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P1‐291: Functional studies of SNPs in the mitochondrial Aβ‐degrading protease, hPreP
Author(s) -
Pinho Catarina M.,
Alikhani Nyosha,
Graff Caroline,
Glaser Elzbieta
Publication year - 2010
Publication title -
alzheimer's and dementia
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 6.713
H-Index - 118
eISSN - 1552-5279
pISSN - 1552-5260
DOI - 10.1016/j.jalz.2010.05.843
Subject(s) - single nucleotide polymorphism , biology , snp , mitochondrion , genetics , genotyping , mitochondrial dna , protease , snp genotyping , genetic association , gene , proteolysis , population , genotype , microbiology and biotechnology , enzyme , biochemistry , medicine , environmental health
formation of the amyloid b (Ab) peptide. Co-immunoprecipitation studies have demonstrated that PrP associates with BACE1. To further establish the molecular mechanism of the interaction, the effect of PrP on BACE1 cellular localisation was investigated in cells and a direct interaction explored in vitro. Methods: SH-SY5Y cells co-expressing PrP with BACE1 and HEK cells co-expressing PrP with BACE1 or BACE1 pro-domain mutants were established. Immunofluorescence microscopy and FACS were used to assess BACE1 localisation in SH-SY5Y cells. Isolated membrane preparations and concentrated media from the HEK cells were subjected to immunoblot analysis and antibodies specific to BACE1, PrP, APP, actin and sAPPb were used. Interaction studies between recombinant BACE1 and PrP were carried out in an ELISA format and by surface plasmon resonance. Results: BACE1 and PrP co-localised and the localisation of BACE1 to the optimal site of Ab production within endosomes was reduced in the presence of PrP. Localisation of BACE1 to the cell surface was also reduced, with an increased localisation of BACE1 to the trans-Golgi network (TGN). A direct interaction between recombinant BACE1 and PrP was confirmed and this interaction was mediated via the pro-domain of BACE1. As PrP also reduces the BACE2 processing of APP, a conserved region in the pro-domain of BACE1 and BACE2 was explored as a potential site of interaction with PrP. The BACE1 pro-domain mutant (P29G) restored sAPPb levels back to control levels, suggesting that PrP mediates its effects on APP processing by interacting with the BACE1 pro-domain within this conserved region. Conclusions: These data provide a better understanding of the mechanism by which PrP inhibits the action of BACE1, and the role that PrP plays in the pathogenesis of Alzheimer’s disease.