Introduction: Parkinson’s Disease (PD) is the 2nd most prevalent neurodegenerative disease. Several genes have been linked with PD including DJ-1 which is associated with the rarer monogenic form of PD. Numerous roles and protein locations have been ascribed to DJ-1 including a role in protection against oxidative stress and localisation to the mitochondria with some conflicting results. We used a simple eukaryotic model, Dictyostelium discoideum, to investigate the role of DJ-1 in mitochondrial function, oxidative stress and how mutations in this protein may lead to PD.
Methodology: I created a range of D. discoideum DJ-1 mutant cell lines including antisense inhibition, overexpression, expression of a C117A mutation (its oxidisable cysteine) and expression of mitochondrially targeted DJ-1 protein. I exposed the cells to oxidative stress to determine if the oxidisable cysteine and localisation was essential for DJ-1’s ‘protective’ roll in cells. Immunofluorescence microscopy and western blots were used to assess localisation and expression of the protein, and Seahorse Respirometry was used to determine whether DJ-1 played a role in mitochondrial function. We also analysed the cell lines phenotypically to determine the normal roles of DJ-1.
Major findings: DJ-1 reduces mitochondrial respiration and has a positive effect on growth. DJ-1 is cytoplasmic and does not localise to the mitochondria regardless of oxidative stress. When DJ-1 was forcibly targeted to the mitochondria, cells mimicked overexpression phenotypes suggesting some potential benefits from mitochondrial localisation. Additionally, DJ-1 C117A mutants have increased sensitivity to oxidative stress and this mutation appears to cause loss of function.
Conclusion: DJ-1 has roles in mitochondrial function and growth and is involved in protection against the consequences of oxidative stress that is dependent on C117. D. discoideum has proven to be a useful model in which to further characterise DJ-1’s normal role.