Expression and analysis of presenilin 1 in a human neuronal system: localization in cell bodies and dendrites.

Abstract

Mutations in the recently identified presenilin 1 gene on chromosome 14 cause early onset familial Alzheimer disease (FAD). Herein we describe the expression and analysis of the protein coded by presenilin 1 (PS1) in NT2N neurons, a human neuronal model system. PS1 was expressed using recombinant Semliki Forest virions and detected by introduced antigenic tags or antisera to PS1-derived peptides. Immunoprecipitation revealed two major PS1 bands of approximately 43 and 50 kDa, neither of which were N-glycosylated or 0-glycosylated. Immunoreactive PS1 was detected in cell bodies and dendrites of NT2N neurons but not in axons or on the cell surface. PS1 was also detected in BHK cells, where it was also intracellular and colocalized with calnexin, a marker for the rough endoplasmic reticulum. A mutant form of PS1 linked to FAD did not differ from the wild-type protein at the light microscopic level. The model system described here will enable studies of the function of PS1 in human neurons and the role of mutant PS1 in FAD.Since mutations in the PS1 and PS2 genes result in early onset FAD that is quite similar to FAD in other pedigrees with APP mutations, the PS mutations may cause AD by altering the metabolism of APP and/or AP3, thereby accelerating amyloidogenesis. Considerable evidence suggests that neurons are a major source of the AP3 that forms amyloid deposits in AD (17, 18), and a recent study showed that within the brain PS1 and PS2 mRNAs are expressed primarily in neurons (19). For these reasons, it is important to study the biosynthesis and processing of these genes in neurons. To accomplish this goal, we developed a strategy to efficiently express and detect PS1 in a well characterized in vitro model of human neurons (NT2N cells) (20). We found that PS1 was localized in the cell body and dendrites of NT2N neurons but not on the cell surface or in axons.

Light and Electron Microscopic Localization of Presenilin-1 in Primate Brain

“…In previous immunofluorescence studies, PS1 and PS2 were localized to ER and Golgi in transiently transfected neuroglioma cells overexpressing epitope-tagged proteins (Kovacs et al, 1996). More recently, PS1 overexpressed in a neuronal cell line infected with recombinant virions was localized to SER and rough ER by confocal microscopy (Cook et al, 1996). At the ultrastructural level, our antibody localizes the N terminus of native PS1 in monkey brain to the cytoplasmic face of membranous organelles in cell bodies and in the neuropil.…”

β‐Amyloid peptide induces formation of actin stress fibers through p38 mitogen‐activated protein kinase

“…These aggregates disrupt distal neurite function and are present in the hippocampus of patients with AD. Both APP and PS1 are richly expressed in dendritic spines (Cook et al 1996). Therefore, Ab peptide accumulates mostly around dendritic spins and synapses in AD.…”

Role of Endoplasmic Reticulum Stress and Unfolded Protein Responses in Health and Diseases

“…The b amyloid proteins are formed as a result of cleavage of amyloid precursor protein (APP) by the action of b-secretase (BACE) and !-secretase. ER stress sensors IRE1 and PERK are greatly influenced by presenilin protein, which is an integral membrane protein and a part of the !-secretase complex, and is widely expressed in both ER and Golgi apparatus [96]. Reports indicate that mutated presenilin reduces the phosphorylation of PERKeIF2a pathway which results in the accumulation of proteins in ER [97].…”