Lecture 2 Parkinson s Disease. David Saffen, Ph.D. Professor/Principal Investigator Dept. Cellular and Genetic Medicine

Transcription

1 Lecture 2 Parkinson s Disease David Saffen, Ph.D. Professor/Principal Investigator Dept. Cellular and Genetic Medicine saffen@fudan.edu.cn

2 Parkinson s disease (PD) James Parkinson ( ) English apothecary surgeon Published essay on Shaking Palsy in 1817 Parkinson s home (London)

3 Characteristics of PD (1) A chronic and progressive neurodegenerative disorder of the brain that affects movement; emotional and cognitive deficits emerge in later stages Usually develops in individuals over 50 (mean = age 60); Rare familial forms are known, but most cases are sporadic. Environmentally induced forms have also been described. PD is the most common motor neurodegenerative disorder and is second most common neurodegenerative disorder after Alzheimer s disease (AD). Prevalence: 1% at age 60; 4-5% at 85; There are currently an estimated 500,000 individuals with PD in the US

4 Characteristics of PD (2) Cardinal symptoms ( parkinsonism ): Resting Tremor in hands, arms, legs, jaw and face Stiffness of limbs and truck; cogwheel rigidity Bradykinesia/freezing: slowness of movement; difficulties in initiating or performing sequential or simultaneous movements Postural instability/stooped posture: impaired balance and coordination; festination (shuffling steps and forwardflexed posture) Additional symptoms: Slow speech, loss of expressiveness in face; diminished sense of smell, speech and swallowing disorders, sleep disorders, rest-less leg syndrome, fatigue, impotence, visual problems, constipation; cognitive deficits, depression, apathy, anxiety, dementia Symptoms produced by medications: dyskinesia, impulse control, hallucinations, delusions, delirium.

5 Characteristics of PD (3) Complete, or nearly complete loss of dopamine-producing neurons in the substantia nigra pars compacta; deposits of aggregated protein (Lewy bodies), containing denatured alpha-synuclein and other proteins, in affected brain areas Recent research as revealed that neuronal loss is more wide-spread than previously suspected, including: dopamine-producing neurons in the ventral tegmental area (VTA), which supply dopamine to the nucleus accumbens (NAc) and prefrontal cortex (PFC); serotonin-producing neurons in the raphe nuclei; norepinephrine-producing neurons in the locus ceruleus and acetylcholine-producing neurons in the nucleus basalis of Meynert; also: neurons in the cortex, striatum, hypothalamus, olfactory bulb, dorsal motor nucleus of the vagus and intestine. There is a growing consensus that PD is heterogeneous, with subtypes distinguished by age of onset, clinical symptoms and progression rates.

6 PD vs parkinsonism The four cardinal signs of parkinsonism (tremor, bradykinesia, rigidity and postural instability) are also observed in diseases other than PD. For example: AIDS, corticobasal degeneration, diffuse Lewy body disease, encephalitis lethargica, multiple system atrophy, progressive supranuclear palsy, and genetic disorders, including Wilson s disease, rapid onset dystonia parkinsonism (DYT12), x-linked dystonia parkinsonism (DYT3) and autosomal recessive juvenile parkinsonism (ARJP) Parkinsonism can also be a symptom resulting from exposure to certain toxins, including carbon monoxide, manganese, paraquat, hexane, toluene and MPTP (a byproduct of synthesis of the opioid-like drug MPPP.) Parkinsonism can also be caused by medications, including: the antipsychotic drugs chlorpromazine, haloperidol and ziprasidone. All of these drugs share the common property of being inhibitors of D 2 dopamine receptors

7 Faces of PD and parkinsonism Owen Chamberlain Salvadore Dali Mohamed Ali Deng Xiaoping Yasir Arafat

8 Brain pathology in PD (1) Dopamineproducing neurons are intrinsically dark brown due to high intracellular levels of neuromelanin, a derivative of the amino acids L-DOPA and dopamine

9 Brain pathology in PD (2)

10 Brain pathology in PD (3) Lewy bodies Lewy body (brown) immunostained for alpha-synuclein in a neuron in the substantia nigra; Lewy bodies are also usually immunopositive for ubiquitin, a low-molecular peptide that is covalently linked to proteins targeted for degradation in proteosomes, and for the ubiquitin binding protein p62.

11 Brain pathology in PD (4) Lewy neurites Ubiquitin immunostained Lewy neurites also positively stain for alpha-synuclein (Prof. Mel Feany, Harvard University)

12 Synucleinopathies Dementia with Lewy bodies (DLB) Diffuse Lewy body disease (DLBD) Lewy body disease (LBD) Multiple system atrophy (MSA)

13 Dopamine neuromelanin (brown polymer) MAO = monoamine oxidase COMT = catechol-o-methyl transferase HVA = homovanillic acid

14 The dopamine nigrostriatal tract Nigrostriatal tract: SNpc to striatum (caudate & putamen) Mesolimbic tract: ventral tegmental area (VTA) to nucleus accumbens & prefrontal cortex Tuberoinfundibular tract: arcuate nucleus to base of pituitary stalk (infundibulum)

15 Dopamine receptors: G protein-coupled receptor (GPCR) family members Subtype Agonists Antagonists Coupling SCH23390 D 1 SKF82958 SKF81297 SKF83566 haloperidol G s D 2 bromocriptine raclopride haloperidol G i/o D 3 quinpirole raclopride G i/o D 4 clozapine G i/o D 5 SKF38393 SCH23390 G s

16 Distribution of dopamine receptors in striatum PET scan showing distribution of binding of the D 2 receptor antagonist [ 11 C] raclopride MRI showing position of putamen (different brain)

17 Changes in striatal dopamine innervation in PD PET scans showing the distribution of dopamine transporter antagonist 11 C-methylphenidate in a healthy individual (left) and in subjects with a LRRK2 mutation (middle and right). Note: the dopamine transporter (DAT) is located in the axonal terminals of dopamine neurons, where it functions to recover dopamine released into the synaptic cleft. Methylphenidate (Ritalin) is a stimulant used in the treatment of attention-deficit/hyperactivity disorder (ADHD) methylphenidate Agarwal PA et al, Movement Disorders 28, 2013

18 Hypothesis: elimination of dopaminergic inputs from the substantia nigra pars compacta to the medium spiny projection neurons in the caudate and putamen weakens the direct pathway and strengthens the indirect pathway. The resulting imbalance produces excessive output from inhibitory GPi projection neurons, resulting in inhibition of voluntary movement Kandel ER, Schwartz JH and Jessel TM, 2000

19 Surgical ablation of the subthalamic nucleus (STN) or globus pallidus, internal segment (GPi) reduces inhibitory output from the basal ganglia, facilitating voluntary movement

20 Epidemiology of PD Rural living, well-water consumption, mining, welding, exposure to pesticides identified as risk factors Male/female prevalence = 1.5 Early twin and epidemiological studies did not indicate a genetic cause for PD More recently, genetic linkage and association studies have demonstrated a strong genetic contribution to PD. About 5-10% of PD cases are inherited, and 21 PD susceptibility chromosomal loci (PARK1-21) and at least 10 specific PD susceptibility genes for familial PD have now been identified. The remaining ~90-95% of PD cases are idiopathic, (i.e., of unknown cause), but PD susceptibility genes have also been implicated in a small percentage of sporadic cases.

21 Suspected environmental causes of PD and parkinsonism Herbicides and insecticides e.g., paraquat, rotenone Manganese Maneb (manganese ethythene-bis-dithiocarbamate) ferro-manganese Metals mercury, copper, lead Organic solvents toluene, N-hexane, carbon disulfide, trichloroethylene Viral infections: post-encephalytic parkinsonism (linked to influenza epidemic of 1918) Physical trauma (brain): e.g., Mohamed Ali (boxer)

22 Environmentally induced PD: MPTP Cyperquat = MPP+ Paraquat MPTP = 1-methyl-4-phenyl- 1,2,3,6-tetrahydropyridine MPP+ = 1-methyl- 4-phenylpyridinium = potent inhibitor of mitochondrial complex I (toxic!) Note: MPTP also produces excessive activation of microglial cells leading to the release of toxic cytokines desmethylprodine = 1-methyl-4-phenylpropionoxypiperidine (MPPP) = an analog of pethidine (Meperidine = Demerol), an opioid analgesic

23 Environmental factors associated with protection from PD Cigarette smoke Caffeinated coffee Non-steroidal anti-inflammatory drugs (NSAIDs)

24 Prevalence of PD differs among ethnic groups United States and Western Europe: US: per 100,000 (~ million persons) Germany: 183 per 100,000 Asia: Japan: per 100,000 China: 130 per 100,000 (~1.7 million persons) Africa: Nigeria: 67 per 100,000 Ethiopia: 7 per 100,000 (due to low life-expectancy?) Special populations: Amish (US): 970 per 100,000!

25 Genetics of Parkinson s disease Familial PD: Monogenetic disorders with dominant or recessive Mendelian Patterns of inheritance Family-based genetic linkage studies have identified 21 chromosomal loci that associate with PD*: PARK1-PARK21. Candidate PD liability genes have been identified in 16 of these loci. (*Note: some of these loci may be false-positives, or contain gene that modify but are insufficient to cause PD when mutated. ) Together familial PD genes account for 5-10% of diagnosed PD cases Idiopathic PD: Sporadic, complex disorders with both genetic and environmental causes.

26 Six major genes implicated in familial PD: Late onset, autosomal dominant inheritance PD: SNCA (PARK1/4*; alpha-synuclein) LRRK2 (Park8; Leucine-rich repeat kinase 2) VPS35 (Park17; vacuolar protein sorting associated protein 35) Early onset, autosomal recessive inheritance PD**: PRKN (PARK2; parkin, : E3 ubiquitin ligase) PINK1 (PARK6; PTEN-induced putative kinase 2) DJ-1 (PARK7; molecular chaperone/anti-oxidant) *PARK1 refers to point mutations in SNCA and PARK4 to SNCA duplications or triplications. **PD is observed in individuals homozygous for the same mutation or individuals harboring different pathogenic mutations in the same gene (termed compound heterozygotes).

27 Common genetic variants in four genes have been consistently replicated in genetic association studies for idiopathic PD SNCA (alpha-synuclein) Rep1: a polymorphic microsatellite repeat located ~10 kb upstream; Rep1(261) associated with increased a-synuclein expression = risk allele; Rep1(259) associate with decreased a-synuclein expression = protective allele. (odds ratio: ) LRRK2 (leucine-rich repeat kinase 2) G2019S accounts for 4-5% of familial PD cases in European populations, but 30-40% of familial PD Arab and Ashkenazi Jewish populations! Mutation is very rare among Asians. G2385R and R1628P = most frequent common risk factors for idiopathic PD in Asian populations. (odds ratios 2.2 and 1.8, respectively) MAPT (microtubule-associated protein Tau) H1 haplotype = risk haplotype (odds ratio: 1.5) GBA (glucocerebrosidase beta) Heterozygous carriers of GBA mutations have ~5-fold increased risk for PD with early onset and 6-fold increased risk of developing dementia. Together, GBA mutations are the most common risk factor for idiopathic PD in non-asian populations.

28 Cellular systems implicated in the etiology of PD (and related candidate PD liability genes) Cell-autonomous (dopamine neurons in SNpc) Protein aggregation (SNCA, DJ-1, ATP13A2, MAPT) Increased Ca2+ influx Mitochondria dysfunction (Parkin, PINK1, DJ-1) Autophagy/Mitophagy (Parkin, PINK1) Proteosomes (Parkin, UCHL1) Endosomes/Lysosomes (LRRK2, VPS35, GBA, ATP13A2) Non-cell-autonomous Excitotoxicity Inflammation, including production of apoptosispromoting cytokines by microglia (possibly potentiated by LRRK2 mutants) and fas-ligand by invasive CD4 T-lymphocytes Prion-like spread of alpha-synuclein aggregation (?)

29 a-synuclein A predominantly 140 aa protein that is highly expressed in neurons, comprising ~1% of cytosolic proteins. Lower levels of 126 and 112 aa isoforms, produced by alternative mrna splicing are also expressed at lower levels Localized in mitochondria, the cytosol and nucleus, but at highest levels in pre-synaptic terminals, where it is proposed to play a role in the clustering of neurotransmitter-filled synaptic vesicles. An intrinsically disordered protein that lacks stable three-dimensional structure. Aggregates to form insoluble fibrils under pathogenic conditions. SNCA is a dosage-sensitive gene: duplication or triplication is sufficient to cause familial, autosomal PD; Genetic regulatory variants that increase SNCA expression are PD risk factors. Rare missense mutations, including A53T, A30P, E46K, H50Q and G51D, cause familial PD.

30 Postulated events in a-synuclein toxicity PLK2 = Polo-like kinase 2 Recent evidence suggests that damage to the ubiquitinproteosome system (UPS) and the autophagy-lysosomal pathway are central to the role of a-synuclein in the etiology of PD CMA = chaperonemediated autophagy

31 Ubiquitin-proteosome system (UPS) lysine E1 = ubiquitin-activating enzyme E2 = ubiquitin-conjugating enzyme E3 = ubiquitin ligase Ubiquitin (76 aa), containing seven lysine residues (yellow) Proteosome

32 Autophagy-lysosome pathways (macro-, micro- and chaperone-mediated autophagy) Lynch-day, et al, CSH Perspectives in Medicine, 2012

33 a-synuclein-mediated inhibition of ubiquitin-proteosome system (USP) and autophagy propels neuronal degeneration in PD CMA = chaperonemediated autophagy Li J et al. Oncotarget, 2015

34 A proposed pathogenic cycle: lysosome dysfunction stimulates the production of aggregated a-synuclein and aggregated a-synuclein promotes lysozyme dysfunction GCase = glucocerebrosidase Xilouri M et al, Movement Disorders, 2016

35 Roles of additional PD-associated proteins in macro-autophagy A53T, E46K = SNCA mutations; G2019S = LRRK2 mutation Li J et al. Oncotarget, 2015

36 Pathogenic mutations in LRRK2 that increase kinase activity may indirectly impair autophagy by excessively phosphorylating Rab proteins, which broadly function in vesicle transport and trafficking. Steger M et al, elife, 2016

37 Drug therapy for PD (1) Symptomatic relief of movement deficits (tremor, rigidity, akinesia) is provided by systemic administration of L-DOPA, a dopamine precursor = levodopa L-DOPA is less successful in improving deficits in gait, balance, swallowing, speech, autonomic functions and cognition. Also, L-DOPA looses efficacy and causes movement and behavioral problems in many patents after long-term treatment.

38 Drug therapy for PD (2) Peripheral dopadecarboxylase inhibitors: carbidopa benserazide selegiline rasagiline entacapone HVA = homovanillic acid Dopamine receptor : agonists bromocriptine, ropinirole, pramipexole piribedil, apomorphine, lisuride, rotigotine MAO = monoamine oxidase COMT = catechol-o-methyl transferase

39 Ablative therapies for PD Lesions of the subthalamic nucleus (STN) or globus pallidus, internal segment (GPi) [to increase voluntary movement] Lesions of specific nuclei in the thalamus [to reduce tremor or dystonia in patients treated with L-DOPA] SNc = Substantia nigra, par compacta GPe = globus pallidus, external segment PPN = pedunculopontine nucleus Green = excitatory pathways Red = inhibitory pathways DeLong MR & Wichmann T, 2007

40 Stimulation of neurons in the STN or GPi can suppress involuntary movements (e.g., chorea, dystonia) that arise as side-effects of L-DOPA therapy. DBS of the cingulate gyrus may also be effective in treating depression in PD patients. Deep brain stimulation (DBS) Future treatments with trans-cranial magnetic stimulation (TMS)? Outlooks: Parkinson s Disease, Nature 466, S17, 2010

41 Cell replacement therapies Dopamine-producing cells from the adrenal cortex Embryonic stem cells Induced pluripotent stem (ips) cells Technical Problems: - availability of cells - possible risk of brain tumors - inability to control the amount of dopamine produced - problems with long-term survival of transplanted cells e.g., Lewy bodies observed in transplanted cells at autopsy (consistent with prion-like behavior of alpha-synuclein)

42 PD as an amyloid disease [SNCA] [MAPT] Schulman JM & De Jager PL, 2010

43 References (1) Hirsch EC et al, Pathogenesis of Parkinson s Disease, Movement Disorders 28, 2013 Kalinderi K, Bostantjjopoulou S and Fidani L, The genetic background of Parkinson s disease: current progress and future prospects, Acta Neurology Scandinavia, 2016 Singleton AB et al, The genetics of Parkinson s Disease: Progress and therapeutic implications, Movement Disorders 28, 2013 Houlden H and Singleton AB, The genetics and neuropathology of Parkinson s disease, ACT Neuropathology 124, , 2012 Obeso JA et al, Missing pieces in the Parkinson s disease puzzle, Nature Medicine 16, , 2010 Vatican Conference on Neuroprotection in Parkinson s disease, Movement Disorders 28, 1-115, 2013 Special supplement, Outlook: Parkinson s Disease, Nature 466, S3-S22, 2010 Kandel ER, Schwartz JN and Jessell TM, Principles of Neural Science, Fourth Edition, McGraw-Hill, 2000 (ISBN ) DeLong MR and Wichmann T, Circuits and Circuit Disorders of the Basal Ganglia, Archives of Neurology 64, 20-24, 2007 Lill CM et al, Comprehensive research synopsis and systematic meta-analysis in Parkinson s disease genetics: the PDGene database, PLoS Genetics 8, 2012

44 References (2) Spatola M and Wider C, Genetics of Parkinson s disease: the yield, Parkinsonism and related disorders, 20S1, S35-S38, 2014 Nuytemans K et al, Genetic etiology of Parkinson disease associated with mutations in the SNCA, PARK2, PINK1, PARK7 and LRRK2 genes: a mutation update, Human Mutation 31, , 2010 Pan PY and Yue Z, Genetic causes of Parkinson s disease and their linkes to autophagy regulation, Parkinsonism and related disorders 20S1, S154-S157, 2014 Simon-Sanchez J, et al, Genome-wide association study reveals genetic risk underlying Parkinson s disease, Nature Genetics 41, , 2009 Cookson MR and Bandmann O, Parkinson s disease: insights from pathways, Human Molecular Genetics 19, R21-R2, 2010 Schulman JM and De Jager PL, Evidence for a common pathway linking neurodegenerative diseases, Nature Genetics 41, , 2010 Matsuda N and Tanaka K, Does Impairment of the UPS or Autophagy-lysosome pathway predispose individuals to neurodegenerative disorders such as Parkinson s disease?, Journal of Alzheimer s Disease 19, 1-9, 2010 Deas E, et al, Mitophagy and Parkinson s disease, Biochimica et Biophysica Acta 1813, , 2011 Tanner CM et al, Rotenone, Paraquat and Parkinson s disease, Environmental Health Perspectives 119, , 2011

45 References (3) Firzmaurice AG et al, Aldehyde dehydrogenase variation enhances effect of pesticides associated with Parkinson s disease, Neurology 82, , 2014 Li JJ et al, Deconvoluting the complexity of autophagy and Parkinson s disease for potentinal therapeutic purpose, Oncotarget 6, , 2015 Lynch-Day MA et al, The role of autophagy in Parkinson s disease, Cold Spring Harbor Perspectives in Medicine, 1-13, 2012 Martini-Stocia H et al, The autophagy-lysosomal pathway in neurodegeneration: A TFEB perspective, Trends in Neuroscience, 1-14, 2014 Migdalska-Richards A, and Schapira HV, The relationship between glucocerebrosidase mutations and Parkinson disease, Journal of Neurochemistry 10, 2016 Sidransky MA et al, Multicenter analysis of glucocerebrosidase mutations in Parkinson s disease, The New England Jouranl of Medicine 361, , 2009 Steger M et al, Phosphoproteomics reveals that Parkinson s disease kinase LRRK2 regulates a subset of Rab GTPases, elife, 2016 Dawson TM et al, Genetic Animal Models of Parkinson s Disease, Neuron 66, , 2010 Images in clinical medicine. Cycling for the freezing of gait, New England Journal of Medicine 362, e46, 2010

46 Journal Presentations Background Siebert M et al., Glucocerebrosidase is shaking up the synucleinopathies, Brain 137, , 2014 Research Fernandes H, et al., ER stress and autophagic perturbations lead to elevated extracellular a-synuclein in GBA-N370S Parkinson s ipsc-derived dopamine neurons, Stem Cell Reports 6,

47 Internet Resources National Institute of Neurodegenerative Disease and stroke (National Institutes of Health): e.htm Parkinson s disease: hope through research (NIH): _disease.htm The Clinicians and Nurses Guide to Parkinson s Disease: National Parkinson Foundation: PDGene database: Viartis: pdmutdb:

48 Appendix Genes and cellular processes implicated in Parkinson s disease Animal models of Parkinson s disease

49 PARK loci and genes associated with familial forms of PD AD =autosomal dominant AR = autosomal recessive EOPD =early onset PD Kalinderi K et al, Acta Neurologica Scandinavia, 2016

50 SNCA (alpha-synuclein; PARK1 and PARK4) 4q21 Nuytemans et al, 2010 First PD susceptibility gene to be discovered: A53T missense mutation associated with PD in large Italian family; A30P and E46K mutations also cause familial PD. Rare SNCA duplications and triplications and rare high-expression promoter variants have also been implicated in familial and sporadic PD. Known SNCA mutations account for about 2.5% of sporadic PD cases. Common variants detected in recent PD-GWA studies. Encodes 144 aa protein; function in brain is unknown, but may play a role in neurotransmitter release and vesicle cycling in presynaptic neurons and/or vesicle trafficking within the endoplasmic reticulum/golgi network. Major component of Lewy bodies (often ubiquitinated); Lewy bodies are also observed in dementia with Lewy bodies (DLB), Diffuse Lewy body disease (DLBD), Lewy body disease (LBD), multiple system atrophy (MSA). Together with PD, these disorders have been termed alpha synucleinopathies. Lewy bodies are also observed in some patients with Gaucher s disease or Alzheimer s disease (AD) patients.

51 LRRK2 (Leucine-rich repeat kinase 2; PARK8) 12q12 Nuytemans et al, 2010.Pathogenic missense mutation G2019S accounts for 4-5% of familial PD cases in European populations, but 30-40% of familial PD Arab and Ashkenazi Jewish populations! Mutation is very rare among Asians. Additional Pathogenic mutations include: N1437H, R1441C, R1441G, Y1699C (located within the Roc-COR GTPase domains) & I2020T. The G2019S and I2020T are located within the LRRK2 kinase domain and increase LRRK2 kinase activity 2-3 fold. G2385R is relatively common in Asian populations and is associated with a ~2-fold increase in PD liability; R1628P variant also common in among Asians. LRRK2 mutations may account for 4% of sporadic PD cases worldwide. Common LRRK2 variants have been detected in recent PD-GWA studies LRRK2 protein contains multiple domains, including domains associated with: i) protein-protein interactions (ankyrin, LRR, COR,WD40), ii) GTPase (Roc and COR domains) and iii) protein kinase activity. Exact physiological functions are still unknown, but are implicated in autophagy and lysosome pathways. Forms dimers under physiological conditions. LRRK2-mediated neurodegeneration is GTPase and kinase dependent. A recent study has shown that LRRK2 phosphorylates and thereby inhibits the activation of a subset of Rab GTPases that function in intracellular vesicle transport (Steger M et al. elife, 2016). Note: Mutations in Rab7L1 (Park 16 locus) and Park39b (PARK 21 locus) have also previously been implicated in PD pathogenesis.

52 VPS35 (Vascular sorting protein 35; PARK17; 16q12) Retromer cargo-recognition complex (VPS35, VPS29 and VPS26a or b) Retromer mediates the transport of the membrane receptor Wntless from endosomes to the trans-golgi network The D620N mutation has been proposed to inhibit autophagy by blocking normal sorting or atg9a. D620N may be a gainof-function mutation that blocks the interaction between the retromer complex and the WASH complex.

53 PRKN (parkin, PARK2) 6q26 Nuytemans et al, aa protein E3 ubiquitin-protein ligase: tags proteins for degradation in proteosomes; also regulates autophagy of dysfunctional mitochondria Many (>100) loss-of-function missense, nonsense, splice-site, indels and CNV mutations have been detected in PARK2; haplotype analysis indicates that most mutations have arisen independently, suggesting a relatively high mutation rate. Loss of dopaminergic neurons in the substantia nigra pars compacta are detected in individuals with two mutant PARK2 genes, but Lewy bodies are not detected. Loss of PARKIN activity may allow the accumulation of PARIS (Parkin interacting substrate), leading to down-regulation of the PGC1a, a transcription factor required for proper functioning of mitochondria. A recent study has found that rapamycin can ameliorate PD like symptoms in a PARKQ311X mouse model by rescuing expression of PGC1a (Siddiqui A et al, J. Neuroscience, 2015).

54 PINK1 (PTEN-induced putative kinase 2; PARK6) The encoded 581 aa protein is a putative serine/threonine kinase that functions in the response of mitochondria to cellular and oxidative stress. May function in the same pathway as parkin. 1p35-36 Many loss-of-function mutations due to missense, nonsense, indels, CNVs have been described. CNVs seem to be largely independent in origin, while many point mutations are inherited. Nuytemans et al, 2010

55 PLINK1 and Parkin may function in mitophagy of dysfunctional mitochondria Deas E et al, 2010 NIX = pro-apoptotic member of Bcl-2 family; VDAC = voltage-dependent anion channel; GABARAP = GABA receptor-associated protein; LC3 = microtubuleassociated protein-1 light chain 3-alpha; p62 = poly-ubiquitin and LC3 binding protein

56 DJ-1 (PARK7) 1p36 Nuytemans et al, 2010 Oncogene DJ-1 encodes a 189 aa protein that is a member of the ThiJ/Pfp1 family of molecular chaperones, which are induced under conditions of oxidative stress. May function as a sensor for oxidative stress and as an antioxidant: C106-SH + ROS -> C106-SO 2 H. DJ-1 dimers localize in mitochondria Reported to modulate alpha-synuclein aggregation

57 Mutation types and frequencies vary among populations Note: LRRK2 G2385R variant is relatively common in Asian populations.

58 Additional familial PD candidate genes Gene Locus Protein Functions Mutations ATP13A2 PARK9 1p36 Putative lysosomal P- type transmembrane cation transporting ATPase PLA2G6 PARK14 Phospholipase A2, group VI Suppresses alphasynuclein toxicity in yeast; may also regulate Mn 2+ toxicity FBXO7 PARK15 F-box protein 7 Component of ubiquitin protein ligase complex DCTN1 2p13 Dynactin complex subunit eif4g1 3q27-qter eif4f complex subunit Long-distant trafficking of proteins and organelles Translation of mrnas with highly structured 5 -ends Recessive autosomal inheritance; homozygous carriers have Kufor- Rakeb disease, with parkinsonism symptoms Recessive autosomal inheritance with early onset and atypical clinical features Recessive autosomal inheritance with early onset and atypical clinical features Dominant mutations in Perry Syndrome: G71R, G71E, G71A, T72P, Q74P A502V, R1205H; Note: 4E-BP is substrate of Lrrk2 kinase; hyperphosphorylated by LRRK2 G2019S; over-expression of 4E-BP suppresses PD phenotype in PINK1 or PRKN lossof-function mutants (Drosophila);

59 Additional PD candidate genes Gene Locus Protein Functions Mutations UCHL1 PARK5 4p14 ubiquitin carboxylterminal esterase L1 Removes ubiquitin from proteins S18Y associated with sporadic, late-onset PD; role in PD still controversial GBA 1q21 lysosomal glucocerebroside MAPT 17q21.1 microtubuleassociated protein-tau BST1 4p15 cyclic ADP-ribose synthesis May be a substrate for the ubiquitin ligase parkin Microtubule stability; axonal transport Ca 2+ homeostasis Dominant missense L444P; N370S; NB: homozygous GBA causes Gaucher s disease MAPT showed genome-level significance in two independent GWAs SNPs associate with PD in Japanese GWA study RAB7L1 1q32 PARK16 intracellular vesicular trafficking including, retromer pathway SNPs in this region showed genome-level significance in two independent GWAs

60 Common genetic variants implicated in Parkinson s disease (Singleton AB et al, Movement Disorders 28, 2013)

61 Major protein degradation pathways in eukaryotic cells Matsuda N and Tanaka K, 2010

62 Intracellular pathways implicated in dominant PD (late onset; Lewy body +) NO = nitric oxide ROS = reactive oxygen species

63 Intracellular pathways implicated in autosomal recessive PD (early onset; Lewy bodies: variable) Dawson TM et al, 2010

64 Animal and in vitro Models (1) In spite of the fact that much of the underlying biology of PD is understood, there is no cure on the horizon. There is therefore a great need to improve current disease-modifying treatments and develop new therapeutic approaches. For this purpose, animal models that reproduce essential aspects of PD will be essential. Mice: Chemical lesion of SN dopamine neurons with 6-hydroxydopamine Rotenone, paraquat or trichloroethylene also induce PD-like symptoms Chronic MPTP exposure induces PD-like symptoms MitoPark mice with selective KO of mitochondrial transcription factor A (Tfam) develop a progressive form of PD that is similar to that observed in human patients Expression of A53T alpha-synuclein under the control of the prion promoter mimics many aspects of PD By contrast, PARK2/PINK2/Dj-1 individual and triple KO mice do not develop overt symptoms of PD, but do show subtle changes in motor and nigral dopamine functions.

65 Animal Models (2) Primates: Chronic MPTP exposure causes PD-like syndrome Drosophila: Over-expression of WT, A53T or A30P alpha-synuclein recapitulates many features of PD, including age-dependent loss of dopamine neurons, formation of Lewy body-like inclusions and dopamine-responsive motor deficits Mutations in PINK1 cause mitochondrial dysfunctions that are rescued by parkin Human: Dopaminergic neurons produced from fibroblasts isolated from PD patients. Fibroblasts ipscs dopamine (+) neurons