Sequalae and reversal of age-dependent alterations in mitochondrial dynamics via autophagy enhancement in...

How aging of human neurons affects dynamics of essential organelle such as mitochondria and autophagosomes remains largely unknown. MicroRNA-induced directly reprogrammed neurons (miNs) derived from adult fibroblasts retain age-associated signatures of the donor, enabling the study of age-dependent features in human neurons, including longitudinal isogenic samples. Transcriptomic analysis revealed that neurons derived from elderly individuals are characterized by gene expression changes associated with the regulation of autophagosomes, lysosomes, and mitochondria, compared to young counterparts. To clarify these changes at the cellular level, we performed live-cell imaging of cellular organelles in miNs from donors of different ages. Older donor miNs exhibit decreased mitochondrial membrane potential, which surprisingly co-occurs with a significant increase in mitochondrial fission and fusion events. We posit that the increased fission and fusion of mitochondria may reflect age-dependent compensation for impaired mitochondrial turnover, perhaps due to changes in macroautophagy/autophagy. We subsequently identified a significant decrease in autophagosome acidification in neurons derived from individuals > 65 years compared to younger donors, and a corresponding age-dependent reduction in neuritic lysosomes resulting in fewer lysosomes available to acidify autophagosomes. This age-dependent deficit in autolysosome flux was rescued by promoting autophagosome generation through TFEB, which also reversed the age-dependent increase in mitochondrial fission and fusion and improved mitochondrial health. Partial organelle recovery occurred after inducing mitophagy or inhibiting mitochondrial fission. Together, this work reveals a mechanism by which aging reduces autophagic flux secondary to a loss of neuritic lysosomes, resulting in mitochondria-intrinsic mechanisms to avoid loss of energy production.Abbreviations: DEG: differentially expressed gene; DMSO: dimethyl sulfoxide; Dox: doxycycline; iPSC: induced pluripotent stem cell; LAMP1: lysosome associated membrane protein 1; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MAPT/tau: microtubule associated protein tau; miNs: microRNA induced neurons; miRNA: microRNA; MMP: mitochondrial membrane potential; PID: post-infection day; RCAN1: regulator of calcineurin 1; TFEB: transcription factor EB; TUBB3: tubulin beta 3 class III; UA: urolithin A.