Recessive parkinsonism, mitochondria and translational regulation

Three genes are known to cause recessive forms of Parkinson disease (PD) in humans: parkin, PINK1 and DJ-1. Of these, the rarest is DJ-1; less than fifty known cases worldwide are due to mutations in DJ-1. Though rare, elucidating the function of DJ-1 and other recessive parkinsonism genes is crucial to understanding pathways involved in pathogenesis. DJ-1 protects cells from oxidative stress in numerous models, an activity likely related to its ability to form a cysteine-sulfinic acid under mildly oxidizing conditions.

In the work described in this thesis, the cellular properties and activities of DJ-1 were investigated with the intent of uncovering the underlying mechanism connecting DJ-1 oxidation to cellular protection. Two pathogenic mutants of DJ-1, M26I and L166P, were found to be unstable in cells, thus accounting for their loss-of-function. Both these and stable pathogenic mutants showed increased mitochondrial localization under oxidative stress.

We also designed mutations to manipulate the oxidative properties of DJ-1 and observed that formation of a cysteine-sulfinic acid at amino acid cysteine 106 was critical for both mitochondrial localization and protection of mitochondria by DJ-1 as indicated by fragmentation of mitochondrial networks. Fragmentation observed in DJ-1 deficient cells was exacerbated by adding oxidative stressors and counteracted by increasing intracellular levels of the antioxidant glutathione, suggesting the mitochondrial phenotype was driven by misregulation of oxidation responses. Expression of other recessive parkinsonism genes, PINK1 or parkin, fully rescued the phenotype, linking all three genes into a single pathway. Since the three genes must all be expressed in substantia nigra neurons to work in concert, in situ hybridization was performed for PINK1 in human and rodent brain to complement previous observations of DJ-1 and parkin expression. Finally, the RNA interaction properties of DJ-1 were investigated.

An interaction between DJ-1 and specific mRNA targets was observed and confirmed using multiple methods. These RNA binding targets included mitochondrial and nuclear encoded components of the oxidative phosphorylation pathway, selenoproteins and other antioxidant proteins including glutathione peroxidases, and components of the PTEN/Akt cell survival pathway. Pathogenic mutants of DJ-1 were deficient in RNA binding activity. This interaction of DJ-1 and RNA was oxidation dependent, as was translational regulation of targets. Increased translation of DJ-1 targets correlated with increased oxidation of DJ-1 in sporadic PD, implicating DJ-1 in the response to PD pathogenesis. This thesis therefore proposes that DJ-1 translationally regulates a localized oxidative stress response that is particularly important in protecting mitochondria.

List of scientific papers

I. Blackinton J, Ahmad R, Miller DW, van der Brug MP, Canet-Avilés RM, Hague SM, Kaleem M, Cookson MR (2005). "Effects of DJ-1 mutations and polymorphisms on protein stability and subcellular localization." Brain Res Mol Brain Res 134(1): 76-83
https://pubmed.ncbi.nlm.nih.gov/15790532

II. Blackinton J, Lakshminarasimhan M, Thomas KJ, Ahmad R, Raza AS, Cookson MR, Wilson MA (2008). "Formation of a stabilized cysteine sulfinic acid is critical for the mitochondrial function of the parkinsonism protein DJ-1." (Manuscript)

III. Blackinton J, Thomas KJ, van der Brug M, Beilina A, Miller D, Cookson MR (2008). "Increased oxidative stress in DJ-1 deficient cells leads to mitochondrial fission." (Manuscript)

IV. Blackinton JG, Anvret A, Beilina A, Olson L, Cookson MR, Galter D (2007). "Expression of PINK1 mRNA in human and rodent brain and in Parkinsons disease." Brain Res 1184: 10-6. Epub 2007 Sep 29
https://pubmed.ncbi.nlm.nih.gov/17950257

V. van der Brug MP, Blackinton J, Chandran J, Hao LY, Lal A, Mazan-Mamczarz K, Martindale J, Xie C, Ahmad R, Thomas KJ, Beilina A, Gibbs JR, Ding J, Myers AJ, Zhan M, Cai H, Bonini NM, Gorospe M, Cookson MR (2008). "RNA binding activity of the recessive parkinsonism protein DJ-1 supports involvement in multiple cellular pathways." Proc Natl Acad Sci U S A 105(29): 10244-9. Epub 2008 Jul 14
https://pubmed.ncbi.nlm.nih.gov/18626009

VI. Blackinton J, van der Brug MP, Kumaran R, Ahmad R, Olson L, Galter D, Lees A, Bandopadhyay R, Cookson MR (2008). "Post-transcriptional regulation of mRNA associated with DJ-1 in Sporadic Parkinson disease." (Manuscript)