Table 1.
Summary of the experiments discussed and a categorization of the evidence as direct or indirect
| Hypothesis | Experiment(s) | Conclusion | Comments | Proposed experiment(s) |
|---|---|---|---|---|
| α-synuclein interacts with and influences microtubule stability Direct | In vitro MT assembly assay with and without α-synuclein, sedimentation assay3 | α-synuclein increases the polymerization of MTs | No clarification on whether the turbidity of the solution or insoluble pellet was caused by polymerization of tubulin and not aggregation | Repeating sedimentation assay with the additional step of viewing the final pellet under electron microscopy |
| Overexpression of α-synuclein and analysis of MTs via fluorescence microscopy,3 acetylation,4 and Golgi fragmentation5 | α-synuclein has a role in the destabilization of microtubules | No MT disruption found in neurons treated with α-synuclein in PFF form: this may be due to the different form of α-synuclein used in this experiment6 | Repeat experiment using PFFs of α-synuclein instead of overexpressing it | |
| α-synuclein acts as an MT dynamase Direct | Turbidity assay with α-synuclein and PD-associated mutants, followed by electron microscopy7 | α-synuclein promotes microtubule polymerization in vitro | Similar experiment showed no effect of α-synuclein on the polymerization of MTs: likely due to procedural differences8 | |
| α-synuclein promotes tau phosphorylation via GSK-3β Indirect | Co-immunoprecipitation of α-synuclein and GSK-3β21 | α-synuclein interacts with GSK-3β | Co-immunoprecipitation does not prove the proteins interact | Use FRET to see whether α-synuclein and GSK-3β interact in vivo |
| α-synuclein promotes the formation of aggregates Indirect | Overexpression of α-synuclein in mice and immunohistochemical staining for α-synuclein, P-tau and pGSK-3β19 | α-synuclein overexpression results in the formation of aggregates that resemble Lewy bodies | No evidence of tauopathy or synucleinopathy was found in a similar model overexpressing α-synuclein under a different promoter25 | |
| α-synuclein acts synergistically with tau to cause neurodegeneration via tau phosphorylation Indirect | α-synuclein and tau overexpressed in dopaminergic neurons and the phosphorylation state of tau analyzed using antibodies30 | α-synuclein has no effect on tau phosphorylation | Only a few phosphorylation sites were targets of the antibody used to analyze tau phosphorylation | Repeat experiment using antibodies recognizing tau phosphorylated at Ser262 or Ser396, the primary targets of PKA and GSK-3β, respectively |
Abbreviations: FRET, fluorescence resonance energy transfer; MTs, microtubules; PD, Parkinson’s disease; PFF, preformed fibril; P-tau, phosphorylated tau.