Autophagy is a conserved, multi-step process of capturing proteolytic cargo in autophagosomes for lysosome degradation. The capacity to remove toxic proteins that accumulate in neurodegenerative disorders attests to the disease-modifying potential of the autophagy pathway. However, neurons respond only marginally to conventional methods for inducing autophagy, limiting efforts to develop therapeutic autophagy modulators for neurodegenerative diseases. The determinants underlying poor autophagy induction in neurons and the degree to which neurons and other cell types are differentially sensitive to autophagy stimuli are incompletely defined. Accordingly, we sampled nascent transcript synthesis and stabilities in fibroblasts, induced pluripotent stem cells (iPSCs), and iPSC-derived neurons (iNeurons), thereby uncovering a neuron-specific stability of transcripts encoding myotubularin-related phosphatase 5 (MTMR5). MTMR5 is an autophagy suppressor that acts with its binding partner, MTMR2, to dephosphorylate phosphoinositides critical for autophagy initiation and autophagosome maturation. We found that MTMR5 is necessary and sufficient to suppress autophagy in iNeurons and undifferentiated iPSCs. Using optical pulse labeling to visualize the turnover of endogenously encoded proteins in live cells, we observed that knockdown of MTMR5 or MTMR2, but not the unrelated phosphatase MTMR9, significantly enhances neuronal degradation of TDP-43, an autophagy substrate implicated in several neurodegenerative diseases. Our findings thus establish a regulatory mechanism of autophagy intrinsic to neurons and targetable for clearing disease-related proteins in a cell-type-specific manner. In so doing, our results not only unravel novel aspects of neuronal biology and proteostasis but also elucidate a strategy for modulating neuronal autophagy that could be of high therapeutic potential for multiple neurodegenerative diseases.

Accumulation of the tau protein in fibrillar intracellular aggregates is a defining feature of multiple neurodegenerative diseases collectively referred to as tauopathies. Despite intensive study of tau, there is limited information on the formation and clearance dynamics of tau inclusions. Using rAAV vectors to mediate expression of Dendra2-tagged human wild-type, P301L and pro-aggregant P301L/S320F tau proteins, with and without the addition of exogenous tau fibrillar seeds, we evaluated tau inclusion dynamics in organotypic brain slice culture (BSC) models using long-term optical pulse labeling methodology. Our studies reveal that tau inclusions typically form in 12-96 h in tauopathy BSC models. Unexpectedly, we demonstrate appreciable turnover of tau within inclusions with an average half-life of ~ 1 week when inclusions are newly formed. When BSCs with inclusions are aged in culture for extended periods, tau inclusions continue to turnover, but their half-lives increase to ~ 2 weeks and ~ 3 weeks after 1 and 2 months in culture, respectively. Individual tau inclusions can be long-lived structures that can persist for months in these BSC models and for even longer in the human brain. However, our data indicate that tau inclusions, are not \'tombstones\', but dynamic structures with appreciable turnover. Understanding the cellular processes mediating this inclusion turnover may lead to new therapeutic strategies that could reverse pathological tau inclusion formation.