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The Non-amyloidal Component Region of ?-Synuclein Is Important for ?-Synuclein Transport Within Axons.

ABSTRACT: Proper transport of the Parkinson's disease (PD) protein, ?-synuclein (?-syn), is thought to be crucial for its localization and function at the synapse. Previous work has shown that defects in long distance transport within narrow caliber axons occur early in PD, but how such defects contribute to PD is unknown. Here we test the hypothesis that the NAC region is involved in facilitating proper transport of ?-syn within axons via its association with membranes. Excess ?-syn or fPD mutant ?-syn^A53T accumulates within larval axons perturbing the transport of synaptic proteins. These ?-syn expressing larvae also show synaptic morphological and larval locomotion defects, which correlate with the extent of ?-syn-mediated axonal accumulations. Strikingly, deletion of the NAC region (?-syn^?71-82) prevented ?-syn accumulations and axonal blockages, and reduced its synaptic localization due to decreased axonal entry and axonal transport of ?-syn, due to less ?-syn bound to membranes. Intriguingly, co-expression ?-syn^?71-82 with full-length ?-syn rescued ?-syn accumulations and synaptic morphological defects, and decreased the ratio of the insoluble higher molecular weight (HMW)/soluble low molecular weight (LMW) ?-syn, indicating that this region is perhaps important for the dimerization of ?-syn on membranes. Together, our observations suggest that under physiological conditions, ?-syn associates with membranes via the NAC region, and that too much ?-syn perturbs axonal transport via aggregate formation, instigating synaptic and behavioral defects seen in PD.

SUBMITTER: Anderson EN

PROVIDER: S-EPMC6984405 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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The Non-amyloidal Component Region of α-Synuclein Is Important for α-Synuclein Transport Within Axons.

Anderson Eric N EN Hirpa Delnessaw D Zheng Kan Hong KH Banerjee Rupkatha R Gunawardena Shermali S

Frontiers in cellular neuroscience 20200110

Proper transport of the Parkinson's disease (PD) protein, α-synuclein (α-syn), is thought to be crucial for its localization and function at the synapse. Previous work has shown that defects in long distance transport within narrow caliber axons occur early in PD, but how such defects contribute to PD is unknown. Here we test the hypothesis that the NAC region is involved in facilitating proper transport of α-syn within axons <i>via</i> its association with membranes. Excess α-syn or fPD mutant ...[more]

PMID: 32038170

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Project description:BACKGROUND: Alpha-synuclein is a presynaptic protein with a proposed role in neurotransmission and dopamine homeostasis. Abnormal accumulation of alpha-synuclein aggregates in dopaminergic neurons of the substantia nigra is diagnostic of sporadic Parkinson's disease, and mutations in the protein are linked to early onset forms of the disease. The folded conformation of the protein varies depending upon its environment and other factors that are poorly understood. When bound to phospholipid membranes, alpha-synuclein adopts a helical conformation that mediates specific interactions with other proteins. RESULTS: To investigate the role of the helical domain in transport and localization of alpha-synuclein, eGFP-tagged constructs were transfected into rat primary hippocampal neurons at 7 DIV. A series of constructs were analyzed in which each individual exon was deleted, for comparison to previous studies of lipid affinity and alpha-helix content. A53T and A30P substitutions, representing Parkinson's disease-associated variants, were analyzed as well. Single exon deletions within the lipid-binding N-terminal domain of alpha-synuclein (exons 2, 3, and 4) partially disrupted its presynaptic localization at 17-21 DIV, resulting in increased diffuse labeling of axons. Similar results were obtained for A30P, which exhibits decreased lipid binding, but not A53T. To examine whether differences in presynaptic enrichment were related to deficiencies in transport velocity, transport was visualized via live cell microscopy. Tagged alpha-synuclein migrated at a rate of 1.85 +/- 0.09 mum/s, consistent with previous reports, and single exon deletion mutants migrated at similar rates, as did A30P. Deletion of the entire N-terminal lipid-binding domain (Delta234GFP) did not significantly alter rates of particle movement, but decreased the number of moving particles. Only the A53TGFP mutant exhibited a significant decrease in transport velocity as compared to ASGFP. CONCLUSIONS: These results support the hypothesis that presynaptic localization involves a mechanism that requires helical conformation and lipid binding. Conversely, the rate of axonal transport is not determined by lipid affinity and is not sufficient to account for differences in presynaptic localization of alpha-synuclein-eGFP variants.

Dataset Information

The Non-amyloidal Component Region of ?-Synuclein Is Important for ?-Synuclein Transport Within Axons.

Publications

The Non-amyloidal Component Region of α-Synuclein Is Important for α-Synuclein Transport Within Axons.

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