Function Of LRRK2 Research Articles

The role of LRRK2 in cell signalling.

Parkinson's disease (PD) is a common late-onset neurodegenerative disorder known primarily for its motor features. Mutations and risk variants in LRRK2 cause familial and idiopathic forms of PD. Mutations segregating with disease are found in the LRRK2 GTPase and kinase domains, affecting catalytic activity and protein-protein interactions. This likely results in an overall gain of LRRK2 cell signalling function contributing to PD pathogenesis. This concept supports the development of LRRK2 kinase inhibitors as disease-modifying treatments, at least for a subset of patients. However, the function of LRRK2 as a cell signalling protein with two catalytic and several protein-protein interaction domains is highly complex. For example, LRRK2 plays important roles in several inflammatory diseases, raising the possibility that it may mediate immune responses in PD. Consistently, LRRK2-mediated cell signalling was not only shown to be important for neuronal function, including neuronal development and homeostasis, but also for peripheral and central immune responses. The catalytic activity of LRRK2 is regulated by autophosphorylation, protein monomer/dimer cycling, and upstream kinases and GTPases, affecting its subcellular localisation and downstream signalling. Part of LRRK2-mediated signalling is likely facilitated by Rab protein phosphorylation, affecting primarily membrane trafficking, including vesicle release at the trans-Golgi network. However, LRRK2 also displays intrinsic GTPase activity and functions as a signalling scaffold. As an example, LRRK2 was suggested to be part of the NRON complex and β-catenin destruction complex, inhibiting NFAT and canonical Wnt signalling, respectively. In summary, continuous research into LRRK2 signalling function contributes to novel diagnostic and therapeutic concepts in PD.

Understanding the role of genetic variability in LRRK2 in Indian population.

Genetic variability in LRRK2 has been unequivocally established as a major risk factor for familial and sporadic forms of PD in ethnically diverse populations. To resolve the role of LRRK2 in the Indian population. We performed targeted resequencing of the LRRK2 locus in 288 cases and 298 controls and resolved the haplotypic structure of LRRK2 in a combined cohort of 800 cases and 402 controls in the Indian population. We assessed the frequency of novel missense variants in the white and East Asian population by leveraging exome sequencing and densely genotype data, respectively. We did computational modeling and biochemical approach to infer the potential role of novel variants impacting the LRRK2 protein function. Finally, we assessed the phosphorylation activity of identified novel coding variants in the LRRK2 gene. We identified four novel missense variants with frequency ranging from 0.0008% to 0.002% specific for the Indian population, encompassing armadillo and kinase domains of the LRRK2 protein. A common genetic variability within LRRK2 may contribute to increased risk, but it was nonsignificant after correcting for multiple testing, because of small cohort size. The computational modeling showed destabilizing effect on the LRRK2 function. In comparison to the wild-type, the kinase domain variant showed 4-fold increase in the kinase activity. Our study, for the first time, identified novel missense variants for LRRK2, specific for the Indian population, and showed that a novel missense variant in the kinase domain modifies kinase activity in vitro. © 2018 International Parkinson and Movement Disorder Society.

Parkinson disease-associated mutations in LRRK2 cause centrosomal defects via Rab8a phosphorylation

BackgroundMutations in LRRK2 are a common genetic cause of Parkinson’s disease (PD). LRRK2 interacts with and phosphorylates a subset of Rab proteins including Rab8a, a protein which has been implicated in various centrosome-related events. However, the cellular consequences of such phosphorylation remain elusive.MethodsHuman neuroblastoma SH-SY5Y cells stably expressing wildtype or pathogenic LRRK2 were used to test for polarity defects in the context of centrosomal positioning. Centrosomal cohesion deficits were analyzed from transiently transfected HEK293T cells, as well as from two distinct peripheral cell types derived from LRRK2-PD patients. Kinase assays, coimmunoprecipitation and GTP binding/retention assays were used to address Rab8a phosphorylation by LRRK2 and its effects in vitro. Transient transfections and siRNA experiments were performed to probe for the implication of Rab8a and its phosphorylated form in the centrosomal deficits caused by pathogenic LRRK2.ResultsHere, we show that pathogenic LRRK2 causes deficits in centrosomal positioning with effects on neurite outgrowth, cell polarization and directed migration. Pathogenic LRRK2 also causes deficits in centrosome cohesion which can be detected in peripheral cells derived from LRRK2-PD patients as compared to healthy controls, and which are reversed upon LRRK2 kinase inhibition. The centrosomal cohesion and polarity deficits can be mimicked when co-expressing wildtype LRRK2 with wildtype but not phospho-deficient Rab8a. The centrosomal defects induced by pathogenic LRRK2 are associated with a kinase activity-dependent increase in the centrosomal localization of phosphorylated Rab8a, and are prominently reduced upon RNAi of Rab8a.ConclusionsOur findings reveal a new function of LRRK2 mediated by Rab8a phosphorylation and related to various centrosomal defects.

The Drosophila hep pathway mediates Lrrk2-induced neurodegeneration.

Although the pathogenesis of Parkinson's disease (PD) remains unclear, mutations in leucine-rich repeat kinase 2 (Lrrk2) are among the major causes of familial PD. Most of these mutations disrupt Lrrk2 kinase and (or) GTPase domain function, resulting in neuronal degeneration. However, the signal pathways underlying Lrrk2-induced neuronal degeneration are not fully understood. There is an expanding body of evidence that suggests a link between Lrrk2 function and MAP kinase (MAPK) cascades. To further investigate this link in vivo, genetic RNAi screens of the MAPK pathways were performed in a Drosophila model to identify genetic modifier(s) that can suppress G2019S-Lrrk2-induced PD-like phenotypes. The results revealed that the knockdown of hemipterous (hep, or JNKK) increased fly survival time, improved locomotor function, and reduced loss of dopaminergic neurons in G2019S-Lrrk2 transgenic flies. Expression of the dominant-negative allele of JNK (JNK-DN), a kinase that is downstream of hep in G2019S-Lrrk2 transgenic flies, elicited a similar effect. Moreover, treatment with the JNK inhibitor SP600125 partially reversed the G2019S-Lrrk2-induced loss of dopaminergic neurons. These results indicate that the hep pathway plays an important role in Lrrk2-linked Parkinsonism in flies. These studies provide new insights into the molecular mechanisms underlying Lrrk2-linked PD pathogenesis and aid in identifying potential therapeutic targets.

Familial knockin mutation of LRRK2 causes lysosomal dysfunction and accumulation of endogenous insoluble α-synuclein in neurons

Missense mutations in the multi-domain kinase LRRK2 cause late onset familial Parkinson's disease. They most commonly with classic proteinopathy in the form of Lewy bodies and Lewy neurites comprised of insoluble α-synuclein, but in rare cases can also manifest tauopathy. The normal function of LRRK2 has remained elusive, as have the cellular consequences of its mutation. Data from LRRK2 null model organisms and LRRK2-inhibitor treated animals support a physiological role for LRRK2 in regulating lysosome function. Since idiopathic and LRRK2-linked PD are associated with the intraneuronal accumulation of protein aggregates, a series of critical questions emerge. First, how do pathogenic mutations that increase LRRK2 kinase activity affect lysosome biology in neurons? Second, are mutation-induced changes in lysosome function sufficient to alter the metabolism of α-synuclein? Lastly, are changes caused by pathogenic mutation sensitive to reversal with LRRK2 kinase inhibitors? Here, we report that mutation of LRRK2 induces modest but significant changes in lysosomal morphology and acidification, and decreased basal autophagic flux when compared to WT neurons. These changes were associated with an accumulation of detergent-insoluble α-synuclein and increased neuronal release of α-synuclein and were reversed by pharmacologic inhibition of LRRK2 kinase activity. These data demonstrate a critical and disease-relevant influence of native neuronal LRRK2 kinase activity on lysosome function and α-synuclein homeostasis. Furthermore, they also suggest that lysosome dysfunction, altered neuronal α-synuclein metabolism, and the insidious accumulation of aggregated protein over decades may contribute to pathogenesis in this late-onset form of familial PD.

LRRK2 promotes the activation of NLRC4 inflammasome during Salmonella Typhimurium infection.

Although genetic polymorphisms in the LRRK2 gene are associated with a variety of diseases, the physiological function of LRRK2 remains poorly understood. In this study, we report a crucial role for LRRK2 in the activation of the NLRC4 inflammasome during host defense against Salmonella enteric serovar Typhimurium infection. LRRK2 deficiency reduced caspase-1 activation and IL-1β secretion in response to NLRC4 inflammasome activators in macrophages. Lrrk2-/- mice exhibited impaired clearance of pathogens after acute S. Typhimurium infection. Mechanistically, LRRK2 formed a complex with NLRC4 in the macrophages, and the formation of the LRRK2-NLRC4 complex led to the phosphorylation of NLRC4 at Ser533. Importantly, the kinase activity of LRRK2 is required for optimal NLRC4 inflammasome activation. Collectively, our study reveals an important role for LRRK2 in the host defense by promoting NLRC4 inflammasome activation.

68 and FX2149 Attenuate Mutant LRRK2-R1441C-Induced Neural Transport Impairment.

Leucine-rich repeat kinase 2 is a large protein with implications in genetic and sporadic causes of Parkinson's disease. The physiological functions of LRRK2 are largely unknown. In this report, we investigated whether LRRK2 alters neural transport using live-cell imaging techniques and human neuroblastoma SH-SY5Y cells. Our results demonstrated that expression of the PD-linked mutant, LRRK2-R1441C, induced mitochondrial, and lysosomal transport defects in neurites of SH-SY5Y cells. Most importantly, recently identified GTP-binding inhibitors, 68 and FX2149, can reduce LRRK2 GTP-binding activity and attenuates R1441C-induced mitochondrial and lysosomal transport impairments. These results provide direct evidence and an early mechanism for neurite injury underlying LRRK2-induced neurodegeneration. This is the first report to show that LRRK2 GTP-binding activity plays a critical role during neurite transport, suggesting inhibition of LRRK2 GTP-binding could be a potential novel strategy for PD intervention.

Mechanisms of Mutant LRRK2 Neurodegeneration.

LRRK2 mutations are associated with the loss of neurons, that is to say toxicity, in patients and in experimental model systems. However, the mechanisms by which mutations can be linked to neurodegeneration are not fully defined. Here I will argue that mechanism in this context encompasses a variety of levels of information. Mutations or alterations in gene expression at a genetic level are one set of mechanisms that are reflected at the biochemical level likely in enhanced or persistent function of LRRK2. By impacting cellular pathways, prominently including changes in autophagy but also microtubule function, mitochondria and protein synthesis, in neurons and immune cells, the LRRK2 brain is primed for neurodegeneration in an age-dependent manner. These concepts have implications for not only modeling LRRK2 disease but also perhaps for Parkinson's disease more generally, including the development of therapeutic modalities.

LRRK2 at the interface of autophagosomes, endosomes and lysosomes.

Over the past 20 years, substantial progress has been made in identifying the underlying genetics of Parkinson’s disease (PD). Of the known genes, LRRK2 is a major genetic contributor to PD. However, the exact function of LRRK2 remains to be elucidated. In this review, we discuss how familial forms of PD have led us to hypothesize that alterations in endomembrane trafficking play a role in the pathobiology of PD. We will discuss the major observations that have been made to elucidate the role of LRRK2 in particular, including LRRK2 animal models and high-throughput proteomics approaches. Taken together, these studies strongly support a role of LRRK2 in vesicular dynamics. We also propose that targeting these pathways may not only be beneficial for developing therapeutics for LRRK2-driven PD, but also for other familial and sporadic cases.

Activation of FADD-Dependent Neuronal Death Pathways as a Predictor of Pathogenicity for LRRK2 Mutations.

BackgroundDespite the plethora of sequence variants in LRRK2, only a few clearly segregate with PD. Even within this group of pathogenic mutations, the phenotypic profile can differ widely.ObjectiveWe examined multiple properties of LRRK2 behavior in cellular models over-expressing three sequence variants described in Greek PD patients in comparison to several known pathogenic and non-pathogenic LRRK2 mutations, to determine if specific phenotypes associated with pathogenic LRRK2 can be observed in other less-common sequence variants for which pathogenicity is unclear based on clinical and/or genetic data alone.MethodsThe oligomerization, activity, phosphorylation, and interaction with FADD was assessed in HEK293T cells over-expressing LRRK2; while the induction of neuronal death was determined by quantifying apoptotic nuclei in primary neurons transiently expressing LRRK2.ResultsOne LRRK2 variant, A211V, exhibited a modest increase in kinase activity, whereas only the pathogenic mutants G2019S and I2020T displayed significantly altered auto-phosphorylation. We observed an induction of detergent-insoluble high molecular weight structures upon expression of pathogenic LRRK2 mutants, but not the other LRRK2 variants. In contrast, each of the variants tested induced apoptotic death of cultured neurons similar to pathogenic LRRK2 in a FADD-dependent manner.ConclusionsOverall, despite differences in some properties of LRRK2 function such as kinase activity and its oligomerization, each of the LRRK2 variants examined induced neuronal death to a similar extent. Furthermore, our findings further strengthen the notion of a convergence on the extrinsic cell death pathway common to mutations in LRRK2 that are capable of inducing neuronal death.

The ubiquitin signal and autophagy: an orchestrated dance leading to mitochondrial degradation

The quality of mitochondria, essential organelles that produce ATP and regulate numerous metabolic pathways, must be strictly monitored to maintain cell homeostasis. The loss of mitochondrial quality control systems is acknowledged as a determinant for many types of neurodegenerative diseases including Parkinson's disease (PD). The two gene products mutated in the autosomal recessive forms of familial early-onset PD, Parkin and PINK1, have been identified as essential proteins in the clearance of damaged mitochondria via an autophagic pathway termed mitophagy. Recently, significant progress has been made in understanding how the mitochondrial serine/threonine kinase PINK1 and the E3 ligase Parkin work together through a novel stepwise cascade to identify and eliminate damaged mitochondria, a process that relies on the orchestrated crosstalk between ubiquitin/phosphorylation signaling and autophagy. In this review, we highlight our current understanding of the detailed molecular mechanisms governing Parkin-/PINK1-mediated mitophagy and the evidences connecting Parkin/PINK1 function and mitochondrial clearance in neurons.

Leucine-rich repeat kinase 2 is a regulator of B cell function, affecting homeostasis, BCR signaling, IgA production, and TI antigen responses

LRRK2 is the causal molecule of autosomal dominant familial Parkinson's disease. B2 cells express a much higher LRRK2 mRNA level than B1 cells. To reveal the function of LRRK2 in B cells, we analyzed B cell functions in LRRK2-knockout (LRRK2−/−) mice. LRRK2−/− mice had significantly higher counts of peritoneal B1 cells than wild-type mice. After BCR stimulation, phosphor-Erk1/2 of splenic B2 cells was enhanced to a higher degree in LRRK2−/− mice. LRRK2−/− mice had a significantly higher serum IgA level, and TNP–Ficoll immunization increased the titer of serum anti-TNP IgM antibody. LRRK2 may play important roles in B cells.

Chronic and acute LRRK2 silencing has no long-term behavioral effects, whereas wild-type and mutant LRRK2 overexpression induce motor and cognitive deficits and altered regulation of dopamine release.

Germline silencing of the PD-related protein LRRK2 does not alter glutamate or dopamine release in adult mice, but some exploratory abnormalities have been reported with ageing. Contrastingly, high levels of human LRRK2 cause locomotor alterations and cognitive deficits accompanied by reduced striatal dopamine levels, with the latter also observed in G2019S mutant mice. Comparative cognitive and motor behavioral testing of LRRK2 KO, overexpressor and mutant overexpressor mice has not previously been reported. Parallel, comparative behavioral characterization was performed assessing motor and cognitive abilities. Striatal antisense oligonucleotide injections were conducted to investigate the effects of acute LRRK2 silencing on behavior and dopamine fiber density. Striatal synaptosomes prepared from hG2019S mice assessed vesicular release of dopamine and its sensitivity to D2 autoreceptor stimulation. Genetic ablation of LRRK2 has no long-term consequences on motor or cognitive function. Consistently, no effects on behavior or dopaminergic fiber density were observed following acute striatal silencing. Conversely, 12-month OE mice show persistent locomotor deficits and worsening of cognitive abilities; whereas, hG2019S mice display early hyperactivity and effective learning and memory that progress to decreased motor and cognitive deficits at older ages. The G2019S mutation does not affect vesicular dopamine release, but decreases its sensitivity to D2-mediated inhibition. LRRK2 silencing is well tolerated in mouse, arguing PD does not result from LRRK2 loss of function. High levels of WT and G2019S LRRK2 produce similar but temporally distinct phenotypes, potentially modeling different stages of disease progression. The data implicate gain of LRRK2 function in the pathogenesis of PD.

Neural stem cell culture model for LRRK2 function determines regulation of dopaminergic and microtubule-associated genes

Leucine-rich repeat kinase 2 (Lrrk2) is a Parkinson’s disease (PD)-related protein involved in sporadic and familial cases of the disease. Lrrk2 is a kinase and may be involved in intracellular signaling pathways. We established a neural stem cell (NSC)-based model system for Lrrk2-associated PD. We investigated its physiological and pathophysiological expression in human and mouse brain and NSCs using different antibodies defining dopaminergic NSCs as a relevant cells population to study Lrrk2 dysfunction. Using small interfering RNA duplexes targeting Lrrk2 in fetal mouse NSCs, we observed higher Tyrosine hydroxylase (Th) mRNA levels. Conversely, when overexpressing human LRRK2 in fetal mouse NSCs, Th, mRNA levels were reduced. Interestingly, in mutant LRRK2 overexpression Map2 and Kif3A mRNA levels were reduced. In conclusion, we demonstrate that overexpressing LRRK2 in neural stem cells results in a cell culture model for LRRK2-mediated PD. Lrrk2 is conversely linked to Th expression and expression of microtubule genes, implicating a role in the pathomechanism towards dopaminergic neurodegeneration.

LRRK2 Pathways Leading to Neurodegeneration.

Mutations in LRRK2 are associated with inherited Parkinson's disease (PD) in a large number of families, and the genetic locus containing the LRRK2 gene contains a risk factor for sporadic PD. The LRRK2 protein contains several domains that suggest a role in cellular signaling, including a kinase domain. It is also clear that LRRK2 interacts, either physically or genetically, with several other important proteins implicated in PD, suggesting that LRRK2 may be a central player in the pathways that underlie parkinsonism. As such, LRRK2 has been proposed to be a plausible target for therapeutic intervention, with kinase inhibition being pursued most actively. However, there are still several fundamental aspects of LRRK2 biology and function that remain unresolved at this time. This review will focus on the key questions of normal function of LRRK2 and how this might be related to the pathophysiology of PD.

LRRK2 a pivotal player in mitochondrial dynamics and lysosomal clustering: highlights to sporadic Parkinson’s disease

Parkinson’s disease (PD) is the second most common progressive neurodegenerative disorder affecting more than 1% of the population over 65 years of age. Dominant mutations in the leucine-rich repeat kinase 2 (lrrk2) gene are found associated with both familial and sporadic cases and represent the most frequent genetic lesions associated with PD. Remarkably, lrrk2 mutations cause PD with age-related penetrance and clinical features similar to late-onset sporadic PD (sPD). LRRK2 is a complex and large protein constituted by multiple domains executing several functions, including GTP hydrolysis, kinase activity and protein binding. Although the cellular function of LRRK2 is largely unknown there is increasing evidence that LRRK2 posits a role in autophagic regulation, microtubule (MT) dynamics, and mitochondrial function. In our recent report, we tackled LRRK2 physiological role taking advantage of a potent and selective inhibitor (LRRK2-IN-1) of LRRK2 kinase activity. We evaluated LRRK2-IN-1 effects on several mechanisms known to be impaired in PD, such as MT network and trafficking, mitochondrial dynamics and function and the autophagic-lysosomal pathway. Importantly, we found that LRRK2 is necessary to maintain the levels of acetylated tubulin, which is a post-translational modification essential for MTs structure and function and with a prevalent role for axonal intracellular trafficking. In addition, we also provided evidence that LRRK2 kinase activity regulates mitochondrial dynamics through phosphorylation of mitochondrial dynamin-like protein (Drp1) increasing mitochondrial fission. Adding up, LRRK2 kinase activity is essential for regulation of lysosomal clustering and distribution through Rab7. Therefore, our recent findings reveal novel insights into the function of LRRK2 in a normal or diseased context.

Computational analysis of the LRRK2 interactome.

LRRK2 was identified in 2004 as the causative protein product of the Parkinson’s disease locus designated PARK8. In the decade since then, genetic studies have revealed at least 6 dominant mutations in LRRK2 linked to Parkinson’s disease, alongside one associated with cancer. It is now well established that coding changes in LRRK2 are one of the most common causes of Parkinson’s. Genome-wide association studies (GWAs) have, more recently, reported single nucleotide polymorphisms (SNPs) around the LRRK2 locus to be associated with risk of developing sporadic Parkinson’s disease and inflammatory bowel disorder. The functional research that has followed these genetic breakthroughs has generated an extensive literature regarding LRRK2 pathophysiology; however, there is still no consensus as to the biological function of LRRK2. To provide insight into the aspects of cell biology that are consistently related to LRRK2 activity, we analysed the plethora of candidate LRRK2 interactors available through the BioGRID and IntAct data repositories. We then performed GO terms enrichment for the LRRK2 interactome. We found that, in two different enrichment portals, the LRRK2 interactome was associated with terms referring to transport, cellular organization, vesicles and the cytoskeleton. We also verified that 21 of the LRRK2 interactors are genetically linked to risk for Parkinson’s disease or inflammatory bowel disorder. The implications of these findings are discussed, with particular regard to potential novel areas of investigation.

Commentary on "Synaptic function is modulated by LRRK2 and glutamate release is increased in cortical neurons of G2019S LRRK2 knock-in mice".

Parkinson's disease (PD) is the most common form of age related motor disorder (Hsu et al., 2010; Ferree et al., 2012). Mutations in leucine-rich repeat kinase 2, LRRK2, gene are considered to be genetic determinants of PD. Among them G2019S is the most prevalent amino acid substitution mutation in LRRK2 and accounts for 1–2% of sporadic PD cases (Healy et al., 2008). LRRK2 is involved in many signaling pathways and its role is different in different cell types. In case of PD the main interest lies in understanding the function of LRRK2 in neuronal physiology. The study of Beccano-Kelly et al. attempts to investigate the role of LRRK2 in synaptic physiology in the context of loss of function and gain of function. Authors compared and contrasted obtained results on normal LRRK2 functions to effects of G2019S mutant LRRK2. To perform experiments they used primary cortical cultures prepared from LRRK2 transgenic overexpressing (OE), knock-out (KO), and knock-in G2019S (KI) mice. Beccano-Kelly et al. report that LRRK2 modulates synaptic function via regulation of glutamatergic activity. This result is a continuation of earlier findings obtained by other research groups in Drosophila model (Lee et al., 2010; Matta et al., 2012) and in mammalian cortical cultures (Piccoli et al., 2011; Parisiadou et al., 2014). Mentioned studies are contradictory in relation to whether deletion of LRRK2 upregulates, or downregulates, glutamatergic synaptic transmission. Beccano-Kelly et al. found out that LRRK2 knock out leads to reduced glutamatergic activity. The current study and previous reports from other groups do not uncover the mechanism of LRRK2 mediated synaptic function. This point should be investigated in the future studies. The most interesting result of Beccano-Kelly et al. was that in the absence of any change to synapse density glutamate release was markedly elevated in knock-in cultures. This indicated that physiological levels of G2019S LRRK2 elevate probability of release. Next observation was that the phosphorylation of synapsin 1 was significantly reduced in KI neurons. Based on obtained results authors concluded that perturbations to the presynaptic release machinery and elevated synaptic transmission are early neuronal effects of LRRK2 G2019S. Taken together, the study of Beccano-Kelly et al. presents novel data about the role of normal and G2019S mutated LRRK2 in regulation of synaptic transmission. On the one hand, discovering the signaling mechanism underlying LRRK2-mediated regulation of synaptic function could lead to the development of PD-preventing therapies.

Interaction of LRRK2 with kinase and GTPase signaling cascades.

LRRK2 is a protein that interacts with a plethora of signaling molecules, but the complexity of LRRK2 function presents a challenge for understanding the role of LRRK2 in the pathophysiology of Parkinson’s disease (PD). Studies of LRRK2 using over-expression in transgenic mice have been disappointing, however, studies using invertebrate systems have yielded a much clearer picture, with clear effects of LRRK2 expression, knockdown or deletion in Caenorhabditis elegans and Drosophila on modulation of survival of dopaminergic neurons. Recent studies have begun to focus attention on particular signaling cascades that are a target of LRRK2 function. LRRK2 interacts with members of the mitogen activated protein kinase (MAPK) pathway and might regulate the pathway action by acting as a scaffold that directs the location of MAPK pathway activity, without strongly affecting the amount of MAPK pathway activity. Binding to GTPases, GTPase-activating proteins and GTPase exchange factors are another strong theme in LRRK2 biology, with LRRK2 binding to rac1, cdc42, rab5, rab7L1, endoA, RGS2, ArfGAP1, and ArhGEF7. All of these molecules appear to feed into a function output for LRRK2 that modulates cytoskeletal outgrowth and vesicular dynamics, including autophagy. These functions likely impact modulation of α-synuclein aggregation and associated toxicity eliciting the disease processes that we term PD.

Leucine-Rich Repeat Kinase 2 Binds to Neuronal Vesicles through Protein Interactions Mediated by Its C-Terminal WD40 Domain

Mutations in the leucine-rich repeat kinase 2 gene (LRRK2) are associated with familial and sporadic Parkinson's disease (PD). LRRK2 is a complex protein that consists of multiple domains, including predicted C-terminal WD40 repeats. In this study, we analyzed functional and molecular features conferred by the WD40 domain. Electron microscopic analysis of the purified LRRK2 C-terminal domain revealed doughnut-shaped particles, providing experimental evidence for its WD40 fold. We demonstrate that LRRK2 WD40 binds and sequesters synaptic vesicles via interaction with vesicle-associated proteins. In fact, a domain-based pulldown approach combined with mass spectrometric analysis identified LRRK2 as being part of a highly specific protein network involved in synaptic vesicle trafficking. In addition, we found that a C-terminal sequence variant associated with an increased risk of developing PD, G2385R, correlates with a reduced binding affinity of LRRK2 WD40 to synaptic vesicles. Our data demonstrate a critical role of the WD40 domain within LRRK2 function.