleavage and the association of PS HL domain-interacting proteins, in amyloid precursor protein metabolism and A␤ production or Notch cleavage.Autosomal dominant mutations in genes encoding presenilin 1 (PS1) 1 and presenilin 2 (PS2) predispose individuals to familial early onset Alzheimer's disease (FAD) (1-3). Alzheimer's disease is pathologically characterized by the cerebral deposition of 40 -42 amino acid ␤-amyloid (A␤) peptides, which are generated by the proteolytic processing of amyloid precursor protein (APP) (4, 5). PSs play an important role in the generation of A␤, as evidenced by the lack A␤ production in cells established from PS1 Ϫ/Ϫ mice (6, 7). Furthermore, FAD-linked mutations in PS increase the production of highly fibrillogenic A␤42 (8 -11). PS are polytopic membrane proteins that undergo regulated endoproteolysis to generate saturable of stable NH 2 -(NTF) and COOH-terminal fragments (CTF), which are the preponderant PS-related species that accumulate in vivo (12,13). Although the precise steps involved in the maturation of synthetic PS polypeptides are not clearly defined, properly folded PS polypeptides are stabilized, undergo endoproteolysis, and form high molecular weight complexes (13-18). The vast majority of overexpressed full-length PS and transgene-derived polypeptides that correspond to the NTF fail to form stable complexes and are rapidly degraded (15,18,19). Whereas endoproteolytic cleavage is not required for biological activity of PS polypeptides (12,20,21), several lines of evidence indicate that the NTF and CTF ...

The aspartate-257 of presenilin 1 is indispensable for mouse development and production of β-amyloid peptides through β-catenin-independent mechanisms

“…All transgenic lines were viable and overtly normal. Consistent with earlier reports (14,26), hPS1⌬cat protein was endoproteolytically cleaved to generate an NTF indistinguishable from that of wild-type hPS1 and a truncated CTF ( Fig. 1 B and D).…”

Functional Analysis of the Transmembrane Domains of Presenilin 1

“…A number of studies have revealed the seminal roles of the TMDs of the ␥-secretase components in the assembly of the active complex and acquisition of its proteolytic activity (12,15,17,18,24,25,32,33). Previously we and others found the requirement of TMD4 in the binding with Pen-2 (12, 33).…”

“…Anti-PS1 NT , anti-PS-C3, and anti-SPP ct antibodies were kindly provided by Drs. G. Thinakaran (University of Chicago, Chicago, IL), A. Takashima (RIKEN, Saitama, Japan), and T. E. Golde (Mayo Clinic, Jacksonville, FL), respectively (17)(18)(19). The monoclonal antibody anti-␣-tubulin AA4.3 developed by Dr. C. Walsh was obtained from the Developmental Studies Hybridoma Bank developed under the auspices of the NICHD, National Institutes of Health, and maintained by The University of Iowa, Department of Biology, Iowa City, IA.…”

“…The formation of stabilized HMW complex containing PS is tightly related to its enzymatic function (15,18). Upon BN-PAGE analysis, all components of the ␥-secretase complex were detected at the position of the HMW complex with an apparent molecular mass of 440 kDa (Fig.…”

Loss of presenilin 1 is associated with enhanced β-catenin signaling and skin tumorigenesis

“…4B, the luciferase activity was significantly higher in PS1-null keratinocytes as compared with control cells when both were transfected with the TOP-flash vector (filled bars in rescue vs. WT). Importantly, accelerated ␤-catenin͞LEF-mediated transcriptional activation in PS1 null cells could be suppressed by reintroducing WT PS1 but not by PS1⌬cat, a PS1 mutant deleting the essential ␤-catenin binding site but retaining full Notch activity (P Ͻ 0.001 between TOP ϩ PS1 vs. TOP ϩ PS1⌬cat; P Ͼ 0.05 between TOP vs. TOP ϩ PS1⌬cat, Student's t test) (30,33). These results demonstrate that the down-regulation of PS1 in ␤-catenin signaling is mediated through direct interaction of the two molecules and that abnormal ␤-catenin signaling in the absence of PS1 cannot be suppressed by restoring the PS1 Notch activity.…”