Investigating the impact of alterations in alpha-synuclein on mitochondrial health, mitochondrial quality control and neuronal differentiation

Abstract

Mitochondrial quality control (mito-QC) co-ordinates dynamic changes in the mitochondrial network that maintain optimal function and cell health. Dysregulated mito-QC has been associated with numerous pathologies, including Parkinson’s disease (PD) which is characterised by degeneration of dopaminergic neurons in the substantia nigra pars compacta (SNpc). The mechanisms of neuronal death in PD are still unknown, but have been linked to the aggregation of a-synuclein; a protein which pathologically forms intracellular inclusions, known as Lewy bodies, in PD brains. A relationship between a-synuclein and mito-QC has been established based on the presence of dysfunctional mitochondria in both PD patients and models of a-synuclein pathology. However, exactly how a-synuclein may impact mito-QC mechanisms is not well understood. Recent research has shown that a-synuclein can use mitochondrial membranes to seed aggregation of toxic oligomeric species, resulting in mitochondrial dysfunction and potentiating disease pathogenesis. Though the majority of research has focused on the hypothesis that pathological a-synuclein may exhibit a toxic gain-of-function that disrupts mito-QC, it is also possible that this could occur through a-synuclein loss-of-function. Since a-synuclein associates with and remodels phospholipid membranes, has been suggested to contain a mitochondrial targeting sequence and can be mitochondrially imported, it may have a physiological role in mito-QC that is lost under pathological circumstances.
In an a-synuclein loss-of-function model, I observe that physiological a-synuclein is not required for mitochondrial energy production or co-ordination of mito-QC. I do not find evidence of a direct functional association between endogenous a-synuclein and mitochondria, however I suggest that a-synuclein may modulate autophagy flux and lysosomal morphology through its action as a cytosolic molecular chaperone. This provides novel insight into a-synuclein’s physiological function in the context of mito-QC. I also investigated the impact of pathological a-synuclein on mito-QC, where I found that a-synuclein overexpression did not disrupt mitochondrial function or damage-induced whole mitochondrial clearance, but did influence autophagosome number and lysosomal size. Since alterations in these phenotypes were also seen in my loss-of-function model, I consider that physiological a-synuclein may have a role in modulating autophagy that is disrupted in pathological circumstances such as PD.Overexpression of ³-synuclein also increased mitochondrial fragmentation and mitochondrial-derivedvesicles (MDVs) at steady state, suggesting that excess ³-synuclein increases the baseline mito-QC response. I suggest that upregulation of these responses may occur due to an a-synuclein-induced increase in oxidative stress. Together, these data indicate that pathological a-synuclein, rather than a lack of physiological a-synuclein, is primarily responsible for dysregulated mitochondrial function in PD.
Since a-synuclein’s pathological impact primarily occurs in dopaminergic neurons of the SNpc, it would be important to further investigate interactions between a-synuclein and mito-QC in a model that better recapitulates this neuronal environment. Accordingly, I generated a neuronal model through differentiation of SH-SY5Y cells that can be used for future work. In this study, I characterised how perturbations in a-synuclein expression impacted neuronal phenotype in differentiated cells, finding that overexpression of a-synuclein A53T, but not wild-type, reduced neuronal morphology and expression of neuronal markers. Interestingly, a-synuclein knockout cells showed a similar but less exaggerated phenotype, suggesting that defects in neuronal differentiation in both cell lines may be explained by loss of physiological a-synuclein activity. This provides insight into a-synuclein’s role in neuronal development.

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Identifiers

ORCID for Naomi Jasmin Thorne: orcid.org/0000-0001-5364-0253

ORCID for David Tumbarello: orcid.org/0000-0002-5169-0561

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