Retrograde Axonal Autophagy and Endocytic Pathways Are Parallel and Separate in Neurons

Abstract

Autophagy and endocytic trafficking are two key pathways that regulate the composition and integrity of the neuronal proteome. Alterations in these pathways are sufficient to cause neurodevelopmental and neurodegenerative disorders. Thus, defining how autophagy and endocytic pathways are organized in neurons remains a key area of investigation. These pathways share many features and converge on lysosomes for cargo degradation, but what remains unclear is the degree to which the identity of each pathway is preserved in each compartment of the neuron. Here, we elucidate the degree of intersection between autophagic and endocytic pathways in axons of primary neurons. Using microfluidic chambers, we labeled newly-generated bulk endosomes and signaling endosomes in the distal axon, and systematically tracked their trajectories, molecular composition, and functional characteristics relative to autophagosomes. We find that newly-formed endosomes and autophagosomes both undergo retrograde transport in the axon, but as distinct organelle populations. Moreover, these pathways differ in their degree of acidification and association with molecular determinants of organelle maturation. These results suggest that the identity of autophagic and newly endocytosed organelles is preserved for the length of the axon. Lastly, we find that expression of a pathogenic form of -synuclein, a protein enriched in presynaptic terminals, increases merging between autophagic and endocytic pathways. Thus, aberrant merging of these pathways may represent a mechanism contributing to neuronal dysfunction in Parkinson's disease and related -synucleinopathies.

Dynein and dynactin move long-range but are delivered separately to the axon tip

“…1 D ). Also in agreement with previous work, endosomes and lysosomes moved primarily in the retrograde direction, whereas mitochondria displayed a high degree of bidirectional movement ( Boecker et al, 2020 ; Kulkarni et al, 2022 ) ( Fig. 1 E ).…”

Disruption of axonal transport in neurodegeneration

“…Further, expression of mutant LRRK2 in primary neurons reduced retrograde autophagic vesicle trafficking and impaired autophagosome maturation via recruitment of the kinesin adaptor JIP4 to increase kinesin association with autophagosomes to alter coordination of axonal trafficking ( 75 ). Intriguingly, while autophagosomes and endosomes are usually retrogradely transported in separate populations, pathogenic αSyn increased the overlap between these populations, indicating that PD may impair sorting of endosomes and autophagosomes as they undergo axonal trafficking ( 91 ).…”

Rab27b promotes lysosomal function and alpha-synuclein clearance in neurons

“…αSyn protein levels are enhanced at synapses, and evidence points to the initiation of αsyn pathology at the axon terminal [43][44][45][46], where degradative capacity is limited [47][48][49]. Protein clearance at distal terminals is dependent on the retrograde transport of autophagosomes that engulf misfolded proteins to the soma where they fuse with mature lysosomes [48][49][50]. Protein clearance is also dependent on the delivery of lysosomes to distal axons where local fusion with autophagic vesicles to initiate clearance occurs [47,49].…”