Serum Starvation Induces BACE1 Processing and Secretion

Background: beta-secretase (BACE1) is a type 1 transmembrane protein implicated in Alzheimer's Disease (AD) pathogenesis. Cleavage of Amyloid Precursor Protein (APP), initiated by BACE1 and followed by gamma-secretase, leads to the formation of toxic A beta peptides. Increased levels of BACE1 have been detected in the CSF of AD patients compared to age-matched healthy controls indicating that neurodegenerative conditions induce shedding of BACE1. Objective: To mimic such conditions, we examined whether serum deprivation stimulates proteolysis-dependent secretion of BACE1. Method: Detection of BACE1 secretion in BACE1 overexpressing cells or ADAM10/ADAM17 knockout fibroblasts cultured under serum deprivation conditions, using western blot analysis. Results: We found that serum deprivation of U251 neuroblastoma or HEK293T cells overexpressing BACE1 stimulated secretion of BACE1. Using ADAM10/ADAM17 knockout fibroblasts and inhibitors of both ADAM10 and ADAM17, we obtained data indicating that these proteases are involved in serum-starvation induced shedding of BACE1. This is unexpected since BACE1 is localized mainly in lipid rafts while ADAM10 is localized mainly in nonlipid raft domains. We hypothesized that serum deprivation results in alterations in the lipid composition of the membrane which can alter the localization of ADAM10 and BACE1. In support, we obtained results indicating that extraction of membrane cholesterol following incubation with methyl beta cyclodextrin potentiated the effect of serum deprivation. Secreted BACE1 was also found to be enzymatically active towards immunoprecipiated full length APP. Conclusion: Serum starvation induces ADAM10-mediated BACE1 secretion.