BACE1 Overexpression Reduces SH-SY5Y Cell Viability Through a Mechanism Distinct from Amyloid-beta Peptide Accumulation: Beta Prime-Mediated Competitive Depletion of sA beta PP alpha

Background: The Alzheimer's disease (AD)-associated amyloid-beta protein precursor (A beta PP) can be cleaved by beta-site A beta PP cleaving enzyme 1 (BACE1) and the gamma-secretase complex to yield neurotoxic amyloid-beta (A beta) peptides. However, A beta PP can also be cleaved in a 'non-amyloidogenic' manner either by beta-secretase to produce soluble A beta PP alpha (sA beta PP alpha) (a fragment with neuroprotective/neurogenic functions) or through alternative BACE1-mediated `beta prime' activity yielding soluble A beta PP beta prime (sA beta PP alpha'). Objective: To determine whether sA beta PP alpha depletion, as opposed to A beta peptide accumulation, contributes to cytotoxicity in AD-relevant SH-SY5Y neuroblastoma cell models. Methods: A beta PP proteolysis was characterized by immunoblotting in mock-, wild-type A beta PP (wtA beta PP)-, BACE1-, and Swedish mutant A beta PP (SweA beta PP)-transfected cells. A beta PP beta prime cleavage was confirmed through secretase inhibitor studies and C-terminal fragment analysis. The roles of sA beta PP alpha and sA beta PP alpha' in cell viability were confirmed by overexpression studies. Results: Despite producing enhanced A beta peptide levels, wtA beta PP- and SweA beta PP-transfected cells did not exhibit reduced viability whereas BACE1-transfected cells did. sA beta PP alpha generation in SH-SY5Y-BACE1 cells was virtually ablated in lieu of BACE1-mediated sA beta PP alpha' production. sA beta PP alpha overexpression in SH-SY5Y-BACE1 cells restored viability whereas sA beta PP alpha' overexpression decreased viability further. The anti-A beta PP 6E10 antibody was shown to cross-react with sA beta PP alpha'. Conclusion: sA beta PP alpha depletion and/or sA beta PP alpha' accumulation, but not elevated A beta peptide levels, represent the cytotoxic mechanism following BACE1 overexpression in SH-SY5Y cells. These data support the novel concept that competitive sA beta PP alpha depletion by BACE1 beta prime activity might contribute to AD. The cross-reactivity of 6E10 with A beta PP alpha'also questions whether previous studies assessing sA beta PP alpha as a biomarker using this antibody should be revisited.