Conversely we find that dysfunction

Conversely, we find that dysfunction in lysosomal-autophagy system observed in human APP and PSEN1 mutant neurons can be reversed by β-secretase inhibition, which reduces the supply of APP-β-CTF to the late endosome and/or lysosome (Jiang et al., 2010). These data argue that accumulation and/or altered turnover of APP-CTF by γ-secretase lead to dysfunction in the neuronal lysosomal-autophagy system. Importantly, the rescue of lysosomal-autophagy defects by CRISPR deletion of APP in a PSEN1 mutant background formally demonstrates that those defects are dependent on APP protein, and not simply convergent independent phenotypes due to mutations in either gene. The lack of detectable defects in lysosome and autophagosome function in PSEN1 Intron4 neurons is surprising, given that these neurons have similar changes in Aβ peptide production as the other PSEN1 mutant neurons. However, the molecular nature of the PSEN1 Intron4 mutation is qualitatively different from the majority of PSEN1 mutations, which are missense protein-coding mutations that result in single amino Etomoxir substitutions (De Jonghe et al., 1999). The PSEN1 Intron4 mutation is a single base deletion within the splice donor site, which has been predicted to generate three different final mRNAs, two of which encode truncated forms of PSEN1, and the third includes an extra amino acid (De Jonghe et al., 1999). If this is the case in cortical neurons, the relative amounts of wild-type and full-length mutant PSEN1 proteins would be much less than 1:1, and relatively higher levels of the wild-type form of PSEN1 might enable short-term compensation for the deleterious effects of the mutant protein. Therefore, it will be essential to further study the effects of this mutation on lysosome and autophagosome function over longer time frames and in neurons homozygous for this mutation. The early stage of axonal lysosome biogenesis is thought to begin in distal regions of axons with the merging of organelles derived from endocytic and autophagic pathways (Maday and Holzbaur, 2014). Further maturation of lysosomes requires retrograde transport to the cell soma where luminal proteases can be delivered from the trans-Golgi network efficiently (Maday et al., 2012). Such a mechanism allows neurons to effectively remove autophagic cargos from axon terminals and synapses. We found that AD mutations in APP and PSEN1 markedly disrupted the axonal transport of lysosomes and those lysosomes accumulate within distal axons contained low levels of cathepsin D. This is similar to previous findings in human AD brain tissue as well as various AD mouse models in which focal axonal swellings containing accumulations of lysosomal dense bodies and autophagic vacuoles have been observed (Nixon, 2017). Although autophagy activation may be a promising therapeutic strategy for AD, its beneficial role in disease pathogenesis is not clear. Several studies showed that increasing autophagy in mouse models of AD ameliorates amyloid pathologies and memory deficits (Spilman et al., 2010, Yang et al., 2011). However, autophagy induction has also been reported to activate the amyloidogenic pathway and increase Aβ secretion (Yu et al., 2005, Nilsson et al., 2013). Similarly, administration of rapamycin after the formation of amyloid plaques failed to rescue the cognitive deficits in mouse model of AD (Majumder et al., 2011). Our data suggest that induction of autophagy may become counterproductive in the face of impaired transport of axonal lysosomes, lysosomal clearance deficits and reduced autophagosome degradation, by further overburdening the already failing lysosomes and thus exacerbating autophagic accumulation in axons. Indeed, recent studies have demonstrated that restoring retrograde transport of autophagic vacuoles by overexpressing Sanpin proteins successfully reduced AD-associated autophagic stress (Tammineni et al., 2017). The present study on human cortical neurons provides further evidence that either reducing input of APP to the lysosomal-autophagy system, enhancing axonal transport or augmenting lysosome function at the early disease stages may represent potential therapeutic strategies to attenuate autophagic defects in AD. Overall, this study establishes a foundation for future investigation into cellular pathways enhancing autophagy and lysosomal proteolytic activity as an approach to ameliorating neurodegeneration in AD.