“Notch signalling impacts on amyloid β metabolism by modulating IDE and BACE1”.

LAURA MORELLI, PHD, EZEQUIEL I. SURACE, MARIA C. LEAL, MARIA P. HOLGADO, CARINA C. FERRARI, RODOLFO TARELLI, FERNANDO PITOSSI, THOMAS WISNIESWSKY, MIGUEL RIUDAVETS, EDUARDO M. CASTAÑO

Honolulu

Conferencia; 10 Alzherimer's Association International Conference on Alzheimer's disease 2010; 2010

Alzheimer Associaction

Abstract:Background:Cerebral amyloid β (Aβ) accumulation is pathogenically associated with sporadic Alzheimer`s disease (SAD). BACE-1 is the ratelimiting enzyme in Aβ generation while insulin-degrading enzyme (IDE) has a major role in the extracellular clearance of naturally secreted Aβ. Vulnerable neurons in AD brains show increased BACE-1 protein levels and enzymatic activity while the opposite occurs with IDE. The mechanisms underlying these observations remain mostly unknown. Other common feature in SAD brains is the overexpression of Notch-1. The role of Notch in neuronal differentiation during development and adult neurogenesis is well established. We hypothesize that Notch pathway activation is implicated in the transcriptional deregulation of genes involved in Aβ metabolism.Methods:With RSAT software we detected HES/Hey-1 (Notch target genes) putative binding sites on IDE promoter. By immunohistochemistry and QRT-PCR we determined Notch activation in brain samples. 125I-insulin and a fluorogenic substrate (R&Dcat#ES004) were used to assay IDE and BACE-1 activities, respectively. N2aSW (neuroblastoma cells stably expressing human Aβ40) were transfected with HA-Notch Intracellular Domain (NICD) or HA-Hey-1 cDNAs and assessment of IDE and BACE-1 transcript levels (by QRT-PCR), extracellular Aβ (by ELISA) and promoter activity (by LUC reporter assay) was performed. Cells were exposed to Jagged (JAG, a Notch ligand) and the parameters evaluated above were determined in the presence of a Notch-specific siRNA. Intracranial injection of JAG in Tg2576 mice, a well-established plaque pathology animal model of AD, was used for Notch signaling activation in vivo.Results:We identified over-expression of NICD and Notch target genes and alterations of BACE-1 and IDE activities in the hippocampus of SAD brains. We demonstrated that over-expression of NICD in N2aSW increased Hey-1 and BACE-1 and reduced IDE mRNA levels, promoting extracellular Aβ accumulation. We showed an opposite effect of NICD on IDE and on BACE promoters and on their proteolytic activities which were reverted by silencing Notch. We reproduced the molecular and pathological effect of Notch activation in Tg2576 mice.Conclusions:Our results support that a Notch-dependent IDE and BACE transcriptional modulation impacts on Aβ metabolism providing a novel functional link between Notch signaling and the amyloidogenic pathway in SAD.