Leucine-rich Repeat Kinase 2 Parkinson's Disease Research Articles

LRRK2 G2019S and Parkinson's disease: insight from Neuroinflammation.

The multiple hypothesis holds that the pathogenesis of Parkinson's disease (PD) requires many factors such as heredity, environment and ageing. Mutations in Leucine-rich repeat kinase 2 (LRRK2) are recognized the risk factors of PD, and closely related to sporadic and familial PD and can regulate a variety of cellular pathways and processes. An Increasing number of studies has shown that glial hyperactivation-mediated neuroinflammation participates in dopaminergic neuron degeneration and pathogenesis of PD. LRRK2 is essential to the regulaton of chronic inflammation, especially for the central nervous system. The review spotlights on the novel development of LRRK2 on microglia and astrocytes, and explore their potential therapeutic targets, in order to provide a new insights in PD. Key messages: What is already known on this topic The G2019S mutation of LRRK2 is now recognised as a pathogenic mutation in PD. Previous studies have focused on the relationship between neurons and LRRK2 G2019S. What this study adds Neuroinflammation mediated by LRRK2 G2019S of glial cells affects the progress and development of PD and attention must be paid to the role of LRRK2 G2019S in glial cells in PD. How this study might affect research, practice or policy Developing anti-inflammatory drugs from the perspective of LRRK2 G2019S of glial cells is a new direction for the treatment of PD.

Modeling Parkinson's disease in LRRK2 rodents.

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are associated with familial and sporadic forms of Parkinson's disease (PD). Sporadic PD and LRRK2 PD share main clinical and neuropathological features, namely hypokinesia, degeneration of nigro-striatal dopamine neurons and α-synuclein aggregates in the form of Lewy bodies. Animals harboring the most common LRRK2 mutations, i.e. p.G2019S and p.R1441C/G, have been generated to replicate the parkinsonian phenotype and investigate the underlying pathogenic mechanisms. Disappointingly, however, LRRK2 rodents did not consistently phenocopy hypokinesia and nigro-striatal degeneration, or showed Lewy body-like aggregates. Instead, LRRK2 rodents manifested non-motor signs and dysregulated transmission at dopaminergic and non-dopaminergic synapses that are reminiscent of behavioral and functional network changes observed in the prodromal phase of the disease. LRRK2 rodents also manifested greater susceptibility to different parkinsonian toxins or stressors when subjected to dual-hit or multiple-hit protocols, confirming LRRK2 mutations as genetic risk factors. In conclusion, LRRK2 rodents represent a unique tool to identify the molecular mechanisms through which LRRK2 modulates the course and clinical presentations of PD and to study the interplay between genetic, intrinsic and environmental protective/risk factors in PD pathogenesis.

ATM mediated DNA double‐strand breaks accumulate in LRRK2 G2019S Parkinson's disease

Parkinson's disease (PD) is the most common neurodegenerative movement disorder, affecting over one million people in the US. Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of inherited and idiopathic PD. We were the first to show that mitochondrial DNA (mtDNA) damage is caused by the most common mutation in LRRK2 (G2019S) and inhibition of LRRK2 kinase activity restores mtDNA integrity in PD models. However, whether aberrant LRRK2 kinase activity due to PD-linked mutations has broad impact on nuclear genome integrity is unknown. Using LRRK2G2019S/G2019S knock-in (KI) human embryonic kidney 293 (HEK293) cells obtained by CRISPR/Cas9 gene editing, our preliminary results indicate nuclear DNA damage is increased, including DNA double-strand breaks (DSBs) as assayed by a neutral comet assay. Consistent with DSB accumulation, we observed significantly increased γ-H2AX and 53BP1 foci. ATM is activated by DSBs and phosphorylates several key proteins that initiate the DNA damage response, cell cycle arrest, DNA repair or apoptosis. We found that basal levels of ATM pS1981 are increased in this in vitro LRRK2 G2019S model, which is an autophosphorylation site that correlates with DNA damage-mediated activation. Additionally, downstream substrates CHK2 (pT68), and P53 (pS15) are similarly increased in LRRK2G2019S/G2019S KI cells compared to isogenic wild-type control cells, substantiating that the ATM-mediated DNA damage response pathway has been up-regulated with the LRRK2 G2019S mutation. Blocking either LRRK2 or ATM kinase activity pharmacologically, significantly reversed LRRK2 G2019S-induced γ-H2AX foci. Overall these results suggest DSBs accumulate in LRRK2 PD, which in turn activate a sustained ATM mediated DNA damage response, which may lead to cell cycle arrest, aberrant DNA repair, and/or cell death. Further understanding of the functional relationship between LRRK2 and ATM offers new molecular insights into PD pathogenesis.

Association of Dual LRRK2 G2019S and GBA Variations With Parkinson Disease Progression

Despite a hypothesis that harboring a leucine-rich repeat kinase 2(LRRK2) G2019S variation and a glucocerebrosidase (GBA) variant would have a combined deleterious association with disease pathogenesis, milder clinical phenotypes have been reported in dual LRRK2 and GBA variations Parkinson disease (PD) than in GBA variation PD alone. To evaluate the association of LRRK2 G2019S and GBA variants with longitudinal cognitive and motor decline in PD. This longitudinal cohort study of continuous measures in LRRK2 PD, GBA PD, LRRK2/GBA PD, and wild-type idiopathic PD used pooled annual visit data ranging from 2004 to 2019 from the Mount Sinai Beth Israel, Parkinson Disease Biomarker Program, Harvard Biomarkers Study, Ashkenazi Jewish-LRRK2-Consortium, Parkinson Progression Marker Initiative, and SPOT-PD studies. Patients who were screened for GBA and LRRK2 variations and completed either a motor or cognitive assessment were included. Data were analyzed from May to July 2020. The associations of LRRK2 G2019S and GBA genotypes on the rate of decline in Montreal Cognitive Assessment (MoCA) and Movement Disorders Society-Unified Parkinson Disease Rating Scale-Part III scores were examined using linear mixed effects models with PD duration as the time scale. Among 1193 individuals with PD (mean [SD] age, 66.6 [9.9] years; 490 [41.2%] women), 128 (10.7%) had GBA PD, 155 (13.0%) had LRRK2 PD, 21 (1.8%) had LRRK2/GBA PD, and 889 (74.5%) had idiopathic PD. Patients with GBA PD had faster decline in MoCA than those with LRRK2/GBA PD (B [SE], -0.31 [0.09] points/y; P < .001), LRRK2 PD (B [SE], -0.33 [0.09] points/y; P < .001), or idiopathic PD (B [SE], -0.23 [0.08] points/y; P = .005). There was a LRRK2 G2019S × GBA interaction in MoCA decline (B [SE], 0.22 [0.11] points/y; P = .04), but not after excluding severe GBA variations (B [SE], 0.12 [0.11] points/y; P = .28). Patients with GBA PD had significantly worse motor progression compared with those with idiopathic PD (B [SE], 0.49 [0.22] points/y; P = .03) or LRRK2 PD (B [SE], 0.77 [0.26] points/y; P = .004). These findings suggest that longitudinal cognitive decline in patients with GBA PD was more severe than in those with LRRK2/GBA PD, which more closely resembled LRRK2 PD. This further supports the notion of a dominant association of LRRK2 on GBA in individuals who carry both and raises the possibility of an LRRK2 × GBA interaction. However, the biological basis of a dominant association or interaction is not clear and is apparently contrary to basic investigations. Study of a larger cohort of individuals with severe GBA variation is warranted.

Differential Inhibition of LRRK2 in Parkinson's Disease Patient Blood by a G2019S Selective LRRK2 Inhibitor.

ABSTRACTBackgroundA common genetic mutation that causes Parkinson's disease (PD) is the G2019S LRRK2 mutation. A precision medicine approach that selectively blocks only excess kinase activity of the mutant allele could yield a safe and effective treatment for G2019S LRRK2 PD.ObjectiveTo determine the activity of a G2019S mutant selective leucine‐rich repeat kinase 2 (LRRK2) kinase inhibitor as compared to a nonselective inhibitor in blood of subjects with genetic and idiopathic PD on two LRRK2 biomarkers, pSer935 LRRK2 and pThr73 Rab10.MethodsBlood was collected from 13 subjects with or without a G2019S LRRK2 mutation with PD and one healthy control. Peripheral blood mononuclear cells were treated ex vivo with a novel G2019S LRRK2 inhibitor (EB‐42168) or the nonselective inhibitor MLi‐2. Quantitative western immunoblot analyses were performed.ResultsEB‐42168 was 100 times more selective for G2019S LRRK2 when compared to wild‐type (WT) LRRK2. Concentrations that inhibited phosphorylation of pSer935 LRRK2 by 90% in homozygous G2019S LRRK2 patients, inhibited pSer935 LRRK2 by 36% in heterozygous patients, and by only 5% in patients carrying only the WT allele. Similar selectivity was seen for pThr73 Rab10. MLi‐2 showed an equivalent level of inhibition across all genotypes.ConclusionsThese findings demonstrate that EB‐42168, a G2019S LRRK2 selective inhibitor, lowers mutant G2019S LRRK2 phosphorylated biomarkers while simultaneously sparing WT LRRK2. Selective targeting of G2019S LRRK2 with a small molecule lays the foundation for a precision medicine treatment of G2019S LRRK2 PD. © 2021 ESCAPE Bio, Inc. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society

Fine-mapping of the non-coding variation driving the Caucasian LRRK2 GWAS signal in Parkinson's disease

IntroductionGenome-wide association studies (GWAS) have confirmed the leucine-rich repeat kinase 2 (LRRK2) gene as a susceptibility locus for idiopathic Parkinson's disease (PD) in Caucasians. Though the rs1491942 and rs76904798 variants have shown the strongest associations, the causal variant(s) remains unresolved. Therefore, the aim of this study was to identify variants that may be driving the LRRK2 GWAS signal by sequencing the entire LRRK2 gene in Caucasian PD patients and controls. MethodsA discovery series (287 PD patients, 294 controls) and replication series (362 PD patients, 168 controls) were included. The entire LRRK2 gene as well as 10 Kb upstream/downstream was sequenced. Candidate potential causal variants were considered to be those that (a) were in at least weak linkage disequilibrium with the two GWAS-nominated variants (rs1491942 and rs76904798), and (b) displayed an association odds ratio (OR) that is stronger than the two GWAS variants. ResultsThirty-four candidate variants (all intronic/intergenic) that may drive the LRRK2 PD GWAS signal were identified in the discovery series. However, examination of the replication series for these variants did not reveal any with a consistently stronger OR than both PD GWAS variants. Evaluation of public databases to determine which candidate variants are most likely to have a direct functional effect on LRRK2 expression was inconclusive. ConclusionThough our findings provide novel insights into the LRRK2 GWAS association, a clear causal variant was not identified. The identified candidate variants can form the basis for future experiments and functional studies that can more definitively assess causal LRRK2 variants.

Low Levels of LRRK2 Gene Expression are Associated with LRRK2 SNPs and Contribute to Parkinson's Disease Progression.

Parkinson's disease (PD) is a chronic neurodegenerative disease that has relatively slow progression with motor symptoms. Leucine-rich repeat kinase 2 (LRRK2) gene mutations and polymorphisms are suggested to be associated with PD. In this study, we aimed to investigate the association between single-nucleotide polymorphisms (SNPs) of the LRRK2 gene, namely, rs11176013, rs10878371, rs11835105, and PD. Genotypes of 132 PD cases and 133 healthy individuals were determined by qRT-PCR. Haplotype analysis was performed. Additionally, LRRK2 mRNA expression levels were determined in 83 PD cases and 55 healthy subjects. The relationship between LRRK2 mRNA levels, the target SNPs, and clinical data was also investigated. Our results indicated that the "GG" genotype and "G" allele of rs11176013 and the "CC" genotype and "C" allele of rs10878371 were more frequent in cases. The "GCG" haplotype was significantly more frequent in cases. LRRK2 mRNA expression levels in patients were significantly lower than those in healthy individuals. The patients with the "CC" genotype for rs10878371 and the "GG" genotype for rs11176013 had decreased LRRK2 mRNA levels. We found that the rs11176013 "GG" genotype and the rs10878371 "CC" genotype were less frequently seen in cases with akinetic rigid or combined akinetic rigid and tremor-dominant initial symptoms. Consequently, our results demonstrate that the rs11176013 and rs10878371 polymorphisms are associated with PD in a Turkish cohort, and moreover, these results suggest that these polymorphisms may affect the expression of the LRRK2 gene and disease progression and thus play a role in the pathogenesis of PD.

Nonsteroidal Anti-inflammatory Use and LRRK2 Parkinson's Disease Penetrance.

The penetrance of leucine rich repeat kinase 2 (LRRK2) mutations is incomplete and may be influenced by environmental and/or other genetic factors. Nonsteroidal anti-inflammatory drugs (NSAIDs) are known to reduce inflammation and may lower Parkinson's disease (PD) risk, but their role in LRRK2-associated PD is unknown. The objective of this study is to evaluate the association of regular NSAID use and LRRK2-associated PD. Symptomatic ("LRRK2-PD") and asymptomatic ("LRRK2-non-PD") participants with LRRK2 G2019S, R1441X, or I2020T variants (definitely pathogenic variant carriers) or G2385R or R1628P variants (risk variant carriers) from 2 international cohorts provided information on regular ibuprofen and/or aspirin use (≥2 pills/week for ≥6 months) prior to the index date (diagnosis date for PD, interview date for non-PD). Multivariate logistic regression was used to evaluate the relationship between regular NSAID use and PD for any NSAID, separately for ibuprofen and aspirin in all carriers and separately in pathogenic and risk variant groups. A total of 259 LRRK2-PD and 318 LRRK2-non-PD participants were enrolled. Regular NSAID use was associated with reduced odds of PD in the overall cohort (odds ratio [OR], 0.34; 95% confidence interval [CI], 0.21-0.57) and in both pathogenic and risk variant carriers (ORPathogenic , 0.38; 95% CI, 0.21-0.67 and ORRiskVariant , 0.19; 95% CI, 0.04-0.99). Similar associations were observed for ibuprofen and aspirin separately (ORIbuprofen , 0.19; 95% CI, 0.07-0.50 and ORAspirin , 0.51; 95% CI, 0.28-0.91). Regular NSAID use may be associated with reduced penetrance in LRRK2-associated PD. The LRRK2 protein is involved in inflammatory pathways and appears to be modulated by regular anti-inflammatory use. Longitudinal observational and interventional studies of NSAID exposure and LRRK2-PD are needed to confirm this association. © 2020 International Parkinson and Movement Disorder Society.

Allosteric modulation of the GTPase activity of a bacterial LRRK2 homolog by conformation-specific Nanobodies

Mutations in the Parkinson's disease (PD)-associated protein leucine-rich repeat kinase 2 (LRRK2) commonly lead to a reduction of GTPase activity and increase in kinase activity. Therefore, strategies for drug development have mainly been focusing on the design of LRRK2 kinase inhibitors. We recently showed that the central RocCOR domains (Roc: Ras of complex proteins; COR: C-terminal of Roc) of a bacterial LRRK2 homolog cycle between a dimeric and monomeric form concomitant with GTP binding and hydrolysis. PD-associated mutations can slow down GTP hydrolysis by stabilizing the protein in its dimeric form. Here, we report the identification of two Nanobodies (NbRoco1 and NbRoco2) that bind the bacterial Roco protein (CtRoco) in a conformation-specific way, with a preference for the GTP-bound state. NbRoco1 considerably increases the GTP turnover rate of CtRoco and reverts the decrease in GTPase activity caused by a PD-analogous mutation. We show that NbRoco1 exerts its effect by allosterically interfering with the CtRoco dimer–monomer cycle through the destabilization of the dimeric form. Hence, we provide the first proof of principle that allosteric modulation of the RocCOR dimer–monomer cycle can alter its GTPase activity, which might present a potential novel strategy to overcome the effect of LRRK2 PD mutations.

Clinical and Dopamine Transporter Imaging Characteristics of Leucine Rich Repeat Kinase 2 (LRRK2) and Glucosylceramidase Beta (GBA) Parkinson's Disease Participants in the Parkinson's Progression Markers Initiative: A Cross-Sectional Study.

There are limited data on the phenotypic and dopamine transporter (DAT) imaging characterization of the Parkinson's disease (PD) patients with leucine rich kinase 2 (LRRK2) and glucosylceramidase beta (GBA) mutations. The objective of this study was to examine baseline clinical and DAT imaging characteristics in GBA and LRRK2 mutation carriers with early PD compared with sporadic PD. The Parkinson's Progression Markers Initiative is an ongoing observational longitudinal study that enrolled participants with sporadic PD, LRRK2 and GBA PD carriers from 33 sites worldwide. All participants are assessed annually with a battery of motor and nonmotor scales, 123-I Ioflupane DAT imaging, and biologic variables. We assessed 158 LRRK2 (89% G2019S), 80 GBA (89 %N370S), and 361 sporadic PD participants with the mean (standard deviation) disease duration of 2.9 (1.9), 3.1 (2.0), and 2.6 (0.6) years, respectively. When compared with sporadic PD, the GBA PD patients had no difference in any motor, cognitive, or autonomic features. The LRRK2 PD patients had less motor disability and lower rapid eye movement behavior disorder questionnaire scores, but no meaningful difference in cognitive or autonomic features. Both genetic cohorts had a higher score on the impulse control disorders scale when compared with sporadic PD, but no difference in other psychiatric features. Both genetic PD cohorts had less loss of dopamine transporter on DAT imaging when compared with sporadic PD. We confirm previous reports of milder phenotype associated with LRRK2-PD. A previously reported more aggressive phenotype in GBA-PD is not evident early in the disease in N370s carriers. This observation identifies a window for potential disease-modifying interventions. Longitudinal data will be essential to define the slope of progression for both genetic cohorts. ClinicalTrials.gov (NCT01141023). © 2020 The Authors. Movement Disorders published by Wiley Periodicals, Inc. on behalf of International Parkinson and Movement Disorder Society.

The In situ Structure of Parkinson's Disease-Linked LRRK2

Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most frequent cause of familial Parkinson's disease (PD). LRRK2 is a large multi-domain protein containing a kinase and GTPase surrounded by multiple domains involved in protein interactions. LRRK2 involved in various biological processes including cytoskeletal integrity, membrane trafficking, autophagy, and mitochondrial homeostasis. LRRK phosphorylates a subset of Rab GTPases and modulates their membrane localization and function. Most of the PD pathogenic mutations are found in the catalytic domains, but all result in hyperactivation of the kinase. This suggests that crosstalk between the GTPase and kinase domains is critical for the regulation of LRRK2 kinase activity. Hyperactivation of LRRK2's kinase is also reported in non-familial PD cases, suggesting that suppression of kinase activity might be beneficial for a wide range of PD patients. Despite being a promising drug target, LRRK2 has eluded structural determination due to the difficulties to purify active full-length LRRK2 protein from cells. Interestingly, LRRK2 has been shown to form filamentous structures surrounding microtubules in cells and several PD mutants or treatment of LRRK2 kinase inhibitor enhance its filament formation. Using in situ cryo-electron tomography and subtomogram averaging, we reveal a 14-Å structure of LRRK2 bearing a pathogenic mutation that oligomerizes as a right-handed double-helix around microtubules, which are left-handed. Using integrative modeling, we determine the architecture of LRRK2, revealing that the GTPase points towards the microtubule, while the kinase is exposed to the cytoplasm. We identify several oligomerization interfaces mediated by non-catalytic regions of LRRK2. Mutation of one of these abolishes LRRK2 microtubule-association in cells. Our work demonstrates the power of cryo-electron tomography to obtain structures of previously unsolved proteins in their cellular environment and provides insights into LRRK2 function and PD pathogenicity.

Long-Term Exposure to PFE-360 in the AAV-α-Synuclein Rat Model: Findings and Implications.

Parkinson's disease (PD) is a progressive neurodegenerative disorder associated with impaired motor function and several non-motor symptoms, with no available disease modifying treatment. Intracellular accumulation of pathological α-synuclein inclusions is a hallmark of idiopathic PD, whereas, dominant mutations in leucine-rich repeat kinase 2 (LRRK2) are associated with familial PD that is clinically indistinguishable from idiopathic PD. Recent evidence supports the hypothesis that an increase in LRRK2 kinase activity is associated with the development of not only familial LRRK2 PD, but also idiopathic PD. Previous reports have shown preclinical effects of LRRK2 modulation on α-synuclein-induced neuropathology. Increased subthalamic nucleus (STN) burst firing in preclinical neurotoxin models and PD patients is hypothesized to be causally involved in the development of the motor deficit in PD. To study a potential pathophysiological relationship between α-synuclein pathology and LRRK2 kinase activity in PD, we investigated the effect of chronic LRRK2 inhibition in an AAV-α-synuclein overexpression rat model. In this study, we report that chronic LRRK2 inhibition using PFE-360 only induced a marginal effect on motor function. In addition, the aberrant STN burst firing and associated neurodegenerative processes induced by α-synuclein overexpression model remained unaffected by chronic LRRK2 inhibition. Our findings do not strongly support LRRK2 inhibition for the treatment of PD. Therefore, the reported beneficial effects of LRRK2 inhibition in similar α-synuclein overexpression rodent models must be considered with prudence and additional studies are warranted in alternative α-synuclein-based models.

Generation of an induced pluripotent stem cell line (CSC-41) from a Parkinson's disease patient carrying a p.G2019S mutation in the LRRK2 gene

The leucine-rich repeat kinase 2 (LRRK2) p.G2019S mutation is the most common genetic cause of Parkinson's disease (PD). An induced pluripotent stem cell (iPSC) line CSC-41 was generated from a 75-year old patient diagnosed with PD caused by a p.G2019S mutation in LRRK2. Skin fibroblasts were reprogrammed using a non-integrating Sendai virus-based technology to deliver OCT3/4, SOX2, c-MYC and KLF4 transcription factors. The generated iPSC line exhibits expression of common pluripotency markers, differentiates into the three germ layers and has a normal karyotype. The iPSC line can be used to explore the association between LRRK2 mutation and PD.

Elevated LRRK2 autophosphorylation in brain-derived and peripheral exosomes in LRRK2 mutation carriers

Missense mutations in the leucine-rich repeat kinase 2 (LRRK2) gene can cause late-onset Parkinson disease (PD). LRRK2 mutations increase LRRK2 kinase activities that may increase levels of LRRK2 autophosphorylation at serine 1292 (pS1292) and neurotoxicity in model systems. pS1292-LRRK2 protein can be packaged into exosomes and measured in biobanked urine. Herein we provide evidence that pS1292-LRRK2 protein is robustly expressed in cerebral spinal fluid (CSF) exosomes. In a novel cohort of Norwegian subjects with and without the G2019S-LRRK2 mutation, with and without PD, we quantified levels of pS1292-LRRK2, total LRRK2, and other exosome proteins in urine from 132 subjects and in CSF from 82 subjects. CSF and urine were collected from the same morning clinic visit in 55 of the participants. We found that total LRRK2 protein concentration was similar in exosomes purified from either CSF or urine but the levels did not correlate. pS1292-LRRK2 levels were higher in urinary exosomes from male and female subjects with a LRRK2 mutation. Male LRRK2 mutation carriers without PD had intermediate pS1292-LRRK2 levels compared to male carriers with PD and controls. However, female LRRK2 mutation carriers without PD had the same pS1292-LRRK2 levels compared to female carriers with PD. pS1292-LRRK2 levels in CSF exosomes were near saturated in most subjects, ten-fold higher on average than pS1292-LRRK2 levels in urinary exosomes, irrespective of LRRK2 mutation status or PD diagnosis. These results provide insights into the effects of LRRK2 mutations in both the periphery and brain in a well-characterized clinical population and show that LRRK2 protein in brain exosomes may be much more active than in the periphery in most subjects.

Relationship between LRRK2 R1628P polymorphism and Parkinson's disease in Asian populations

Although the leucine-rich repeat kinase 2 (LRRK2) R1628P polymorphism has been associated with the risk of Parkinson's disease (PD) in Taiwan, China, and Singapore, there are conflicting findings regarding this relationship. Thus, the aim of the present meta-analysis was to evaluate the associations between the LRRK2 R1628P polymorphism (rs33949390) and PD in Asian populations. A search for eligible studies was performed in PubMed, Embase, SinoMed, and the China Knowledge Resource Integrated Database, and pooled odds ratios and 95% confidence intervals were used to evaluate the strength of the association between the R1628P polymorphism and PD. This meta-analysis assessed 19 studies from 14 papers that involved a total of 9,927 PD patients and 8,602 controls and found that the R1628P polymorphism was significantly associated with the risk of PD in Asian populations. Moreover, stratification analyses indicated that the R1628P polymorphism was significantly associated with an increased risk of PD among Chinese as well as non-Chinese Asian populations and an increased risk of PD in Chinese patients from China, Taiwan, and Singapore. In a stratified analysis conducted according to age, significant associations were found for both late-onset PD and early-onset PD. The present data indicate that the R1628P polymorphism of the LRRK2 gene contributes to PD susceptibility in Asian, especially Chinese, populations.

LRRK2 delays degradative receptor trafficking by impeding late endosomal budding through decreasing Rab7 activity.

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene cause late-onset autosomal dominant Parkinson's disease (PD), and sequence variations at the LRRK2 locus are associated with increased risk for sporadic PD. LRRK2 contains both GTPase and kinase domains flanked by protein interaction motifs, and mutations associated with familial PD have been described for both catalytic domains. LRRK2 has been implicated in diverse cellular processes, and recent evidence pinpoints to an important role for LRRK2 in modulating a variety of intracellular membrane trafficking pathways. However, the underlying mechanisms are poorly understood. Here, by studying the classical, well-understood, degradative trafficking pathway of the epidermal growth factor receptor (EGFR), we show that LRRK2 regulates endocytic membrane trafficking in an Rab7-dependent manner. Mutant LRRK2 expression causes a slight delay in early-to-late endosomal trafficking, and a pronounced delay in trafficking out of late endosomes, which become aberrantly elongated into tubules. This is accompanied by a delay in EGFR degradation. The LRRK2-mediated deficits in EGFR trafficking and degradation can be reverted upon coexpression of active Rab7 and of a series of proteins involved in bridging the EGFR to Rab7 on late endosomes. Effector pulldown assays indicate that pathogenic LRRK2 decreases Rab7 activity both in cells overexpressing LRRK2, as well as in fibroblasts from pathogenic mutant LRRK2 PD patients when compared with healthy controls. Together, these findings provide novel insights into a previously unknown regulation of Rab7 activity by mutant LRRK2 which impairs membrane trafficking at very late stages of the endocytic pathway.

Presynaptic dysfunction in Parkinson's disease: a focus on LRRK2

PD (Parkinson's disease) is a common neurodegenerative disease clinically characterized by bradykinesia, rigidity and resting tremor. Recent studies have proposed that synaptic dysfunction, implicated in numerous studies of animal models of PD, might be a key factor in PD. The molecular defects that lead to PD progression might be hidden at the presynaptic neuron: in fact accumulating evidence has shown that the majority of the genes linked to PD play a critical role at the presynaptic site. In the present paper, we focus on the presynaptic function of LRRK2 (leucine-rich repeat kinase 2), a protein that mutated represents the main genetic cause of familial PD described to date. Neurotransmission relies on proper presynaptic vesicle trafficking; defects in this process, variation in dopamine flow and alteration of presynaptic plasticity have been reported in several animal models of LRRK2 mutations. Furthermore, impaired dopamine turnover has been described in presymptomatic LRRK2 PD patients. Thus, given the pathological events occurring at the synapses of PD patients, the presynaptic site may represent a promising target for early diagnostic therapeutic intervention.

Mouse models for LRRK2 Parkinson's disease

Parkinson's disease (PD) is the second most common neurodegenerative disease. Mutations in Leucine-rich-repeat-kinase 2 (LRRK2), the causative gene for PARK8 type PD with autosomal dominant inheritance, are the most prevalent genetic causes of both familial and sporadic PD. Animal models are critical tools in the attempt to understand the mechanisms of LRRK2-mediated pathogenesis. We have generated human Bacterial Artificial Chromosome (BAC) mediated transgenic mouse models expressing mutant LRRK2 that robustly recapitulate the behavioral, neurochemical and pathological features of PD. These mice develop an age-dependent decrease in motor activity that is progressive and responds to treatment with levodopa. Pathologically, the most salient phenotype is early axonopathy of nigrostriatal dopaminergic neurons, accompanied by hyperphosphorylated tau. The mice also exhibit a consistent dopamine transmission deficit in both acute brain slices and live freely moving animals. Here we will discuss LRRK2 mouse models from several laboratories, their commonalities and differences, and offer scientific insights drawn from these studies.

LRRK2 Parkinson disease mutations enhance its microtubule association

Dominant missense mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common genetic causes of Parkinson disease (PD) and genome-wide association studies identify LRRK2 sequence variants as risk factors for sporadic PD. Intact kinase function appears critical for the toxicity of LRRK2 PD mutants, yet our understanding of how LRRK2 causes neurodegeneration remains limited. We find that most LRRK2 PD mutants abnormally enhance LRRK2 oligomerization, causing it to form filamentous structures in transfections of cell lines or primary neuronal cultures. Strikingly, ultrastructural analyses, including immuno-electron microscopy and electron microscopic tomography, demonstrate that these filaments consist of LRRK2 recruited onto part of the cellular microtubule network in a well-ordered, periodic fashion. Like LRRK2-related neurodegeneration, microtubule association requires intact kinase function and the WD40 domain, potentially linking microtubule binding and neurodegeneration. Our observations identify a novel effect of LRRK2 PD mutations and highlight a potential role for microtubules in the pathogenesis of LRRK2-related neurodegeneration.

The Parkinson's disease protein LRRK2 impairs proteasome substrate clearance without affecting proteasome catalytic activity

Leucine-rich repeat kinase 2 (LRRK2) mutations are the most common known cause of Parkinson's disease (PD). The clinical features of LRRK2 PD are indistinguishable from idiopathic PD, with accumulation of α-synuclein and/or tau and/or ubiquitin in intraneuronal aggregates. This suggests that LRRK2 is a key to understanding the aetiology of the disorder. Although loss-of-function does not appear to be the mechanism causing PD in LRRK2 patients, it is not clear how this protein mediates toxicity. In this study, we report that LRRK2 overexpression in cells and in vivo impairs the activity of the ubiquitin-proteasome pathway, and that this accounts for the accumulation of diverse substrates with LRRK2 overexpression. We show that this is not mediated by large LRRK2 aggregates or sequestration of ubiquitin to the aggregates. Importantly, such abnormalities are not seen with overexpression of the related protein LRRK1. Our data suggest that LRRK2 inhibits the clearance of proteasome substrates upstream of proteasome catalytic activity, favouring the accumulation of proteins and aggregate formation. Thus, we provide a molecular link between LRRK2, the most common known cause of PD, and its previously described phenotype of protein accumulation.