Asparagine endopeptidase cleaves tau at N167 after uptake into microglia

Background Tau cleavage by different proteolytic enzymes generates short, aggregation-prone fragments that have been implicated in the pathogenesis of Alzheimer’s disease (AD). Asparagine endopeptidase (AEP) activity in particular has been associated with tau dysfunction and aggregation, and the activity of the protease is increased in both aging and AD. Methods and Results Using a mass spectrometry approach we identified a novel tau cleavage site at N167 and confirmed its processing by AEP. In combination with the previously known site at N368, we show that AEP cleavage yields a tau fragment that is present in both control and AD brains at similar levels. AEP is a lysosomal enzyme, and our data suggest that it is expressed in microglia rather than in neurons. Accordingly, we observe tau cleavage at N167 and N368 after endocytotic uptake into microglia, but not neurons. However, tau168-368 does not accumulate in microglia and we thus conclude that the fragment is part of a proteolytic cascade leading to tau degradation. Conclusions While we confirm previous studies showing increased overall AEP activity in AD brains, our data suggests that AEP-mediated cleavage of tau is a physiological event occurring during microglial degradation of the secreted neuronal protein. The disease-associated increase in active AEP may thus be related to pro-inflammatory conditions in AD brains, and our findings argue against AEP inhibition as a therapeutic approach in AD. * aa : amino acid AAD : age at death AD : Alzheimer’s disease AEP : asparagine endopeptidase APP : amyloid precursor protein approx. : approximately COD : cause of death ctrl : control ELISA : Enzyme-linked immunosorbent assay fl : full-length HEK : human embryonic kidney cells iPSC : induced pluripotent stem cell kDa : kilodalton MW : molecular weight Ngn2 : Neurogenin 2 PMI : post-mortem interval p.p.m. : parts per million ThT : Thioflavin T wt : wild-type