The Genes of Parkinson’s Disease

For most of the twentieth century, genetic predisposition was thought to play a negligible role in the disease. The strongest evidence was epidemiological and seemed to point to environmental factors such as pesticides, which both selectively kill dopamine-producing neurons and induce Parkinson’s-like symptoms. It wasn’t until 1997 that the debate opened up and researchers started to consider the genetics of the disease.

While a hereditary cause can still only be linked to about 5 percent of Parkinson’s cases, these genes are starting to shed light on the universal processes that might cause dopaminergic neurons to die. By probing these genes, a new story is beginning to emerge—one that suggests environmental factors may exacerbate disease in individuals who are susceptible, because genetic factors curtail their ability to cope with toxic damage. Indeed, many of the genes mutated in hereditary Parkinson’s disease play a pathogenic role in mitochondria as well as in the ubiquitin proteasome system and the autophagy-lysosome machinery, both of which remove misfolded proteins and damaged organelles. However, some of the implicated proteins have functions that do not necessarily fit into these scenarios, so more work is needed to understand whether and how the various disease-associated genes interact. Recent studies have identified Parkinson’s-related genes containing either gain-of-function mutations (those that generate a gene product with a novel function) or loss-of-function mutations (those that render the gene product nonfunctional). These genes demonstrate that we are getting closer to understanding the mechanisms of neuronal damage in Parkinson’s disease.

Gain-of-function mutations

The a-synuclein gene mutation. The first big breakthrough came in 1997 when researchers were studying several families with the inherited dominant form of Parkinson’s disease—a form that required a toxic gain-of-function mutation in the a-synuclein gene locus (reviewed in reference [1. B. Thomas, M.F. Beal, “Parkinson’s disease,” Hum Mol Genet, 16:R183-94, 2007.]). Later, a-synuclein was identified as a major component of Lewy bodies, the fibrous bundles within neurons seen in autopsied brains of Parkinson’s patients. Normally, the a-synuclein protein regulates the release of neurotransmitters; however, mutations to the a-synuclein gene, SNCA—both point mutations and multiplications—occurred in these familial cases of Parkinson’s disease, pointing to a role for this gene in PD. The finding also suggested that individuals with no mutations, but who simply produced a higher than average level of a-synuclein, had a greater risk of developing PD than those with a lower a-synuclein levels.[2. S. Winkler et al., “a-Synuclein and Parkinson disease susceptibility,” Neurology, 69:1745-50, 2007.]

A colored transmission electron micrograph (TEM) of a neuronal section containing a Lewy body made up of a-synuclein filaments in pink and blue.
LYSIA FORNO / PHOTO RESEARCHER, INC.

Both pathogenic mutations in the gene and elevated concentrations of the protein will give a-synuclein a propensity to develop a ß-sheet structure. These structures readily polymerize into oligomers and higher-order aggregates, such as the fibrils that are characteristic of Lewy bodies.¹ Not only do genetic alterations instigate disease, but a-synuclein aggregation is also worsened by various types of post-translational modifications to the normal protein,[3. A. Oueslati et al., “Role of post-translational modifications in modulating the structure, function and toxicity of a-synuclein: Implications for Parkinson’s disease pathogenesis and therapies,” Prog Brain Res, 183:115-45, 2010.]potentially expanding the number of factors involved in propagating Parkinson’s.

Parkinson’s researchers began to look beyond the genes that code for a-synuclein. For example, when investigators inhibited mitochondrial complex I—a protein critical to the mitochondrial electron transport that powers the cell—a-synuclein began to form aggregates and became toxic to the cell. The results suggested that disrupted mitochondria might themselves spur a-synuclein bundle formation and, in theory, also cause Parkinson’s disease.[4. R. Banerjee et al., “Mitochondrial dysfunction in the limelight of Parkinson’s disease pathogenesis,” Biochim Biophys Acta, 1792:651-63, 2009.]

Curiously, however, researchers also found evidence that a-synuclein overexpression induces mitochondrial dysfunction, resulting in electron leakage that causes oxidative stress in the cell, ultimately leading to cell death.[5. L.J. Hsu et al., “a-Synuclein promotes mitochondrial deficit and oxidative stress,” Am J Pathol, 157:401-10, 2000.] It’s unclear whether the chicken comes before the egg or vice versa: is it the overabundance of a-synuclein that damages the mitochondria, or is it a damaged mitochondrial electron pump mechanism that initiates the process that leads to a-synuclein bundling? There appears to be evidence for both possibilities.

It wasn’t until 1997 that researchers started to consider the genetics of Parkinson’s disease.

With a-synuclein a proven player, researchers started working on explaining how the mutations that either changed the shape of the protein or increased its expression might cause neuronal death. These investigations have yielded no shortage of hypotheses. Some studies suggest that a-synuclein oligomers bound together in a doughnut shape might form pores in the plasma membrane, allowing calcium ions to accumulate in the cell at toxic levels.[6. K. Furukawa et al., “Plasma membrane ion permeability induced by mutant a-synuclein contributes to the degeneration of neural cells,” J Neurochem, 97:1071-77, 2006.] (The idea isn’t so far-fetched, considering that a-synuclein monomers alone readily associate with the plasma membrane.) Yet others suggest that an overabundance of a-synuclein tends to gunk up a neuron’s ability to release and recycle neurotransmitters that are stored in the vesicles of firing neurons.[7. V.M. Nemani et al., “Increased expression of alpha-synuclein reduces neuro- transmitter release by inhibiting synaptic vesicle reclustering after endocytosis,” Neuron, 65:66-79, 2010. Free F1000 Evaluation] (See graphic below.) The thinking is that the buildup of this protein interferes with synaptic transmission by preventing docked vesicles at the presynaptic membrane from releasing into the synaptic space. This reduces the overall dopamine being transmitted by these neurons, thereby increasing intracellular dopamine to toxic levels and permanently damaging the dopamine-producing cells.[8. A.A. Cooper et al., “a-Synuclein blocks ER-Golgi traffic and Rab1 rescues neuron loss in Parkinson’s models,” Science, 313:324-28, 2006.]

Finally, some research suggests that it is not an overabundance of a-synuclein, but rather the mutant forms of the protein that cause damage by throwing several monkey wrenches into the cell’s ability to get rid of all misfolded proteins. Mutant species of a-synuclein are poor substrates for degradation by the proteasome, the cell’s recycling complex; they inhibit the digestion of proteins by proteolysis; they block lysosomes, which fuse with other vesicles to break up their contents as well as blocking the vesicles that are tagged with chaperone proteins for fusion with lysosomes during autophagy⁸; and they disrupt ER-Golgi trafficking, which is vital to vesicle transport and a number of secretory pathways.

Are all of these mechanisms triggered when a-synuclein becomes abundant or mutated in the cell? Is there a hierarchy—one mechanism that predominates over the others? Would we need to disable all of them in order to prevent or slow disease in susceptible individuals? These are still the big questions that require further testing, but whatever the exact mechanism(s) involved, it is clear that a-synuclein aggregates exert toxic effects on several important cellular functions necessary for the survival of dopaminergic neurons.

The LRRK2 gene mutation. A second autosomal dominant Parkinson’s gene involving a toxic gain-of-function mutation was discovered in 2004 and named LRRK2 (leucine-rich repeat kinase 2). Mutations in this gene are found in about 5-6 percent of all familial cases as well as 2 percent of cases with no known cause. Interestingly, this mutation can cause early-onset Parkinson’s in families from diverse ethnic backgrounds,¹ in a form that is identical in clinical symptoms to late-onset Parkinson’s. LRRK2 encodes a protein that is part of a larger multidomain protein with characteristic GTPase and kinase domains. While a great deal of effort has gone into identifying the protein’s normal physiological functions in the cell, only a few of its substrates—which appear to promote neuron growth by activating protein translation or cytoskeletal reorganization—have so far been tentatively identified. Most of LRRK2’s substrates, its binding partners and its regulators have yet to be confirmed or clarified, and consequently its role in disease is still a mystery.

Many of the LRRK2 transgenic mouse models of PD develop cardinal abnormalities of the disease, such as stimulated dopamine neurotransmission or behavioral deficits, and progressive age-dependent motor deficits leading to immobility that are responsive to the dopaminergic drugs L-DOPA and apomorphine.[9. T.M. Dawson et al., “Genetic animal models of Parkinson’s disease,” Neuron, 66:646-61, 2010.]Strikingly, there is an additive effect when both LRRK2 and the a-synuclein gene are mutated in mice, whereas a deficiency of LRRK2 limits the toxic effects of mutant a-synuclein, suggesting that the two proteins are involved in common disease development pathways.

Infographic: Death or Damage of Dopamine Neurons
View full size JPG | PDF

LUCY READING-IKKANDA

Recently, investigators were able to determine that in vivo, mutated LRRK2 exerted toxicity resulting from its enhanced kinase activity. Overexpression of normal LRRK2 or a “kinase-dead” version of the enzyme did not produce these ill effects. What’s more, the mutant-LRRK2-mediated neurotoxicity was blocked by selective LRRK2 kinase inhibitors, suggesting that the latter could become a new treatment for PD.[10. B.D. Lee et al.,

“Inhibitors of leucine-rich repeat kinase-2 protect against models of Parkinson’s disease,” Nat Med, 16:998-1000, 2010. Free F1000 Evaluation]

Loss-of-function mutations

There is compelling evidence suggesting that loss-of-function mutations rendering the three genes, parkin, PINK1 and DJ-1, functionally inactive, underlie common forms of autosomal-recessive PD. Patients with loss-of-function parkin (also known as PARK2) mutations account for an estimated 40-50 percent of all familial early-onset cases of PD, whereas mutations in PINK1 (PARK6 ) and DJ-1 (PARK7 ) are less common.¹Several studies have demonstrated that products of all three recessive genes preserve mitochondrial functions, protect against reactive oxygen species, or play a role in protein degradation pathways. Normally, parkin tags proteins with ubiquitin, a protein that acts as an address label targeting tagged proteins either for destruction via the proteasome, or for other signaling pathways such as DNA repair, endocytosis, transcriptional regulation, and protein trafficking.¹ Mutations in the parkin gene lead to a loss of this tagging ability. Although a number of the proteins that parkin tags for degradation have been identified, no consensus has emerged on which of these may (if not tagged) accumulate in the cell, leading to neurodegeneration in PD.

Recent studies point towards both the parkin and PINK1 proteins playing a prominent role in preserving mitochondrial functions. Mutant flies that lack parkin or PINK1 genes exhibit abnormal mitochondria, as well as enhanced sensitivity to oxidative stress, muscle degeneration, and significant loss of dopamine-producing neurons. Mice lacking parkin and PINK1 also exhibit mitochondrial dysfunction that results in heightened oxidative stress.

Interestingly, researchers were able to rescue mitochondrial dysfunctions in flies lacking PINK1 by increasing parkin expression. On the other hand, increasing expression of PINK1 had no effect on dysfunctions in flies lacking parkin.¹ This neatly puts parkin and PINK1 in a common pathway, with PINK1 functioning upstream from parkin. Several recent studies also found parkin and PINK1 involvement in facilitating autophagic clearance of damaged mitochondria, by a process known as mitophagy.[11. R. Banerjee et al., “Autophagy in neurodegenerative disorders: pathogenic roles and therapeutic implications,” Trends Neurosci, 33:541-49, 2010.] In cellular models, parkin is selectively targeted to impaired mitochondria, a process that relies on the expression of PINK1 through phosphorylation of parkin.¹¹ These data suggest that a failure to activate efficient mitophagy may serve as an important pathogenic mechanism of cell death in PD.

The third recessive gene, DJ-1, produces a molecular chaperone that aids in protein folding, in addition to other functions. It is found in the cytosol, the mitochondrial matrix, and mitochondrial intermembrane space. In cellular models, it regulates redox-dependent signaling pathways and acts as a regulator of antioxidant gene expression, while gene deletion studies show that it counters oxidative stress in mitochondria. Recent studies indicate that DJ-1 deficiency is associated with perturbed mitochondrial dynamics and autophagic dysregulation,[12. I. Irrcher et al., “Loss of the Parkinson’s disease-linked gene DJ-1 perturbs mitochondrial dynamics,” Hum Mol Genet, 19:3734-46, 2010.]linking its functions with those mediated by parkin and PINK1. Despite all these advances, we still lack a clear picture of how exactly these three proteins (parkin, PINK1, and DJ-1) fit in a common pathway for disease development.

Future focus

The identification and characterization of familial PD-linked genes has sparked an extremely fruitful line of research, delineating molecular pathways that are involved in the pathogenesis of PD. Recent evidence suggests that these molecular pathways are not only relevant to the rare familial variants of PD, but also to the more common noninherited version of the disease. Genome-wide association studies are also contributing by providing a new tool for searching out other PD-associated genes. So far, the proteins that have been linked to PD by genetic studies have roles in lipid and vesicle dynamics (a-synuclein), the ubiquitin-proteasome system (parkin), abnormal kinase function (LRRK2), oxidative stress, and mitochondrial dysfunction (DJ-1, PINK1, parkin). It appears as though these disparate functions must converge, leading to the dysfunction and death of dopaminergic neurons.

While we have made great strides in understanding all these functions, the connections between them are not immediately apparent. Therefore, the major focus of future research should be to identify common underlying mechanisms by which familial PD-linked genes impact the survival of dopaminergic neurons—mechanisms which will offer new and tractable targets for developing drugs to prevent and treat PD.

F1000 Members Bobby Thomas and M. Flint Beal are at Weill Cornell Medical School in New York City.