Function Of LRRK2 Research Articles

CalDAG-GEFI acts as a guanine nucleotide exchange factor for LRRK2 to regulate LRRK2 function and neurodegeneration.

Mutations in LRRK2 are the most common genetic cause of Parkinson's disease (PD). LRRK2 protein contains two enzymatic domains: a GTPase (Roc-COR) and a kinase domain. Disease-causing mutations are found in both domains. Now, studies have focused largely on LRRK2 kinase activity, while attention to its GTPase function is limited. LRRK2 is a guanine nucleotide-binding protein, but the mechanism of direct regulation of its GTPase activity remains unclear and its physiological GEF is not known. Here, we identified CalDAG-GEFI (CDGI) as a physiological GEF for LRRK2. CDGI interacts with LRRK2 and increases its GDP to GTP exchange activity. CDGI modulates LRRK2 cellular functions and LRRK2-induced neurodegeneration in both LRRK2 Drosophila and mouse models. Together, this study identified the physiological GEF for LRRK2 and provides strong evidence that LRRK2 GTPase is regulated by GAPs and GEFs. The LRRK2 GTPase, GAP, or GEF activities have the potential to serve as therapeutic targets, which is distinct from the direct LRRK2 kinase inhibition.

Evaluation of Neuroinflammatory Contribution to Neurodegeneration in LRRK2 Drosophila Models.

Pathological mutations in the LRRK2 gene are the major genetic cause of Parkinson's disease (PD). Although several animal models with either LRRK2 down- or over-expression have been developed, the physiological function of LRRK2 remains elusive. LRRK2 is constitutively expressed in various tissues including neurons and glial cells, but importantly, it is expressed at low levels in dopaminergic neurons, further contributing to the cryptic function of LRRK2. Significant levels of LRRK2 protein and mRNA have been detected in peripheral blood mononuclear cells, lymph nodes, the spleen, and primary microglia, strongly suggesting the contribution of inflammatory cells to neuronal degeneration. In this research article, using Drosophila LRRK2 models, we were able to demonstrate a significant contribution of glial cells to the LRRK2 pathological phenotype. Furthermore, in Drosophila, neurodegeneration is associated with a significant and important increase in specific inflammatory peptides. Finally, levetiracetam, a compound widely used in human therapy to treat epilepsy, was able to rescue both neuronal degeneration and neuroinflammation.

G2019S selective LRRK2 kinase inhibitor abrogates mitochondrial DNA damage

Pathogenic mutations in LRRK2 cause Parkinson’s disease (PD). The G2019S variant is the most common, which results in abnormally high kinase activity. Compounds that target LRRK2 kinase activity are currently being developed and tested in clinical trials. We recently found that G2019S LRRK2 causes mitochondrial DNA (mtDNA) damage and treatment with multiple classes of LRRK2 kinase inhibitors at concentrations associated with dephosphorylation of LRRK2 reversed mtDNA damage to healthy control levels. Because maintaining the normal function of LRRK2 in heterozygous G2019S LRRK2 carriers while specifically targeting the G2019S LRRK2 activity could have an advantageous safety profile, we explored the efficacy of a G2019S mutant selective LRRK2 inhibitor to reverse mtDNA damage in G2019S LRRK2 models and patient cells relative to non-selective LRRK2 inhibitors. Potency of LRRK2 kinase inhibition by EB-42168, a G2019S mutant LRRK2 kinase inhibitor, and MLi-2, a non-selective inhibitor, was determined by measuring phosphorylation of LRRK2 at Ser935 and/or Ser1292 using quantitative western immunoblot analysis. The Mito DNADX assay, which allows for the accurate real-time quantification of mtDNA damage in a 96-well platform, was performed in parallel. We confirmed that EB-42168 selectively inhibits LRRK2 phosphorylation on G2019S LRRK2 relative to wild-type LRRK2. On the other hand, MLi-2 was equipotent for wild-type and G2019S LRRK2. Acute treatment with EB-42168 inhibited LRRK2 phosphorylation and also restored mtDNA damage to healthy control levels. We further investigated the relationship between LRRK2 kinase activity, mtDNA damage and mitophagy. Levels of mtDNA damage caused by G2019S LRRK2 were fully re-established within 2 h of a LRRK2 inhibitor wash out and recovery experiment, indicating the mtDNA damage phenotype is highly dynamic. G2019S LRRK2 mitophagy defects were not alleviated with LRRK2 kinase inhibition, suggesting that mitophagy is not mechanistically regulating LRRK2 kinase-mediated reversal of mtDNA damage in this acute timeframe. Abrogation of mtDNA damage with the mutant selective tool inhibitor EB-42168 demonstrates the potential of a precision medicine approach for LRRK2 G2019S PD. Levels of mtDNA damage may serve as a potential pharmacodynamic biomarker of altered kinase activity that could be useful for small molecule development and clinical trials.

LRRK2 and Parkinson's disease: from genetics to targeted therapy.

LRRK2 variants are implicated in both familial and sporadic PD. LRRK2-PD has a generally benign clinical presentation and variable pathology, with inconsistent presence of Lewy bodies and marked Alzheimer's disease pathology. The mechanisms underlying LRRK2-PD are still unclear, but inflammation, vesicle trafficking, lysosomal homeostasis, and ciliogenesis have been suggested, among others. As novel therapies targeting LRRK2 are under development, understanding the role and function of LRRK2 in PD is becoming increasingly important. Here, we outline the epidemiological, pathophysiological, and clinical features of LRRK2-PD, and discuss the arising therapeutic approaches targeting LRRK2 and possible future directions for research.

Regulation of LRRK2 dynamics and conformation states by small molecules.

Mutations in LRRK2 is a risk factor for Parkinson's disease (PD), and the aberrant kinase activity is the main cause for pathological functions of LRRK2. Research have been focused on developing selective inhibitors for the LRRK2 kinase. However, we now know that in addition to the intrinsic kinase activity, the conformation of the LRRK2 protein plays important roles in regulate LRRK2 function. We have shown that the kinase domain serves as a central hub for inter-domain communication, and is the major driver for LRRK2 conformation transitions. In this study, we used GaMD simulations coupled with HDX-MS to analyze the dynamic changes and the inter-domain allosteric communications of C-terminal half of the LRRK2, LRRK2RCKW. Specifically, when the kinase domain is locked into a closed, active conformation or an open, inactive conformation by binding of the type I (MLi-2) and type II (Rebastinib) inhibitors, respectively. While both type I and type II inhibitor reduced the kinase activity effectively, they drive the LRRK2 in different conformation. Binding of MLi-2 stabilizes the kinase domain, and reduces the dynamic of LRRK2RCKW globally, thus stabilizes the compact domain orientations of LRRK2RCKW. In contrast, binding of Rebastinib resulted in a more dynamic kinase domain, which promoted the extended conformation of LRRK2RCKW. We also reveal the importance of the Dk-helix, which plays crucial roles in bridging the kinase and GTPase domain. The dynamic changes in the kinase domain that propagate through the DK-helix would lead to different conformations of the ROC domain, which potentially affect the GTPase activity, which is crucial for the cellular function of LRRK2.

A New Target for Parkinson’s Disease

LRRK2 blockade has emerged as a potential target for developing disease-modifying therapies against Parkinson’s disease. Both in cellular and animal models, DNL201- a first-in-class central nervous system–penetrant LRRK2 kinase inhibitor, successfully mitigated the LRRK2 function resulting in re-establishing lysosomal activity as per the findings of Jennings and colleagues. In both healthy volunteers and patients with Parkinson’s Disease, LRRK2 kinase was inhibited by DNL201 and the drug candidate was also shown to be safe and tolerable at doses that improved lysosomal functioning. These findings argue for advanced studies on LRRK2 inhibitors including early as well as late-stage clinical investigations in patients with Parkinson's disease. Sci. Transl. Med.14, eabj2658 (2022).

Abrogation of LRRK2 dependent Rab10 phosphorylation with TLR4 activation and alterations in evoked cytokine release in immune cells

LRRK2 protein is expressed prominently in immune cells, cell types whose contribution to LRRK2-associated genetic Parkinson's disease (PD) is increasingly being recognised. We investigated the effect of inflammatory stimuli using RAW264.7 murine macrophage cells as model systems. A detailed time course of TLR2 and TLR4 stimulation was investigated through measuring LRRK2 phosphorylation at its specific phospho-sites, and Rab8 and Rab10 phosphorylation together with cytokine release following treatment with LPS and zymosan. LRRK2 phosphorylation at Ser935, Ser955 and Ser973 was increased significantly over untreated conditions at 4–24h in both WT-LRRK2 and T1348N-LRRK2 cell lines to similar extents although levels of Ser910 phosphorylation were maintained at higher levels throughout. Importantly we demonstrate that LPS stimulation significantly decreased phospho-Rab10 but not phospho-Rab8 levels over 4–24h in both WT-LRRK2 and T1348N-LRRK2 cell lines. The dephosphorylation of Rab10 was not attributed to its specific phosphatase, PPM1H as the levels remained unaltered with LPS treatment. MAPK phosphorylation occurred prior to LRRK2 phosphorylation which was validated by blocking TLR4 and TLR2 receptors with TAK242 or Sparstolonin B respectively. A significant decrease in basal level of TNFα release was noted in both T1348N-LRRK2 and KO-LRRK2 cell lines at 48h compared to WT-LRRK2 cell line, however LPS and zymosan treatment did not cause any significant alteration in the TNFα and IL-6 release between the three cell lines. In contrast, LPS and zymosan caused significantly lower IL-10 release in T1348N-LRRK2 and KO-LRRK2 cell lines. A significant decrease in phospho-Rab10 levels was also confirmed in human IPS-derived macrophages with TLR4 activation. Our data demonstrates for the first time that LRRK2-dependent Rab10 phosphorylation is modulated by LPS stimulation, and that cytokine release may be influenced by the status of LRRK2. These data provide further insights into the function of LRRK2 in immune response, and has relevance for understanding cellular dysfunctions when developing LRRK2-based inhibitors for clinical treatment.

Genetic and Environmental Factors Influence the Pleomorphy of LRRK2 Parkinsonism.

Missense mutations in the LRRK2 gene were first identified as a pathogenic cause of Parkinson’s disease (PD) in 2004. Soon thereafter, a founder mutation in LRRK2, p.G2019S (rs34637584), was described, and it is now estimated that there are approximately 100,000 people worldwide carrying this risk variant. While the clinical presentation of LRRK2 parkinsonism has been largely indistinguishable from sporadic PD, disease penetrance and age at onset can be quite variable. In addition, its neuropathological features span a wide range from nigrostriatal loss with Lewy body pathology, lack thereof, or atypical neuropathology, including a large proportion of cases with concomitant Alzheimer’s pathology, hailing LRRK2 parkinsonism as the “Rosetta stone” of parkinsonian disorders, which provides clues to an understanding of the different neuropathological trajectories. These differences may result from interactions between the LRRK2 mutant protein and other proteins or environmental factors that modify LRRK2 function and, thereby, influence pathobiology. This review explores how potential genetic and biochemical modifiers of LRRK2 function may contribute to the onset and clinical presentation of LRRK2 parkinsonism. We review which genetic modifiers of LRRK2 influence clinical symptoms, age at onset, and penetrance, what LRRK2 mutations are associated with pleomorphic LRRK2 neuropathology, and which environmental modifiers can augment LRRK2 mutant pathophysiology. Understanding how LRRK2 function is influenced and modulated by other interactors and environmental factors—either increasing toxicity or providing resilience—will inform targeted therapeutic development in the years to come. This will allow the development of disease-modifying therapies for PD- and LRRK2-related neurodegeneration.

The Functional Assessment of LRRK2 in Caenorhabditis elegans Mechanosensory Neurons.

The nematode Caenorhabditis elegans (C. elegans) is a powerful model organism to systematically analyze the functions of genes of interest in vivo. Especially, C. elegans nervous system is suitable for morphological and functional analyses of neuronal genes due to its optical transparency of the body and the well-established anatomy including neural connections. The C. elegans ortholog of Parkinson's disease-associated gene LRRK2, named lrk-1, has been shown to play a role in the regulation of axonal morphology in a subset of neurons. Here I describe the detailed methodologies for the assessment of LRK-1/LRRK2 function as well as the analysis of genetic interaction involving lrk-1/LRRK2 by performing live imaging of C. elegans mechanosenrory neurons.

LRRK2 mediates tubulation and vesicle sorting from lysosomes.

Genetic variation around the LRRK2 gene affects risk of both familial and sporadic Parkinson's disease (PD). However, the biological functions of LRRK2 remain incompletely understood. Here, we report that LRRK2 is recruited to lysosomes after exposure of cells to the lysosome membrane-rupturing agent LLOME. Using an unbiased proteomic screen, we identified the motor adaptor protein JIP4 as an LRRK2 partner at the lysosomal membrane. LRRK2 can recruit JIP4 to lysosomes in a kinase-dependent manner via the phosphorylation of RAB35 and RAB10. Using super-resolution live-cell imaging microscopy and FIB-SEM, we demonstrate that JIP4 promotes the formation of LAMP1-negative tubules that release membranous content from lysosomes. Thus, we describe a new process orchestrated by LRRK2, which we name LYTL (LYsosomal Tubulation/sorting driven by LRRK2), by which lysosomal tubulation is used to release vesicles from lysosomes. Given the central role of the lysosome in PD, LYTL is likely to be disease relevant.

Protein synthesis is suppressed in sporadic and familial Parkinson's disease by LRRK2.

Gain of function LRRK2-G2019S is the most frequent mutation found in familial and sporadic Parkinson's disease. It is expected therefore that understanding the cellular function of LRRK2 will provide insight on the pathological mechanism not only of inherited Parkinson's, but also of sporadic Parkinson's, the more common form. Here, we show that constitutive LRRK2 activity controls nascent protein synthesis in rodent neurons. Specifically, pharmacological inhibition of LRRK2, Lrrk2 knockdown or Lrrk2 knockout, all lead to increased translation. In the rotenone model for sporadic Parkinson's, LRRK2 activity increases, dopaminergic neuron translation decreases, and the neurites atrophy. All are prevented by LRRK2 inhibitors. Moreover, in striatum and substantia nigra of rotenone treated rats, phosphorylation changes are observed on eIF2α-S52(↑), eIF2s2-S2(↓), and eEF2-T57(↑) in directions that signify protein synthesis arrest. Significantly, translation is reduced by 40% in fibroblasts from Parkinson's patients (G2019S and sporadic cases alike) and this is reversed upon LRRK2 inhibitor treatment. In cells from multiple system atrophy patients, translation is unchanged suggesting that repression of translation is specific to Parkinson's disease. These findings indicate that repression of translation is a proximal function of LRRK2 in Parkinson's pathology.

A Critical LRRK at the Synapse? The Neurobiological Function and Pathophysiological Dysfunction of LRRK2.

Since the discovery of LRRK2 mutations causal to Parkinson’s disease (PD) in the early 2000s, the LRRK2 protein has been implicated in a plethora of cellular processes in which pathogenesis could occur, yet its physiological function remains elusive. The development of genetic models of LRRK2 PD has helped identify the etiological and pathophysiological underpinnings of the disease, and may identify early points of intervention. An important role for LRRK2 in synaptic function has emerged in recent years, which links LRRK2 to other genetic forms of PD, most notably those caused by mutations in the synaptic protein α-synuclein. This point of convergence may provide useful clues as to what drives dysfunction in the basal ganglia circuitry and eventual death of substantia nigra (SN) neurons. Here, we discuss the evolution and current state of the literature placing LRRK2 at the synapse, through the lens of knock-out, overexpression, and knock-in animal models. We hope that a deeper understanding of LRRK2 neurobiology, at the synapse and beyond, will aid the eventual development of neuroprotective interventions for PD, and the advancement of useful treatments in the interim.

The Current State-of-the Art of LRRK2-Based Biomarker Assay Development in Parkinson's Disease.

Evidence is mounting that LRRK2 function, particularly its kinase activity, is elevated in multiple forms of Parkinson’s disease, both idiopathic as well as familial forms linked to mutations in the LRRK2 gene. However, sensitive quantitative markers of LRRK2 activation in clinical samples remain at the early stages of development. There are several measures of LRRK2 activity that could potentially be used in longitudinal studies of disease progression, as inclusion/exclusion criteria for clinical trials, to predict response to therapy, or as markers of target engagement. Among these are levels of LRRK2, phosphorylation of LRRK2 itself, either by other kinases or via auto-phosphorylation, its in vitro kinase activity, or phosphorylation of downstream substrates. This is advantageous on many levels, in that multiple indices of elevated kinase activity clearly strengthen the rationale for targeting this kinase with novel therapeutic candidates, and provide alternate markers of activation in certain tissues or biofluids for which specific measures are not detectable. However, this can also complicate interpretation of findings from different studies using disparate measures. In this review we discuss the current state of LRRK2-focused biomarkers, the advantages and disadvantages of the current pallet of outcome measures, the gaps that need to be addressed, and the priorities that the field has defined.

The Michael J. Fox Foundation's Strategies for Accelerating Translation of LRRK2 into Therapies for Parkinson Disease.

Since 2005, The Michael J. Fox Foundation for Parkinson’s Research (MJFF) has invested significant funding and non-funding effort to accelerate research and drug development activity around the Parkinson disease (PD)-associated protein LRRK2. MJFF has spearheaded multiple public/private pre-competitive collaborations that have contributed to our understanding of LRRK2 function; de-risked potential safety questions around the therapeutic use of LRRK2 kinase inhibitors; and generated critical research tools, biosamples, and data for the field. Several LRRK2-targeted therapies are now in human testing due to the hard work of so many in the PD community. In this perspective, we present a holistic description and model of how our Foundation’s support targeted important barriers to LRRK2 research and helped move the field into clinical trials.

Lrrk2 modulation of Wnt signaling during zebrafish development.

Mutations in leucine-rich repeat kinase 2 (lrrk2) are the most common genetic cause of Parkinson's disease. Difficulty in elucidating the pathogenic mechanisms resulting from disease-associated Lrrk2 variants stems from the complexity of Lrrk2 function and activities. Lrrk2 contains multiple protein-protein interacting domains, a GTPase domain, and a kinase domain. Lrrk2 is implicated in many cellular processes including vesicular trafficking, autophagy, cytoskeleton dynamics, and Wnt signaling. Here, we generated a zebrafish lrrk2 allelic series to study the requirements for Lrrk2 during development and to dissect the importance of its various domains. The alleles are predicted to encode proteins that either lack all functional domains (lrrk2sbu304 ), the GTPase, and kinase domains (lrrk2sbu71 ) or the kinase domain (lrrk2sbu96 ). All three lrrk2 mutants are viable, morphologically normal, and display wild-type-like locomotion. Because Lrrk2 modulates Wnt signaling in some contexts, we assessed Wnt signaling in all three mutant lines. Analysis of Wnt signaling by studying the expression of target genes using whole mount RNA in situ hybridization and a transgenic Wnt reporter revealed wild-type domains of Wnt activity in each of the mutants. However, we found that Wnt pathway activation is attenuated in lrrk2sbu304/sbu304 , which lacks both scaffolding and catalytic domains, but not in the other alleles during late embryogenesis. This supports a model in which Lrrk2 scaffolding functions are key to a context-dependent role in promoting canonical Wnt signaling.

LRRK2 Is Recruited to Phagosomes and Co-recruits RAB8 and RAB10 in Human Pluripotent Stem Cell-Derived Macrophages.

SummaryThe Parkinson's disease-associated gene, LRRK2, is also associated with immune disorders and infectious disease and is expressed in immune subsets. Here, we characterize a platform for interrogating the expression and function of endogenous LRRK2 in authentic human phagocytes using human induced pluripotent stem cell-derived macrophages and microglia. Endogenous LRRK2 is expressed and upregulated by interferon-γ in these cells, including a 187-kDa cleavage product. Using LRRK2 knockout and G2019S isogenic repair lines, we find that LRRK2 is not involved in initial phagocytic uptake of bioparticles but is recruited to LAMP1+/RAB9+ “maturing” phagosomes, and LRRK2 kinase inhibition enhances its residency at the phagosome. Importantly, LRRK2 is required for RAB8a and RAB10 recruitment to phagosomes, implying that LRRK2 operates at the intersection between phagosome maturation and recycling pathways in these professional phagocytes.

The LRRK2 N-terminal domain influences vesicle trafficking: impact of the E193K variant

The LRRK2 protein consists of multiple functional domains, including protein-binding domains at its N and C-terminus. Mutations in the Leucine-rich repeat kinase 2 gene (LRRK2) have been linked to familial and sporadic Parkinson’s disease (PD). We have recently described a novel variant falling within the N-terminal armadillo repeats, E193K. Herein, our aim is to investigate the functional impact of LRRK2 N-terminal domain and the E193K variant on vesicle trafficking. By combining Total Internal Reflection Fluorescence (TIRF) microscopy and a synaptopHluorin assay, we found that expression of a construct lacking the N-terminal domain increases the frequency and amplitude of spontaneous synaptic events. Complementary biochemical approaches showed that the E193K variant alters the binding properties of LRRK2, decreases LRRK2 binding to synaptic vesicles, and promotes vesicle fusion. Our results confirm the physiological and pathological relevance of the nature of the LRRK2-associated macro-molecular complex solidifying the idea that different pathological mutations critically alter the scaffolding function of LRRK2 resulting in a perturbation of the vesicular trafficking as a common denominator.

Loss of LrrK2 Function Potentiates Dopamine Signaling in the Striatum to Promote Compulsive Alcohol Use

Long-lasting plastic changes in the striatum play a central role in driving the transition from hedonic to compulsive alcohol drinking, a hallmark of alcohol use disorders. It is known that the propensity to undergo this transition varies with the individual, yet the underlying molecular mechanisms are not well understood. Here, we report that the Parkinson’s-related protein Leucine-rich repeat kinase 2 (LRRK2) plays a role in in this transition to compulsive alcohol drinking by modulating the function of dopamine D1 receptors within the striatonigral pathway. We find that alcohol decreases LRRK2 kinase activity in the dorsal striatum and that deletion of the Lrrk2 gene specifically from D1-expressing striatal neurons promotes compulsive-like alcohol drinking in mice by strengthening D1 receptor signaling and function. These findings identify a novel role for Lrrk2 in the striatum and suggest that Lrrk2 function promotes resilience for compulsive alcohol drinking and the development of alcohol use disorders.

Identification of PP2A and S6 Kinase as Modifiers of Leucine-Rich Repeat Kinase-Induced Neurotoxicity

Mutations in LRRK2 are currently recognized as the most common monogenetic cause of Parkinsonism. The elevation of kinase activity of LRRK2 that frequently accompanies its mutations is widely thought to contribute to its toxicity. Accordingly, many groups have developed LRRK2-specific kinase inhibitors as a potential therapeutic strategy. Given that protein phosphorylation is a reversible event, we sought to elucidate the phosphatase(s) that can reverse LRRK2-mediated phosphorylation, with the view that targeting this phosphatase(s) may similarly be beneficial. Using an unbiased RNAi phosphatase screen conducted in a Drosophila LRRK2 model, we identified PP2A as a genetic modulator of LRRK2-induced neurotoxicity. Further, we also identified ribosomal S6 kinase (S6K), a target of PP2A, as a novel regulator of LRRK2 function. Finally, we showed that modulation of PP2A or S6K activities ameliorates LRRK2-associated disease phenotype in Drosophila.

LRRK2 binds to the Rab32 subfamily in a GTP-dependent manner via its armadillo domain

ABSTRACT LRRK2 is a multi-domain Ser/Thr kinase that is associated with inherited and sporadic cases of Parkinson’s disease. Many mutations linked to disease are associated within a central ROC-COR regulatory region and the subsequent kinase domain, leading to enhanced catalytic activity. The N-terminus of human LRRK2 consists of armadillo repeat motifs (ARMs) followed by ankyrin repeats (ANKs). Recently, Rab GTPases have emerged as key players in LRRK2 function, both as substrates of the kinase, and as regulators of the catalytic activity. Rabs recruit effector proteins via their GTP-dependent switch 1 and 2 regions to distinct sub-cellular compartments to regulate membrane trafficking. LRRK2 phosphorylates Rab8, Rab10 and Rab12 in switch 2, and this activity is regulated via interactions with Rab29. Furthermore, the related Rab32-subfamily GTPases, Rab32 and Rab38, have also been shown to interact with LRRK2. Here, we have mapped the interactions of the Rab32-subfamily to the ARM domain of LRRK2. The complexes are dependent on the GTP state of the Rabs in vitro, implying that LRRK2 may be an effector of the Rab32-subfamily of small GTPases. X-ray crystal structures of the Rab32-family GTPases and subsequent mutational studies reveal that a positively charged residue in switch 1 is critical for binding of Rab32/38 to LRRK2. Homology modelling and mutational analyses of the ARM domain point to a patch of negatively charged residues that contribute to complex formation. These structural and biochemical studies provide a framework for understanding the molecular basis for Rab regulation of LRRK2 and its role in Parkinson’s disease.