Molecular neurology

Parkinson's disease

Matthias Georg Ziller

R4 neurology
January 30th 2008
 Parkinson's disease
• Clinical picture first
described in 1817

• Motor features
– Resting tremor
– Bradykinesia
– Rigidity
– Postural instability

• Non motor features

– Autonomic
– Cognitive
– psychiatric
 Burden of Parkinson's disease

• ~ 500.000 patients in US
• 1 % of > 50 yo.
• 2nd most common MD

• Lifetime risk
– 2 % for men, 1.3 % for women

• Projected increase
– Patient numbers will double to
8-9 million worldwide in 2030

Dorsey et al., NEUROLOGY 2007;68:384-386 Projected number

of people with Parkinson disease in the most populous nations, 2005 through 2030
 PD: A history of understanding
• For a long time assumed to be a non hereditary disorder … but :

– 140 years ago – description of early familial cases

– 90 years ago – link between parkinsonism and cases of encephalitis

and formation of inclusion bodies was exlored

– 45 years ago – demonstration of dopamine deficiency, discovery of

nigro-striatal pathway and first symptomatic use of dopamine

– 25 years ago – observation of MPTP-induced toxicology

– 10 years ago – The molecular genetic revolution … monogenic forms

… a "Nature versus nurture" debate

Pathophysiology of motor manifestations

D1

Result of the direct circuit is activation of cortex and facilitation of movement

 Pathophysiology of motor manifestations

D2

Result of the indirect circuit is inhibition of thalamus, decreased activation of cortex

and inhibition of movement – Action of SNpc is antagonistic to this
 Pathophysiology of motor manifestations

Loss of dopaminergic stimulation of striatum results in greater inhibition of thalamus

and decreased cortical activation  bradykinesia
 Neuropathology

Loss of dopaminergic pigmented neurons in substantia nigra pars compacta

 Neuropathology
• Cell loss in other pigmented
nuclei
– Ventral tegmental area
– Locus coeruleus
– Raphe nucleus

• Intraneuronal eosinophilic
inclusion bodies (Lewy)
– Brainstem
– Diffuse distribution in cortex
– Staining + for alpha-synuclein

• Reactive gliosis
 Lewy body and Lewy neurites

-SYNUCLEINE +
• 2003 Braak et al.:
• Staging of PD pathology
– Allows to understand
presymptomatic disease and
non motor manifestations

• Inclusion bodies ascend in the

brainstem
– Stage 2: coeruleus
• Sleep, mood
– At stage 3 reach SNpc
• Motor manifestations

– > stage 4 cortex involved

• Dementia
• Behavioural symptoms
Progression of a-
synuclein immunopositive
labelling from stages 3 to
6, Braak 2006
 "Nurture": environmental contribution
• Factors that increase the risk

• Influenza epidemic - van Economo’s disease

• Being raised on a farm with well water
• Pesticide / herbicide / manganese exposure
• MPTP toxin exposure

• Factors that modify (maybe decrease) the risk

• Regular tobacco use and recent smoking

• Caffeinated beverages (but not EtOH)
• Physical exercise
• Some NSAIDs (Ibuprofen, but not ASA)
• Increased risk with high dairy intake in men
• Higher levels of uric acid in peripheral blood
 "Nature": Genetic forms
– Discovered during past 10 years

– Monogenic forms may be clinically indistinguishable

from "sporadic PD"

– 2-3 % of sporadic cases probably monogenic

– May lead to understanding of pathophysiology

Genetic forms
 Familial forms - Summary

• PARK 1-13
• 5 autosomal dominant:
– PARK 1 (=4), 4 is the triplication form of PARK1
– 3, 5, 8, 13

• 4 recessive:
– PARK 2,6,7,9

• 1 X-linked:
– PARK 12

• 2 with unknown mode of transmission: 10,11

• additional mutations in small families without assigned PARK-

numbers
 Alpha-synuclein (PARK1+4)
• First described + associated with familial parkinsonism 1997
– SNCA gene, chrom 4q

• 3 known point mutations

• single-allele duplication or triplication in some families
– initially named PARK4

• Penetrance low, 33 %
• Severity depends on gene dosage,
– patients with duplication resemble “idiopathic” PD more than triplication
cases

• Mutations and multiplications are rare

• Increased expression of wild type SNCA copies may play a role in

"idiopathic disease"
 What is SNCA ?
• 140-residue cytosolic and lipid-binding
protein
• Natively unfolded, presynaptic protein
– synaptic vesicle recycling, storage of
transmitters
– negative coregulator of transmitter
release

• Identical with Non-amyloid β

component precursor

• Oligomer-forming SNCA in LBs

– truncated, oxidized and phosphorylated
variants

• Mutants have greater nuclear targeting

in cell culture
Speculative Model of the Interactions among Proteins Implicated in Parkinson's Disease

Feany M. N Engl J Med 2004;351:1937-1940

 LRRK-2 (PARK8)
• Leucin-rich repeat kinase 2 gene, identified by 2 groups in 2004
• Mutations often associated with late-onset disease
– contrary to previous beliefs

• Large gene, 51 exons

• encoding 2.527 AA- protein with several functional domains
• Certain genotypes may be susceptibility factors
• 16 sequence changes clearly pathogenic, in only 10 of 51 exons
– clustering in C-terminal region of protein
– pathology linked to kinase function but unclear*

• most frequent mutation is c.6055G

–  A, 1.5 % of index cases with late-onset “idiopathic” PD
– same mutation found in 40% of Arab PD patients and 20 % of Ashkenazi Jewish
PD  likely founder effect

• homozygous forms of this mutation exist, but are not more severe than
heterozygous

*Greggio et al., Neurobiol Dis 2006,

Kinase activity is required for the toxic effects of mutant LRRK2/dardarin
 Parkin (PARK2)
• Identified 1998, earlier age at onset, slower progression
• most common factor (10-20%) for early onset disease, all ethnic groups
• large number of known mutations
• 12 exons, chrom 6

• Gene product Parkin: 465 aa protein,

– ubiquitin ligase, activity disrupted in mutants

• Mediates ubiquitilation of target proteins

– loss of ligase activity leads to pathology
– May lead to accumulation of substrates

• Parkin essential in formation of Lewy bodies ?

• Possible role in mitochondrial integrity
• Effector mechanisms
– Rescues mitochondrial dysfunction in flies after inactivation of PINK
– SNCA induced dysfunction is enhanced due to lack of Parkin activity
– Posttranslational modification of Parkin due to oxidative or nitrosative stress
compromises ligase activity
Speculative Model of the Interactions among Proteins Implicated in Parkinson's Disease

Feany M. N Engl J Med 2004;351:1937-1940

Ubiquitin Proteasome System
 Ubiquitine-Proteasome System in PD
• Parkin, E3-UB ligase
– Loss of function in mutants
– Mediates ubiquitylation of synphilin, SNCA-interacting protein

• UCHL-1 (PARK5), deubiquitylating enzyme

– Impaired UB-hydrolysis

• Alpha-synuclein inhibits proteasome

– But no generalized dysfunction in brain regions with LBs

• Precise mechanism of UPS dysfunction in PD unclear

– Decreased function with toxin exposure ?
– MPTP inhibits proteasome
– Changes linked to mitochondrial function, aging ?
 PINK1(PARK6)
• =PTEN-induced kinase 1, mutations found in 2004

• 3 families with consanguinity and autosomal recessive parkinsonism

– 1-8 % of populations, considerable variation between ethnic groups

• Mutations near kinase domain

–  loss of function in vivo

• 581 aa-protein, mitochondrial localization

– Highly conserved kinase domain, mitochondrial targeting motif
– no known substrate for kinase activity

• Mutations lead to different phosphorylation patterns

• Proteasomal stress enables altered cleavage of PINK and possible
accumulation in LBs

• Interaction with DJ-1: recruited to pathway after oxidative damage

due to PINK dysfunction
 DJ-1 (PARK7)
• Described 2003, chrom 1, 1-2 % of early onset cases

• Localization:
– ubiquitous expression
– cytosolic, may translocate to mitochondria with pH changes
• Function:
– chaperone-like activity,
– intracellular sensor of oxidative stress
– regulates D2 receptor signaling
– Direct action as antioxidant ?

• DJ-1 mutants
– show increased vulnerability to energy metabolism changes

• Overexpression of DJ-1 protects against mitochondrial complex 1 inhibitors

– effect is abrogated by mutants
Speculative Model of the Interactions among Proteins Implicated in Parkinson's Disease

Feany M. N Engl J Med 2004;351:1937-1940

Oxidative stress and mitochondrial dysfunction

• Mitochondrial complex 1 inhibitors MPTP

– rotenone, paraquat
– Reproduce parkinsonism with dopaminergic neuron loss
– Absence of LB in these cases

• Chronic infusion reproduces all features, including LBs

– Supports theory of environmental toxin interaction by inhibition
respiratory chain

• Inhibition of complex I:
– Depletion of ATP, impairment of dependent processes
– Generation of free radicals causing oxidative stress

• Post-mortem evidence of ox stress in PD brains

– Elevated lipid peroxidation and nitration levels in SN and LBs
– Reduced levels of glutathione and oxidized glutathione (antioxidants)
– Reduced complex 1 activity in muscle, brain, platelets of PD patients
 More questions: Role of susceptibility genes

1. Exact role is controversial:

- genes connected to monogenic forms may also act as susceptibility genes
- Single mutations in “recessive” genes are common (Parkin, DJ-1, PINK1)

2. Heterozygous mutations
- may be associated with PET, MRI, ultrasound changes
- Subtle motor signs in “asymptomatic” carriers
- Unclear if these are developmental changes, early disease
markers or adaptive

3. Are modifications of wild-type forms linked to parkinsonism ?

ubiquitylation studies of wild-type Parkin linked to sporadic PD …

4. Genetic polymorphisms may be associated with disease

- pG2385 increases risk in Chinese population
 References
• Continuum 2004, No.3 Movement Disorders, Suchowersky and
Furtado
• Schlossmacher & Klein, Neurology, 2007, 69, 2093, PD – 10 years
after its genetic revolution
• Lim et al, BMC Biochemistry 2007, 8, S1, Role of ubiquitin
proteasome system in Parkinson's disease
• Thomas & Flint Beal, Parkinson's disease, Human Molecular
Genetics 2007, Vol 16, Review Issue 2, 183-194
• Gandhi & Wood, Molecular pathogenesis of PD, Human Molecular
Genetics 2005, 14, 18, 2749-2755
• Braak et al., Stanley Fahn Lecture: The staging procedure for
Inclusion Body Pathology, Mov Disorders 21, no.12, 2006, 2042-
2051
• Feany M. N Engl J Med 2004;351:1937-1940
• 16th Annual National Residents’ Seminar on Movement Disorders
Course Notes, Ottawa 2008