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1.
Nat Rev Neurol ; 16(2): 97-107, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31980808

RESUMO

One of the most common monogenic forms of Parkinson disease (PD) is caused by mutations in the LRRK2 gene that encodes leucine-rich repeat kinase 2 (LRRK2). LRRK2 mutations, and particularly the most common mutation Gly2019Ser, are observed in patients with autosomal dominant PD and in those with apparent sporadic PD, who are clinically indistinguishable from those with idiopathic PD. The discoveries that pathogenic mutations in the LRRK2 gene increase LRRK2 kinase activity and that small-molecule LRRK2 kinase inhibitors can be neuroprotective in preclinical models of PD have placed LRRK2 at the centre of disease modification efforts in PD. Recent investigations also suggest that LRRK2 has a role in the pathogenesis of idiopathic PD and that LRRK2 therapies might, therefore, be beneficial in this common subtype of PD. In this Review, we describe the characteristics of LRRK2-associated PD that are most relevant to the development of LRRK2-targeted therapies and the design and implementation of clinical trials. We highlight strategies for correcting the effects of mutations in the LRRK2 gene, focusing on how to identify which patients are the optimal candidates and how to decide on the timing of such trials. In addition, we discuss challenges in implementing trials of disease-modifying treatment in people who carry LRRK2 mutations.


Assuntos
Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Doença de Parkinson/genética , Antiparkinsonianos/farmacologia , Ensaios Clínicos como Assunto , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/efeitos dos fármacos , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/epidemiologia
2.
Biochem Soc Trans ; 47(6): 1581-1595, 2019 12 20.
Artigo em Inglês | MEDLINE | ID: mdl-31769472

RESUMO

Mutations in the leucine-rich-repeat kinase 2 (LRRK2) gene are associated with familial and sporadic cases of Parkinson's disease but are also found in immune-related disorders such as inflammatory bowel disease, tuberculosis and leprosy. LRRK2 is highly expressed in immune cells and has been functionally linked to pathways pertinent to immune cell function, such as cytokine release, autophagy and phagocytosis. Here, we examine the current understanding of the role of LRRK2 kinase activity in pathway regulation in immune cells, drawing upon data from multiple diseases associated with LRRK2 to highlight the pleiotropic effects of LRRK2 in different cell types. We discuss the role of the bona fide LRRK2 substrate, Rab GTPases, in LRRK2 pathway regulation as well as downstream events in the autophagy and inflammatory pathways.


Assuntos
Sistema Imunitário/fisiologia , Inflamação/fisiopatologia , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/fisiologia , Animais , Humanos , Inflamação/imunologia , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Mutação
3.
BMC Neurol ; 19(1): 260, 2019 Oct 28.
Artigo em Inglês | MEDLINE | ID: mdl-31660902

RESUMO

BACKGROUND: Parkinsonism is a complex multifactorial neurodegenerative disorder, in which genetic and environmental risk factors may both play a role. Among environmental risk factors cocaine was earlier ambiguously linked to Parkinsonism. Former single case reports described Parkinsonism in chronic cocaine users, but an epidemiological study did not confirm an increased risk of Parkinson's disease. Here we report a patient, who developed Parkinsonism in young age after chronic cocaine use, in whom a homozygous LRRK2 risk variant was also detected. CASE PRESENTATION: The patient was investigated because of hand tremor, which started after a 1.5-year period of cocaine abuse. Neurological examination suggested Parkinsonism, and asymmetrical pathology was confirmed by the dopamine transporter imaging study. The genetic investigations revealed a homozygous risk allele in the LRRK2 gene. After a period of cocaine abstinence, the patient's symptoms spontaneously regressed, and the dopamine transporter imaging also returned to near-normal. CONCLUSIONS: This case report suggests that cocaine abuse indeed might be linked to secondary Parkinsonism and serves as an example of a potential gene-environmental interaction between the detected LRRK2 risk variant and cocaine abuse. The reversible nature of the DaTscan pathology is a unique feature of this case, and needs further evaluation, whether this is incidental or can be a feature of cocaine related Parkinsonism.


Assuntos
Transtornos Relacionados ao Uso de Cocaína/complicações , Interação Gene-Ambiente , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Doença de Parkinson/etiologia , Adulto , Encéfalo/diagnóstico por imagem , Encéfalo/patologia , Proteínas da Membrana Plasmática de Transporte de Dopamina/metabolismo , Humanos , Masculino , Neuroimagem/métodos , Doença de Parkinson/diagnóstico por imagem , Doença de Parkinson/patologia , Fatores de Risco , Tomografia Computadorizada com Tomografia Computadorizada de Emissão de Fóton Único
5.
Med Hypotheses ; 131: 109302, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31443765

RESUMO

Parkinson's disease (PD) patients have higher rates of melanoma and vice versa, observations suggesting that the two conditions may share common pathogenic pathways. ß-Catenin is a transcriptional cofactor that, when concentrated in the nucleus, upregulates the expression of canonical Wnt target genes, such as Nurr1, many of which are important for neuronal survival. ß-Catenin-mediated activity is decreased in sporadic PD as well as in leucine-rich repeat kinase 2 (LRRK2) and ß-glucosidase (GBA) mutation cellular models of PD, which is the most common genetic cause of and risk for PD, respectively. In addition, ß-catenin expression is significantly decreased in more aggressive and metastatic melanoma. Multiple observational studies have shown smokers to have significantly lower rates of PD as well as melanoma implying that tobacco may contain one or more elements that protect against both conditions. In support, smoker's brains have significantly reduced levels of α-synuclein, a pathological intracellular protein found in PD brain and melanoma cells. Tobacco contains very high lithium levels compared to other plants. Lithium has a broad array of neuroprotective actions, including enhancing autophagy and reducing intracellular α-synuclein levels, and is effective in both neurotoxin and transgenic preclinical PD models. One of lithium's neuroprotective actions is enhancement of ß-catenin-mediated activity leading to increased Nurr1 expression through its ability to inhibit glycogen synthase kinase-3 ß (GSK-3ß). Lithium also has anti-proliferative effects on melanoma cells and the clinical use of lithium is associated with a reduced incidence of melanoma as well as reduced melanoma-associated mortality. This is the first known report hypothesizing that inhaled lithium from smoking may account for the associated reduced rates of both PD and melanoma and that this protection may be mediated, in part, through lithium-induced GSK-3ß inhibition and consequent enhanced ß-catenin-mediated activity. This hypothesis could be directly tested in clinical trials assessing lithium therapy's ability to affect ß-catenin-mediated activity and slow disease progression in patients with PD or melanoma.


Assuntos
Lítio/farmacologia , Melanoma/prevenção & controle , Modelos Biológicos , Fármacos Neuroprotetores/farmacologia , Doença de Parkinson/prevenção & controle , Fumantes , Tabaco/química , Via de Sinalização Wnt/efeitos dos fármacos , beta Catenina/fisiologia , Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/epidemiologia , Doença de Alzheimer/prevenção & controle , Autofagia/efeitos dos fármacos , Química Encefálica/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Glicogênio Sintase Quinase 3 beta/antagonistas & inibidores , Glicogênio Sintase Quinase 3 beta/fisiologia , Humanos , Incidência , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Lítio/análise , Lítio/uso terapêutico , Carbonato de Lítio/uso terapêutico , Melanoma/epidemiologia , Mutação , Fármacos Neuroprotetores/análise , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares/biossíntese , Membro 2 do Grupo A da Subfamília 4 de Receptores Nucleares/genética , Doença de Parkinson/epidemiologia , Transtornos Parkinsonianos/tratamento farmacológico , Água/química , Via de Sinalização Wnt/fisiologia , alfa-Sinucleína/metabolismo , beta-Glucosidase/genética
6.
Nat Neurosci ; 22(8): 1248-1257, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31346295

RESUMO

Studies of patients afflicted by neurodegenerative diseases suggest that misfolded proteins spread through the brain along anatomically connected networks, prompting progressive decline. Recently, mouse models have recapitulated the cell-to-cell transmission of pathogenic proteins and neuron death observed in patients. However, the factors regulating the spread of pathogenic proteins remain a matter of debate due to an incomplete understanding of how vulnerability functions in the context of spread. Here we use quantitative pathology mapping in the mouse brain, combined with network modeling to understand the spatiotemporal pattern of spread. Patterns of α-synuclein pathology are well described by a network model that is based on two factors: anatomical connectivity and endogenous α-synuclein expression. The map and model allow the assessment of selective vulnerability to α-synuclein pathology development and neuron death. Finally, we use quantitative pathology to understand how the G2019S LRRK2 genetic risk factor affects the spread and toxicity of α-synuclein pathology.


Assuntos
Encéfalo/patologia , Conectoma/psicologia , alfa-Sinucleína/genética , Animais , Mapeamento Encefálico , Morte Celular , Feminino , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Modelos Lineares , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Neurológicos , Neurônios/patologia
7.
Int J Oncol ; 55(1): 21-34, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31180559

RESUMO

Emerging studies have indicated that leucine­rich repeat kinase 2 (LRRK2) is associated with thyroid cancer (TC). The present study investigated the effect of LRRK2 on the cell cycle and apoptosis in TC, and examined the underlying mechanisms in vitro. To screen TC­associated differentially expressed genes, gene expression microarray analysis was conducted. Retrieval of pathways associated with TC from the Kyoto Encyclopedia of Genes and Genomes database indicated that the c­Jun N­terminal kinase (JNK) signaling pathway serves an essential role in TC. SW579, IHH­4, TFC­133, TPC­1 and Nthy­ori3­1 cell lines were used to screen cell lines with the highest and lowest LRRK2 expression for subsequent experiments. The two selected cell lines were transfected with pcDNA­LRRK2, or small interfering RNA against LRRK2 or SP600125 (a JNK inhibitor). Subsequently, flow cytometry, terminal deoxynucleotidyl transferase­mediated dUTP­biotin nick end labeling, a 5­ethynyl­2'­deoxyuridine assay and a scratch test was conducted to detect the cell cycle distribution, apoptosis, proliferation and migration, respectively, in each group. The LRRK2 gene was determined to be elevated in TC based on the microarray data of the GSE3678 dataset. The SW579 cell line was identified to exhibit the highest LRRK2 expression, while IHH­4 cells exhibited the lowest LRRK2 expression. LRRK2 silencing, through inhibiting the activation of the JNK signaling pathway, increased the expression levels of genes and proteins associated with cell cycle arrest and apoptosis in TC cells, promoted cell cycle arrest and apoptosis, and inhibited cell migration and proliferation in TC cells, indicating that LRRK2 repression could exert beneficial effects through the JNK signaling pathway on TC cells. These observations demonstrate that LRRK2 silencing promotes TC cell growth inhibition, and facilitates apoptosis and cell cycle arrest. The JNK signaling pathway may serve a crucial role in mediating the anti­carcinogenic activities of downregulated LRRK2 in TC.


Assuntos
Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/biossíntese , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Sistema de Sinalização das MAP Quinases , Neoplasias da Glândula Tireoide/enzimologia , Neoplasias da Glândula Tireoide/genética , Antracenos/farmacologia , Apoptose/fisiologia , Pontos de Checagem do Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Regulação para Baixo , Ativação Enzimática , Humanos , MAP Quinase Quinase 4/antagonistas & inibidores , MAP Quinase Quinase 4/metabolismo , Neoplasias da Glândula Tireoide/patologia , Transfecção
8.
Pharmacol Res Perspect ; 7(3): e00484, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31149340

RESUMO

Mutations in leucine-rich repeat kinase 2 (LRRK2) gene have been pathogenically linked to Parkinson's disease, and pharmacological inhibition of LRRK2 is being pursued to tackle nigro-striatal dopaminergic neurodegeneration. However, LRRK2 kinase inhibitors may have manifold actions, affecting not only pathological mechanisms in dopaminergic neurons but also physiological functions in nondopaminergic neurons. Therefore, we investigated whether LRRK2 kinase inhibitors differentially modulate dopamine and glutamate release from the mouse striatum and cerebral cortex. Spontaneous and KCl-evoked [3H]-dopamine and glutamate release from superfused synaptosomes obtained from wild-type and LRRK2 knock-out, kinase-dead or G2019S knock-in mice was measured. Two structurally unrelated inhibitors, LRRK2-IN-1 and GSK2578215A, were tested. LRRK2, phosphoSerine1292 and phosphoSerine935 LRRK2 levels were measured in all genotypes, and target engagement was evaluated by monitoring phosphoSerine935 LRRK2. LRRK2-IN-1 inhibited striatal glutamate but not dopamine release; GSK2578215A inhibited striatal dopamine and cortical glutamate but enhanced striatal glutamate release. LRRK2-IN-1 reduced striatal and cortical phosphoSerine935 levels whereas GSK2578215A inhibited only the former. Neither LRRK2 inhibitor affected neurotransmitter release in LRRK2 knock-out and kinase-dead mice; however, they facilitated dopamine without affecting striatal glutamate in G2019S knock-in mice. GSK2578215A inhibited cortical glutamate release in G2019S knock-in mice. We conclude that LRRK2-IN-1 and GSK2578215A modulate exocytosis by blocking LRRK2 kinase activity, although their effects vary depending on the nerve terminal examined. The G2019S mutation unravels a dopamine-promoting action of LRRK2 inhibitors while blunting their effects on glutamate release, which highlights their positive potential for the treatment of PD, especially of LRRK2 mutation carriers.


Assuntos
Aminopiridinas/farmacologia , Benzamidas/farmacologia , Benzodiazepinonas/farmacologia , Corpo Estriado/citologia , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Pirimidinas/farmacologia , Córtex Visual/citologia , Animais , Corpo Estriado/efeitos dos fármacos , Corpo Estriado/metabolismo , Dopamina/metabolismo , Exocitose , Técnicas de Introdução de Genes , Ácido Glutâmico/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/antagonistas & inibidores , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/química , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Masculino , Camundongos , Fosforilação , Serina/metabolismo , Sinaptossomos/efeitos dos fármacos , Sinaptossomos/metabolismo , Córtex Visual/efeitos dos fármacos , Córtex Visual/metabolismo
9.
Drugs ; 79(10): 1037-1051, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31161537

RESUMO

Kinase activating missense mutations in leucine-rich repeat kinase 2 (LRRK2) are pathogenically linked to neurodegenerative Parkinson's disease (PD). Over the past decade, substantial effort has been devoted to the development of potent and selective small molecule inhibitors of LRRK2, as well as their preclinical testing across different Parkinson's disease models. This review outlines the genetic and biochemical evidence that pathogenic missense mutations increase LRRK2 kinase activity, which in turn provides the rationale for the development of small molecule inhibitors as potential PD therapeutics. An overview of progress in the development of LRRK2 inhibitors is provided, which in particular indicates that highly selective and potent compounds capable of clinical utility have been developed. We outline evidence from rodent- and human-induced pluripotent stem cell models that support a pathogenic role for LRRK2 kinase activity, and review the substantial experiments aimed at evaluating the safety of LRRK2 inhibitors. We address challenges still to overcome in the translational therapeutic pipeline, including biomarker development and clinical trial strategies, and finally outline the potential utility of LRRK2 inhibitors for other genetic forms of PD and ultimately sporadic PD. Collective evidence supports the ongoing clinical translation of LRRK2 inhibitors as a therapeutic intervention for PD is greatly needed.


Assuntos
Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/antagonistas & inibidores , Doença de Parkinson/tratamento farmacológico , Inibidores de Proteínas Quinases/química , Animais , Biomarcadores , Linhagem Celular , Regulação da Expressão Gênica , Humanos , Células-Tronco Pluripotentes Induzidas , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Terapia de Alvo Molecular/métodos , Mutação , Inibidores de Proteínas Quinases/metabolismo , Roedores
10.
J Neurol ; 266(7): 1796-1800, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31041581

RESUMO

BACKGROUND: Recently, rs2421947 in DNM3 (dynamin 3) was reported as a genetic modifier of age at onset (AAO) of LRRK2 G2019S-related Parkinson's disease (PD) in a genome-wide association study in Arab-Berber population. Rs356219 in SNCA (α-synuclein) was also reported to regulate the AAO of LRRK2-related PD in European populations, and GAK (Cyclin G-associated kinase) rs1524282 was reported to be associated with an increased PD risk with an interaction with SNCA rs356219. G2019S variant is rare in Asian populations, whereas two other Asian-specific LRRK2 variants, G2385R and R1628P, are more frequent with a twofold increased risk of PD. METHODS: In this study, we investigated whether rs2421947, rs356219 and rs1524282 modified AAO in LRRK2-related PD patients in Han Chinese population. We screened LRRK2 G2385R and R1628P variants in 732 PD patients and 1992 healthy controls, and genotyped DNM3 rs2421947, SNCA rs356219 and GAK rs1524282 among the LRRK2 carriers. RESULTS: The SNCA rs356219-G allele was found to increase the risk of PD in LRRK2 carriers (OR 1.50, 95%CI 1.08-2.01, P = 0.016), and the AAO of AG + GG genotypes was 4 years earlier than AA genotype (P = 0.006). Nonetheless, no similar association was found in DNM3 rs2421947 and GAK rs1524282. CONCLUSIONS: Our results show that SNCA but not DNM3 or GAK is associated with AAO of LRRK2-PD patients in Chinese population.


Assuntos
Dinamina III/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Doença de Parkinson/epidemiologia , Doença de Parkinson/genética , Proteínas Serina-Treonina Quinases/genética , alfa-Sinucleína/genética , Idade de Início , Idoso , China/epidemiologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Doença de Parkinson/diagnóstico , Vigilância da População
11.
Folia Neuropathol ; 57(1): 1-5, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31038182

RESUMO

Pathogenesis and the development of Alzheimer's disease (AD) are subject to several environmental and genetic factors. This study was aimed to estimate the frequency of mutations in leucine-rich repeat kinase 2 (LRRK2) gene to examine the association between these mutations and risk of AD. For finding the articles, four databases including PubMed, Web of Science, Scopus, and Cochrane Library were checked up to August 2018. An analysis was done by RevMan 5.3 using crude odds ratio (OR) and 95% confidence intervals (CIs) to determine the association between LRRK2 polymorphisms and the risk of AD. Of 359 articles identified in the databases, 13 studies were included and analysed in the meta-analysis. There was no significant risk of AD related to five LRRK2 polymorphisms (rs33949390, rs34778348, rs7308720, rs34637584, and rs35870237). The results showed that LRRK2 variants (p.R1628P, p.G2385R, p.N551K, p.G2019S, and p.I2020T) were not associated with the risk of AD and were not a common cause of AD in populations. Nevertheless, p.R1628P can be examined in patients with AD in other populations in the future studies.


Assuntos
Doença de Alzheimer/genética , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Idoso , Feminino , Predisposição Genética para Doença/genética , Humanos , Masculino , Polimorfismo de Nucleotídeo Único
12.
Neurosci Lett ; 706: 140-145, 2019 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-31085292

RESUMO

INTRODUCTION: Parkinson's disease is the second most common neurodegenerative disease. Lifestyle, environmental effects and several genetic factors have been proposed to contribute to its development. Though the majority of PD cases do not have a family history of disease, genetic alterations are proposed to be present in 60 percent of the more common sporadic cases. OBJECTIVE: The aim of this study is to evaluate the frequency of PD related specific risk variants of LRRK2, MAPT, SNCA and PARK10 genes in the Hungarian population. Out of the ten investigated polymorphisms three are proposed to have protective effect and seven are putative risk factors. METHODS: For genotyping, TaqMan allelic discrimination and restriction fragment length polymorphism method was used. LRRK2 mutations were investigated among 124 sporadic PD patients and 128 healthy controls. MAPT and SNCA variant frequencies were evaluated in a group of 123 patients and 122 controls, while PARK10 variant was studied in groups of 121 patients and 113 controls. RESULTS: No significant difference could be detected in the frequencies of the investigated MAPT and PARK10 variants between the studied Hungarian PD cases and controls. The minor allele of the risk factor S1647T LRRK2 variant was found to be more frequent among healthy male individuals compared to patients. Moreover, in the frequency of one of the investigated SNCA variant a significant intergroup difference was detected. The minor allele (A) of rs356186 is proposed to be protective against developing the disease. In accord with data obtained in other populations, the AA genotype was significantly more frequent among Hungarian healthy controls compared to patients. Similarly, a significant difference in genotype distribution was also found in comparison of patients with late onset disease to healthy controls, which was due to the higher frequency of AG genotype among patients. CONCLUSION: The frequencies of different gene variants show great differences in populations. Assessment of the frequency of variants of PD related genes variants is important in order to uncover the pathomechanisms underlying the disease, and to identify potential therapeutic targets. This is the first comprehensive study focusing on these genetic variants in the population of East-Central European region. Our results extend the knowledge on the world wide occurrence of these polymorphisms by demonstrating the occurrence of specific alleles and absence of others in Hungarian PD patients.


Assuntos
Predisposição Genética para Doença , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Doença de Parkinson/genética , alfa-Sinucleína/genética , Proteínas tau/genética , Adulto , Idoso , Alelos , Feminino , Frequência do Gene , Genótipo , Humanos , Hungria , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único
13.
Neurobiol Dis ; 127: 512-526, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30954703

RESUMO

BACKGROUND: Mutations in LRRK2 are the most common cause of autosomal dominant Parkinson's disease, and the relevance of LRRK2 to the sporadic form of the disease is becoming ever more apparent. It is therefore essential that studies are conducted to improve our understanding of the cellular role of this protein. Here we use multiple models and techniques to identify the pathways through which LRRK2 mutations may lead to the development of Parkinson's disease. METHODS: A novel integrated transcriptomics and proteomics approach was used to identify pathways that were significantly altered in iPSC-derived dopaminergic neurons carrying the LRRK2-G2019S mutation. Western blotting, immunostaining and functional assays including FM1-43 analysis of synaptic vesicle endocytosis were performed to confirm these findings in iPSC-derived dopaminergic neuronal cultures carrying either the LRRK2-G2019S or the LRRK2-R1441C mutation, and LRRK2 BAC transgenic rats, and post-mortem human brain tissue from LRRK2-G2019S patients. RESULTS: Our integrated -omics analysis revealed highly significant dysregulation of the endocytic pathway in iPSC-derived dopaminergic neurons carrying the LRRK2-G2019S mutation. Western blot analysis confirmed that key endocytic proteins including endophilin I-III, dynamin-1, and various RAB proteins were downregulated in these cultures and in cultures carrying the LRRK2-R1441C mutation, compared with controls. We also found changes in expression of 25 RAB proteins. Changes in endocytic protein expression led to a functional impairment in clathrin-mediated synaptic vesicle endocytosis. Further to this, we found that the endocytic pathway was also perturbed in striatal tissue of aged LRRK2 BAC transgenic rats overexpressing either the LRRK2 wildtype, LRRK2-R1441C or LRRK2-G2019S transgenes. Finally, we found that clathrin heavy chain and endophilin I-III levels are increased in human post-mortem tissue from LRRK2-G2019S patients compared with controls. CONCLUSIONS: Our study demonstrates extensive alterations across the endocytic pathway associated with LRRK2 mutations in iPSC-derived dopaminergic neurons and BAC transgenic rats, as well as in post-mortem brain tissue from PD patients carrying a LRRK2 mutation. In particular, we find evidence of disrupted clathrin-mediated endocytosis and suggest that LRRK2-mediated PD pathogenesis may arise through dysregulation of this process.


Assuntos
Neurônios Dopaminérgicos/metabolismo , Endocitose/genética , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Mutação , Animais , Perfilação da Expressão Gênica , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Proteômica , Ratos , Ratos Transgênicos , Vesículas Sinápticas/genética
14.
Neuroscience ; 409: 169-179, 2019 06 15.
Artigo em Inglês | MEDLINE | ID: mdl-31029729

RESUMO

Parkinson's disease (PD) is the most common neurodegenerative movement disorder and is characterized by the loss of neurons in the substantia nigra that project to the striatum and release dopamine (DA), which is required for normal movement. Common non-motor symptoms likely involve abnormalities with other neurotransmitters, such as serotonin, norepinephrine, acetylcholine, glycine, glutamate and gamma-aminobutyric acid (GABA). As part of a broad effort to provide better PD research tools, the Michael J. Fox Foundation for Parkinson's Research funded the generation and characterization of knockout (KO) rats for genes with PD-linked mutations, including PINK1, Parkin, DJ-1 and LRRK2. Here we extend the phenotypic characterization of these lines of KO rats to include in vivo microdialysis to measure both basal and potassium-induced release of the above neurotransmitters and their metabolites in the striatum of awake and freely moving rats at ages 4, 8 and 12 months compared to wild-type (WT) rats. We found age-dependent abnormalities in basal DA, glutamate and acetylcholine in PINK1 KO rats and age-dependent abnormalities in basal DA metabolites in Parkin and LRRK2 KO rats. Parkin KO rats had increased glycine release while DJ-1 KO rats had decreased glutamate release and increased acetylcholine release compared to WT rats. All lines except DJ-1 KO rats showed age-dependent changes in release of one or more neurotransmitters. Our data suggest these rats may be useful for studies of PD-related synaptic dysfunction and neurotransmitter dynamics as well as studies of the normal and pathogenic functions of these genes with PD-linked mutations.


Assuntos
Acetilcolina/metabolismo , Encéfalo/metabolismo , Dopamina/metabolismo , Ácido Glutâmico/metabolismo , Doença de Parkinson/metabolismo , Animais , Neurônios Dopaminérgicos/metabolismo , Técnicas de Inativação de Genes , Glicina/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Masculino , Doença de Parkinson/genética , Proteína Desglicase DJ-1/genética , Proteína Desglicase DJ-1/metabolismo , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Ratos , Serotonina/metabolismo , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/metabolismo , Ácido gama-Aminobutírico/metabolismo
15.
Biochem Soc Trans ; 47(2): 651-661, 2019 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-30837320

RESUMO

The past two decades in research has revealed the importance of leucine-rich repeat kinase 2 (LRRK2) in both monogenic and sporadic forms of Parkinson's disease (PD). In families, mutations in LRRK2 can cause PD with age-dependent but variable penetrance and genome-wide association studies have found variants of the gene that are risk factors for sporadic PD. Functional studies have suggested that the common mechanism that links all disease-associated variants is that they increase LRRK2 kinase activity, albeit in different ways. Here, we will discuss the roles of LRRK2 in areas of inflammation and vesicular trafficking in the context of monogenic and sporadic PD. We will also provide a hypothetical model that links inflammation and vesicular trafficking together in an effort to outline how these pathways might interact and eventually lead to neuronal cell death. We will also highlight the translational potential of LRRK2-specific kinase inhibitors for the treatment of PD.


Assuntos
Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Doença de Parkinson/enzimologia , Doença de Parkinson/genética , Proteínas Serina-Treonina Quinases/genética , Animais , Estudo de Associação Genômica Ampla/métodos , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Mutação/genética
16.
Biochem Soc Trans ; 47(2): 663-670, 2019 04 30.
Artigo em Inglês | MEDLINE | ID: mdl-30837321

RESUMO

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene are a frequent genetic cause of late-onset Parkinson's disease (PD) and a target for therapeutic approaches. LRRK2 protein can influence vesicle trafficking events in the cytosol, with action both in endosomal and lysosomal pathways in different types of cells. A subset of late endosomes harbor intraluminal vesicles that can be secreted into the extracellular milieu. These extracellular vesicles, called exosomes, package LRRK2 protein for transport outside the cell into easily accessed biofluids. Both the cytoplasmic complement of LRRK2 as well as the exosome-associated fraction of protein appears regulated in part by interactions with 14-3-3 proteins. LRRK2 inside exosomes have disease-linked post-translational modifications and are relatively stable compared with unprotected proteins in the extracellular space or disrupted cytosolic compartments. Herein, we review the biology of exosome-associated LRRK2 and the potential for utility in diagnosis, prognosis, and theragnosis in PD and other LRRK2-linked diseases.


Assuntos
Biomarcadores/metabolismo , Exossomos/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Animais , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Doença de Parkinson/genética , Doença de Parkinson/metabolismo
17.
PLoS One ; 14(3): e0213527, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30861027

RESUMO

Mitochondrial function has been implicated and studied in numerous complex age-related diseases. Understanding the potential role of mitochondria in disease pathophysiology is of importance due to the rise in prevalence of complex age-related diseases, such as type 2 diabetes (T2D) and Alzheimer's disease (AD). These two diseases specifically share common pathophysiological characteristics which potentially point to a common root cause or factors for disease exacerbation. Studying the shared phenomena in Mexican Americans is of particular importance due to the disproportionate prevalence of both T2D and AD in this population. Here, we assessed the potential role of mitochondria in T2D and cognitive impairment (CI) in a Mexican American cohort by analyzing blood-based indices of mitochondrial DNA copy number (mtDNACN) and cell-free mitochondrial DNA (CFmtDNA). These mitochondrial metrics were also analyzed for correlation with relevant neuropsychological variables and physiological data collected as indicators of disease and/or disease progression. We found mtDNACN to be significantly decreased in individuals with CI, while CFmtDNA was significantly elevated in T2D; further, CFmtDNA elevation was significantly exacerbated in individuals with both diseases. MtDNACN was found to negatively correlate with age and fatty acid binding protein concentration, while positively correlating with CFmtDNA as well as CERAD total recall score. Candidate gene SNP-set analysis was performed on genes previously implicated in maintenance and control of mitochondrial dynamics to determine if nuclear variants may account for variability in mtDNACN. The results point to a single significant locus, in the LRRK2/MUC19 region, encoding leucine rich repeat kinase 2 and mucin 19. This locus has been previously implicated in Parkinson's disease, among others; rs7302859 was the driver SNP. These combined findings further indicate that mitochondrial dysfunction (as assessed by proxy via mtDNACN) is intimately linked to both T2D and CI phenotypes as well as aging.


Assuntos
Ácidos Nucleicos Livres , Disfunção Cognitiva , DNA Mitocondrial , Diabetes Mellitus Tipo 2 , Americanos Mexicanos , Idoso , Doença de Alzheimer/sangue , Doença de Alzheimer/etnologia , Doença de Alzheimer/genética , Ácidos Nucleicos Livres/sangue , Ácidos Nucleicos Livres/genética , Disfunção Cognitiva/sangue , Disfunção Cognitiva/etnologia , Disfunção Cognitiva/genética , DNA Mitocondrial/sangue , DNA Mitocondrial/genética , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/etnologia , Diabetes Mellitus Tipo 2/genética , Feminino , Loci Gênicos , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Masculino , Pessoa de Meia-Idade , Mucinas/genética , Polimorfismo de Nucleotídeo Único
18.
Int J Mol Sci ; 20(6)2019 Mar 26.
Artigo em Inglês | MEDLINE | ID: mdl-30917570

RESUMO

Alzheimer's disease (AD), Parkinson's disease (PD), frontotemporal dementia (FTD), amyotrophic lateral sclerosis (ALS), Huntington's disease (HD), and prion diseases have a certain degree of clinical, pathological, and molecular overlapping. Previous studies revealed that many causative mutations in AD, PD, and FTD/ALS genes could be found in clinical familial and sporadic AD. To further elucidate the missing heritability in early-onset Alzheimer's disease (EOAD), we genetically characterized a Thai EOAD cohort by Next-Generation Sequencing (NGS) with a high depth of coverage, capturing variants in 50 previously recognized AD and other related disorders' genes. A novel mutation, APP p.V604M, and the known causative variant, PSEN1 p.E184G, were found in two of the familiar cases. Remarkably, among 61 missense variants were additionally discovered from 21 genes out of 50 genes, six potential mutations including MAPT P513A, LRRK2 p.R1628P, TREM2 p.L211P, and CSF1R (p.P54Q and pL536V) may be considered to be probably/possibly pathogenic and risk factors for other dementia leading to neuronal degeneration. All allele frequencies of the identified missense mutations were compared to 622 control individuals. Our study provides initial evidence that AD and other neurodegenerative diseases may represent shades of the same disease spectrum, and consideration should be given to offer exactly embracing genetic testing to patients diagnosed with EOAD. Our results need to be further confirmed with a larger cohort from this area.


Assuntos
Doença de Alzheimer/genética , Adulto , Idade de Início , Doença de Alzheimer/patologia , Precursor de Proteína beta-Amiloide/genética , Feminino , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Masculino , Glicoproteínas de Membrana/genética , Pessoa de Meia-Idade , Mutação de Sentido Incorreto , Presenilina-1/genética , Receptores de Fator Estimulador das Colônias de Granulócitos e Macrófagos/genética , Receptores Imunológicos/genética , Proteínas tau/genética
19.
J Biol Chem ; 294(15): 5907-5913, 2019 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-30796162

RESUMO

Mutation in leucine-rich repeat kinase 2 (LRRK2) is a common cause of familial Parkinson's disease (PD). Recently, we showed that a disease-associated mutation R1441H rendered the GTPase domain of LRRK2 catalytically less active and thereby trapping it in a more persistently "on" conformation. However, the mechanism involved and characteristics of this on conformation remained unknown. Here, we report that the Ras of complex protein (ROC) domain of LRRK2 exists in a dynamic dimer-monomer equilibrium that is oppositely driven by GDP and GTP binding. We also observed that the PD-associated mutations at residue 1441 impair this dynamic and shift the conformation of ROC to a GTP-bound-like monomeric conformation. Moreover, we show that residue Arg-1441 is critical for regulating the conformational dynamics of ROC. In summary, our results reveal that the PD-associated substitutions at Arg-1441 of LRRK2 alter monomer-dimer dynamics and thereby trap its GTPase domain in an activated state.


Assuntos
Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina , Mutação de Sentido Incorreto , Doença de Parkinson , Multimerização Proteica , Substituição de Aminoácidos , Guanosina Difosfato/química , Guanosina Difosfato/genética , Guanosina Trifosfato/química , Guanosina Trifosfato/genética , Células HEK293 , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/química , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Doença de Parkinson/enzimologia , Doença de Parkinson/genética , Domínios Proteicos
20.
J Biol Chem ; 294(13): 4738-4758, 2019 03 29.
Artigo em Inglês | MEDLINE | ID: mdl-30709905

RESUMO

Mutations in the gene encoding for leucine-rich repeat kinase 2 (LRRK2) are a common cause of hereditary Parkinson's disease. LRRK2 regulates various intracellular vesicular trafficking pathways, including endolysosomal degradative events such as epidermal growth factor receptor (EGFR) degradation. Recent studies have revealed that a subset of RAB proteins involved in secretory and endocytic recycling are LRRK2 kinase substrates in vivo However, the effects of LRRK2-mediated phosphorylation of these substrates on membrane trafficking remain unknown. Here, using an array of immunofluorescence and pulldown assays, we report that expression of active or phosphodeficient RAB8A variants rescues the G2019S LRRK2-mediated effects on endolysosomal membrane trafficking. Similarly, up-regulation of the RAB11-Rabin8-RAB8A cascade, which activates RAB8A, also reverted these trafficking deficits. Loss of RAB8A mimicked the effects of G2019S LRRK2 on endolysosomal trafficking and decreased RAB7A activity. Expression of pathogenic G2019S LRRK2 or loss of RAB8A interfered with EGFR degradation by causing its accumulation in a RAB4-positive endocytic compartment, which was accompanied by a deficit in EGFR recycling and was rescued upon expression of active RAB7A. Dominant-negative RAB7A expression resulted in similar deficits in EGF degradation, accumulation in a RAB4 compartment, and deficits in EGFR recycling, which were all rescued upon expression of active RAB8A. Taken together, these findings suggest that, by impairing RAB8A function, pathogenic G2019S LRRK2 deregulates endolysosomal transport and endocytic recycling events.


Assuntos
Endossomos/metabolismo , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/metabolismo , Lisossomos/metabolismo , Mutação de Sentido Incorreto , Proteínas rab de Ligação ao GTP/metabolismo , Substituição de Aminoácidos , Endossomos/genética , Receptores ErbB/genética , Receptores ErbB/metabolismo , Células HEK293 , Células HeLa , Humanos , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Lisossomos/genética , Transporte Proteico/genética , Proteínas Serina-Treonina Quinases/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteólise , Proteínas rab de Ligação ao GTP/genética
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