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 PROGRESS IN BRAIN RESEARCH

VOLUME 183

RECENT ADVANCES IN PARKINSON’S

DISEASE: BASIC RESEARCH

EDITED BY

ANDERS BJÖRKLUND
Wallenberg Neuroscience Centre
Division of Neurobiology
Lund University
Lund, Sweden

M. ANGELA CENCI
Basal Ganglia Pathophysiology Unit
Department of Experimental Medical Science
Lund University
Lund, Sweden

AMSTERDAM – BOSTON – HEIDELBERG – LONDON – NEW YORK – OXFORD

PARIS – SAN DIEGO – SAN FRANCISCO – SINGAPORE – SYDNEY – TOKYO
Elsevier
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First edition 2010

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 List of Contributors

D.M. Alessi, Departments of Neurology, Pathology and Cell Biology, and the Center for Motor Neuron
Biology and Disease, Columbia University, New York, NY, USA
C. Baunez, Laboratoire de Neurobiologie de la Cognition (LNC), UMR6155 CNRS/Aix-Marseille
Université, Marseille, France
H. Bergman, The Interdisciplinary Center for Neural Computation; Institute for Medical Research Israel-
Canada (IMRIC), Department of Medical Neurobiology (Physiology), The Hebrew University,
Jerusalem, Israel
R.E. Burke, Departments of Neurology, Pathology and Cell Biology, Columbia University, New York,
NY, USA
P. Calabresi, Fondazione Santa Lucia IRCCS, Rome, Italy; Clinica Neurologica, Università degli Studi di
Perugia, Ospedale S. Maria della Misericordia, Perugia, Italy
A.R. Carta, Department of Toxicology, University of Cagliari, Cagliari, Italy
M.A. Cenci, Basal Ganglia Pathophysiology Unit, Department of Experimental Medical Science, Lund
University, Lund, Sweden
S. Chan, Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago, IL,
USA
M.R. Cookson, Cell Biology and Gene Expression Unit, Laboratory of Neurogenetics, National Institute
on Aging, Bethesda, MD, USA
T.M. Dawson, NeuroRegeneration and Stem Cell Programs, Institute for Cell Engineering, Johns
Hopkins University School of Medicine, Baltimore, MD, USA; Department of Neurology, Johns
Hopkins University School of Medicine, Baltimore, MD, USA; Solomon H. Snyder Department of
Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, USA
V.L. Dawson, NeuroRegeneration and Stem Cell Programs, Institute for Cell Engineering; Department of
Neurology; Department of Physiology; Solomon H. Snyder Department of Neuroscience, Johns Hopkins
University School of Medicine, Baltimore, MD, USA
M. Day, Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago, IL,
USA
R.L.A. deVries, Departments of Neurology, Pathology and Cell Biology, and the Center for Motor
Neuron Biology and Disease, Columbia University, New York, NY, USA
M. di Luca, Department of Pharmacological Sciences, University of Milano, Milano, Italy
S. Elias, Institute for Medical Research Israel-Canada (IMRIC), Department of Medical Neurobiology
(Physiology), The Hebrew University, Jerusalem, Israel
M. Fournier, Laboratory of Molecular Neurobiology and Neuroproteomics, Brain Mind Institute,
The Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
M.J. Frank, Department of Cognitive, Linguistic, and Psychological Sciences, Department of Psychiatry
and Human Behavior, and Brown Institute for Brain Science, Brown University, Providence, RI, USA

v
vi

F. Gardoni, Department of Pharmacological Sciences, University of Milano, Milano, Italy

T. Gasser, Hertie Institute for Clinical Brain Research, Department of Neurodegenerative Diseases,
Tübingen, Germany; DZNE, German Center for Neurodegenerative Diseases, Tübingen
T. Gertler, Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago,
IL, USA
V. Ghiglieri, Fondazione Santa Lucia IRCCS, Rome, Italy
J.A. Goldberg, Department of Physiology, Feinberg School of Medicine, Northwestern University,
Chicago, IL, USA
P. Gubellini, Institut de Biologie du Développement de Marseille-Luminy (IBDML), UMR6216 CNRS/
Aix-Marseille Université, Marseille, France
J.N. Guzman, Department of Physiology, Feinberg School of Medicine, Northwestern University,
Chicago, IL, USA
G. Heimer, Institute for Medical Research Israel-Canada (IMRIC), Department of Medical Neurobiology
(Physiology), The Hebrew University, Jerusalem, Israel
V. Jackson-Lewis, Departments of Neurology, Pathology and Cell Biology, and the Center for Motor
Neuron Biology and Disease, Columbia University, New York, NY, USA
A. Kachroo, MassGeneral Institute for Neurodegenerative Disease, Massachusetts General Hospital,
Boston, MA, USA
C. Konradi, Center for Molecular Neuroscience and Kennedy Center for Research on Human Development,
Departments of Pharmacology and Psychiatry, Vanderbilt University, Nashville, TN, USA
H.A. Lashuel, Laboratory of Molecular Neurobiology and Neuroproteomics, Brain Mind Institute, The
Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
I. Martin, NeuroRegeneration and Stem Cell Programs, Institute for Cell Engineering, Johns Hopkins
University School of Medicine, Baltimore, MD, USA; Department of Neurology, Johns Hopkins
University School of Medicine, Baltimore, MD, USA
M. Morelli, Department of Toxicology, University of Cagliari, Cagliari, Italy
A. Oueslati, Laboratory of Molecular Neurobiology and Neuroproteomics, Brain Mind Institute, The
Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
J.L. Plotkin, Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago,
IL, USA
S. Przedborski, Departments of Neurology, Pathology and Cell Biology, and the Center for Motor Neuron
Biology and Disease, Columbia University, New York, NY, USA
M. Rivlin-Etzion, The Interdisciplinary Center for Neural Computation; Institute for Medical Research
Israel-Canada (IMRIC), Department of Medical Neurobiology (Physiology), The Hebrew University,
Jerusalem, Israel
J. Sanchez-Padilla, Department of Physiology, Feinberg School of Medicine, Northwestern University,
Chicago, IL, USA
M.A. Schwarzschild, MassGeneral Institute for Neurodegenerative Disease, Massachusetts General
Hospital, Boston, MA, USA
W. Shen, Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago, IL,
USA
D.J. Surmeier, Department of Physiology, Feinberg School of Medicine, Northwestern University,
Chicago, IL, USA
X. Tian, Department of Physiology, Feinberg School of Medicine, Northwestern University, Chicago, IL,
USA
 vii

M. Tocilescu, Departments of Neurology, Pathology and Cell Biology, and the Center for Motor Neuron
Biology and Disease, Columbia University, New York, NY, USA
C. Vives-Bauza, Departments of Neurology, Pathology and Cell Biology, and the Center for Motor
Neuron Biology and Disease, Columbia University, New York, NY, USA
T.V. Wiecki, Department of Cognitive Linguistic, and Psychological Sciences, Department of Psychiatry
and Human Behavior, and Brown Institute for Brain Science, Brown University, Providence, RI, USA
 Preface

Research on Parkinson´s disease (PD) is one of the most dynamic fields of modern neuroscience. It is an
excellent example of how clinical and basic research can fruitfully interact and inspire each other in a truly
translational way. During the decades after the discovery of dopamine in the late 1950s the field was
dominated by pharmacological and neurochemical approaches. Since the discovery of the role of alpha-
synuclein and its role in PD pathogenesis in the late 1990s, PD research has entered a new exciting phase
of development, and its scope has broadened to include dynamic molecular and genetic approaches. This
had led to the discovery of further genetic mutations accounting for familial forms of the disease (Parkin,
DJ-1, Pink-1, and leucine-rich repeat kinase 2), and spurred intense molecular biological investigations on
the mechanisms of neurodegeneration in PD. In addition to molecular genetics, other fields of PD
research have undergone a dramatic development during the past 20 years. Significant progress has
been made modelling PD in animals both on a symptomatic level and on a mechanistic perspective. The
current availability of a diversified range of models in different species provides neuroscientists with
articulate tools to study molecular mechanisms, test pathophysiological hypotheses and identify new
treatment principles. The discovery that PD motor symptoms and treatment-induced dyskinesias are
dramatically ameliorated by high-frequency stimulation of some deep basal ganglia nuclei has prompted
efforts on the part of both neurophysiologists and computational neuroscientists to decipher the basic
neural operations of the basal ganglia in health and disease. Finally, technological developments in the
area of brain imaging have provided exciting new opportunities for pathophysiological investigations,
differential diagnosis and treatment monitoring in PD patients.
These two companion volumes of Progress in Brain Research were composed to capture all the richness
and complexity of PD as a topic for basic, translational and clinical investigation. A year ago, when we
approached leading researchers in the different subfields to contribute, the vast majority of the invited
authors enthusiastically accepted the invitation and delivered contributions that turned out to represent
the utmost state-of-the art in each given field. It is with great pleasure and pride that we now present this
collection of review chapters to a broad audience of readers.
The chapters have been grouped into two volumes and five sections. The first volume covers basic and
molecular investigations of the mechanisms of neurodegeneration in PD (Section I: Genetic and
molecular mechanisms of neurodegeneration in PD) and the secondary adaptations that affect the basal
ganglia at both the single cell level and the system level (Section II: Cellular and system-level pathophy-
siology of the basal ganglia in PD). The second volume focuses on translational and clinical aspects of PD
research reviewing animal models of PD from drosophila to non-human primate species (Section I:
Animal models of PD) the very dynamic area of functional neuroimaging (Section II: Exploring PD with
brain imaging) and the most challenging therapeutic developments (Section III: Frontiers in PD
treatment).
Like no other neurological disease, PD is inspiring enormously diversified research themes and
approaches in a way that would have been impossible to foresee some 10 years ago. An increasing number

ix
x

of investigators, also from areas outside neuroscience, have joined the PD research community and are
now contributing to the richness and diversity of this field. Over the last 15 years, several international PD
patient-initiated, non-profit organizations have dramatically improved the funding possibilities for this
area of research which is now advancing at an extremely rapid pace. It is our hope that this formidable
development of knowledge and technologies will deliver novel options for treatment – and eventually a
cure – for all who suffer from this disease.
In closing we would like to express our warmest thanks to all the authors for their outstanding
contributions and to Gayathri Venkatasamy, our Developmental Editor at Elsevier, for her expert and
patient assistance.

Lund, June 11th 2010

Anders Björklund
M. Angela Cenci
 SECTION I

Genetic and molecular mechanisms of

neurodegeneration in PD
A. Bjorklund and M. A. Cenci (Eds.)
Progress in Brain Research, Vol. 183
ISSN: 0079-6123
Copyright
2010 Elsevier B.V. All rights reserved.

CHAPTER 1

Identifying PD-causing genes and genetic

susceptibility factors: current approaches and future
prospects

Thomas Gasser

Hertie Institute for Clinical Brain Research, Department of Neurodegenerative Diseases, T€

ubingen, Germany, and
DZNE, German Center for Neurodegenerative Diseases, T€ ubingen

Abstract: Over the last years, a plethora of genetic findings have completely changed our views on the
aetiology of Parkinson’s disease (PD). Linkage studies and positional cloning strategies have identified
mutations in a growing number of genes which cause monogenic autosomal-dominant or autosomal-
recessive forms of the disorder. While these Mendelian forms of PD are relatively rare, high-throughput
genotyping and sequencing technologies have more recently provided evidence that low-penetrance
variants in at least some of these genes also play a direct role in the aetiology of the common sporadic
disease. In addition, rare variants in other genes, such as the Gaucher’s disease-associated
glucocerebrosidase A, have also been found to be important risk factors at least in subgroups of patients.
Thus, an increasingly complex network of genes contributing in different ways to disease risk and
progression is emerging. These findings provide the ‘genetic entry points’ to identify molecular targets
and readouts necessary to design rational disease-modifying treatments.

Keywords: Parkinson’s disease; Genetics; Genetic risk factors; DNA polymorphisms

Introduction innovations of the 1980s, such as polymerase chain

reaction amplification of DNA fragments and the
The progress of molecular genetic technology over discovery of polymorphic micro-satellite repeat
the last 25 years has revolutionized our understand- elements in the genome (usually consisting of
ing of Parkinson’s disease (PD) and many other repetitive di-, tri- or tetranucleotide sequences
common complex disorders. The technological which proved to be extremely useful as landmarks
(‘DNA markers’) to map the genome) hand in

hand with the development of appropriate statis­
Corresponding author.
Tel.: 07071/2986529; Fax: 07071/294839; tical tools and computer programmes, led to the
E-mail: [email protected] mapping and cloning of a large number of genes

DOI: 10.1016/S0079-6123(10)83001-8 3
4

which cause – when mutated – monogenic dis­ The advent of micro-array technology which
eases, that is inherited disorders following a Men­ provided the opportunity to study hundreds
delian mode of transmission (Gasser, 2009a). of thousands or even millions of genetic variants
Most of these classic neurogenetic disorders such (usually single-nucleotide polymorphisms, ‘SNPs’)
as Huntington’s disease, myotonic dystrophy or in a large cohort of patients and controls has
spinal muscular atrophy, to name just a few, are provided the basis for a new generation of genetic
relatively rare. Nevertheless, with the identifica­ studies, genome-wide association studies (GWAS)
tion of these rare disease genes ‘neurogenetics’ in order to go beyond the analysis of single candi­
has become part of the mainstream of neurology date genes and to systematically analyse, in an
(Harbo et al., 2009). unbiased way, the genetic risk profile of complex
During the 1990s it became apparent that in some diseases. While most GWAS evaluate the role of
rare cases the much more common and typically common genetic variability as risk factors for a
sporadic neurologic disorders such as Parkinson’s disease, this approach has already been success­
disease (PD) (Denson and Wszolek, 1995), fully applied also to quantitative traits such as age
Alzheimer’s disease (AD) (Goate et al., 1991) or of onset in PD (Latourelle et al., 2009) or to
amyotrophic lateral sclerosis (Rosen et al., 1993) laboratory values such as serum levels of uric
could also run in families following a Mendelian acid (Dehghan et al., 2008). Another extension
pattern of inheritance. In fact, it turned out that of this technology is the combination with gen­
many of these monogenic variants of common dis­ ome-wide transcriptome analysis (Elstner et al.,
eases resemble the typical sporadic forms to a large 2009), promising a deeper insight into relevant
degree both clinically (with the exception that age of gene regulation networks.
onset is often younger in patients with inherited The next technological revolution is already
forms of these disorders) and pathologically, sug­ under way. Massive-parallel sequencing will
gesting that the molecular pathways discovered make large-scale whole exome or even whole gen­
through the relevant genes in hereditary forms may ome sequencing feasible. This will be necessary to
also be of importance in sporadic cases. The same identify the suspected multitude of rare genetic
gene identification strategies used in classical neuro­ variants in different genes which are thought to
genetic diseases were successfully used to show that explain another substantial fraction of the genetic
the Mendelian forms of the respective common dis­ risk for common diseases.
orders were also caused by mutations in single Methods are being developed to deal with the
genes. The identification of those disease genes, increasingly complex and vast amount of informa­
such as SNCA, the gene encoding alpha-synuclein tion generated by current and future sequencing
(aSYN) for PD, or APP, the gene coding for technology.
the amyloid precursor protein in AD, was a crucial
step in the elucidation of the chain of molecular
events which lead to neurodegeneration in these Identification of monogenic forms of PD by
disorders. positional cloning strategies
It was then only in recent years that accumulating
evidence suggested that the close resemblance Autosomal-dominant forms of inherited PD
between familial and sporadic forms on the clinical
and pathologic level also has its correspondence on The classic approaches of linkage analysis and
the genetic level: common genetic variants in genes positional cloning have been the uniquely success­
identified in monogenic forms or in genes belonging ful strategies to identify genes causing the autoso­
to the identified pathways have been found to mal-dominantly inherited diseases including the
modify the risk to develop a sporadic disease. major forms of familial PD.
 5

This strategy relies on the availability of large individuals were identified with a relatively early-
and clinically well-characterized families, usually onset form of PD (average age of onset was 46
with at least 8–10 affected family members. By years), a high rate of dementia and an unusually
studying the co-segregation of genetic (DNA)mar­ severe and rapid course (average disease duration
kers (most often highly polymorphic dinucleotide less than 10 years), segregating as an autosomal-
repeat elements, above-mentioned micro-satellite dominant trait (Golbe et al., 1990). Only a year
polymorphisms, were used), the genetic locus of later, the same group identified a putative disease-
the disease-causing gene in a given family can be causing mutation in a known gene of the region,
narrowed down to a region of several million base alpha-synuclein (the gene is abbreviated as
pairs (megabases, Mb) of DNA. The statistical SNCA, the protein as aSYN). It was a single
method to estimate the likelihood that a particular base-pair change (a G to A transition) at position
set of neighbouring DNA markers (a so-called 209 of the coding sequence, leading to the change
haplotype) are co-inherited with a disease gene from alanine to threonine at position 53 of the
as a result of its physical proximity on the chromo­ aSYN protein (A53T) (Polymeropoulos et al.,
some (i.e. that DNA markers and disease gene are 1997). As expected, the mutation co-segregated
‘linked’) is called linkage analysis. The most with the disease in the affected family, but this in
important prerequisite for this type of study, in itself is no proof of pathogenicity. Depending on
addition to the availability of sufficiently large the size of a linked genetic region, the affected
families, is the unequivocal classification of affected members in a family will share a large number of
and unaffected family members. Erroneous classi­ candidate genes and consequently all genetic var­
fication, which in many age-related complex dis­ iants that are located in this region. In fact, the
eases is a real possibility, will lead to false linkage causative role of the SNCA mutation initially was
results (Gasser, 2008). When a disease locus is doubted by some researchers based on the surpris­
identified with sufficient confidence (a so-called ing fact that the highly homologous mouse SNCA
lod score of >3 is equivalent to a genome-wide p- gene contains the threonine thought to be patho­
value of 0.05 and is considered to be significant genic in humans at position 53. It was therefore of
evidence), all the genes in the identified region great importance to find additional independent
have to be sequenced and analysed for potentially sequence variants segregating with PD in other
disease-causing mutations. Of course, not all of the families. Eventually, those variants were found,
identified sequence variants in a linked region are although they are extremely rare: only two further
pathogenic. This means that either the demonstra­ pathogenic point mutations in SNCA have
tion of mutations in several independent families been recognized, leading to the exchange A30P
co-segregating with a disease is necessary (amount­ (Krüger et al., 1998) and E46K (Zarranz et al.,
ing in effect to a replication of the initial finding) or 2004), respectively, each in a single, large, domi­
the careful functional studies in model systems are nant family, reflecting the high penetrance of
required to prove pathogenicity. these mutations. SNCA point mutations are very
rare and have not been found in large cohorts of
patients with sporadic PD (Berg et al., 2005).
PARK1 (alpha-synuclein) Further important insight into the link between
SNCA and PD came from the discovery of gene
It was by classic linkage analysis that Polymero­ multiplication mutations. A family with autoso­
poulos et al. mapped the disease locus in a large mal-dominant parkinsonism, also frequently
Italian family with autosomal-dominant PD to the accompanied by dementia, had been mapped to
long arm of chromosome 4 (Polymeropoulos the short arm of chromosome 4 (4p15) and had
et al., 1996). In this family, more than 40 therefore been thought to be genetically distinct
6

from the families with SNCA mutations (Farrer protein aggregates which had long been recog­
et al., 1999). The affected members resembled nized as the pathologic hallmark in familial as
those with SNCA mutations both clinically and well as sporadic cases of the disease (Fig. 1). The
pathologically to a remarkable degree (see currently favoured hypothesis states that the
below). It was therefore not too surprising when amino acid changes in aSYN lead to an increased
it became apparent that the assignment to chro­ tendency of the protein to form oligomers and
mosome 4p was in fact due to a genotyping error fibrillar aggregates (Goedert et al., 1998; Karpinar
and that the disease also co-segregated with the et al., 2009), eventually resulting in neuronal dys­
SNCA locus in this family. However, direct function and cell death, although the precise rela­
sequencing of the SNCA gene did, however, not tionship between mutations, aggregate formation
reveal any putative pathogenic mutations. Instead, and their deleterious effects on neurons is still
Singleton and co-workers found a triplication of unknown. Many studies favour the hypothesis
the entire locus containing the SNCA gene in the that the mature aggregates (i.e. Lewy bodies and
affected members of this pedigree (Singleton Lewy neuritis detected on the light microscopy
et al., 2003). This finding is of particular relevance level) are not themselves the toxic moiety, but
with respect to the molecular pathogenesis of PD, rather an attempt of the cell to clear much smaller
as it suggests that not only structurally altered toxic oligomers (Cookson and van der Brug,
aSYN can cause PD but that also the wild-type 2007), while the elusive oligomers are the true
aSYN protein is pathogenic, if it is over- toxic moiety (Conway et al., 2000).
expressed. In fact, the triplication is associated Mutation carriers from the first family described
with a roughly twofold over-expression of the with an SNCA mutation (the ‘Contursi’ kindred)
aSYN protein in the brain of affected individuals,
as shown by Western blotting on autopsy material
(Singleton et al., 2003).
Subsequently several additional families with
SNCA triplications, and also with SNCA duplica­
tions, have been found (Ibanez et al., 2009). In
those gene locus multiplication families, a clear
dose dependence of the pathogenic effect was
observed: SNCA duplications, that is a 50%
increase of the gene dosage (three instead of the
usual two gene copies) leads to relatively late-
onset dopa-responsive parkinsonism resembling
typical PD, while triplications are associated with
an earlier disease onset, a high prevalence of
dementia and a rapid disease course. This could
be even shown in a single family in whom different
branches segregated a duplication and a triplica­
tion of a 1.5 Mb genomic fragment containing the
SNCA gene (Fuchs et al., 2007).
The identification of the first SNCA mutations Fig. 1. aSYN immunopositive neuronal inclusions in the
dorsal motor vagal nucleus of a patient with an A30P SNCA
by Polymeropoulos and co-workers soon leads to
mutation. Marked aSYN pathology is obvious with numerous
the discovery that the encoded protein (aSYN) is Lewy bodies (arrowheads) and Lewy neurites (arrows). Figure
the major fibrillar component of the Lewy body kindly provided by Prof. R. Krüger, Hertie Institute for Clinical
and Lewy neurites (Spillantini et al., 1997), the Brain Research, Tübingen.
 7

clinically had relatively early onset of a mostly showed typical L-dopa-responsive parkinsonism
akinetic/rigid form of PD with rapid progression with onset in their fifties (Funayama et al., 2002).
and commonly also suffered from dementia. By positional cloning, missense mutations in the
The clinical spectrum was later confirmed and gene for leucine-rich repeat kinase 2 (LRRK2)
extended in additional families carrying the same (Paisan-Ruiz et al., 2004; Zimprich et al., 2004)
mutation, in whom prominent autonomic distur­ were found to be disease causing. The gene
bances were also noted (Spira et al., 2001). spans a genomic region of 144 kb, with 51 exons
A very similar clinical picture with early dementia encoding 2527 amino acids, and to date, at least six
and autonomic disturbances, thereby more resem­ verified disease-causing mutations have been
bling a variant of PD called ‘dementia with Lewy identified.
bodies’ (DLB) rather than typical PD, was Mutations in the LRRK2 gene are clearly the
described in the family with the E46K mutation most common cause of dominantly inherited PD
(Zarranz et al., 2004). On the other hand, a more discovered so far. In a number of studies across
typical late onset of parkinsonian symptoms with several different populations between 5 and 15%
only late development of relatively mild dementia of dominant families carry mutations in LRRK2
was described in a German family with the A30P (Berg et al., 2005; Di Fonzo et al., 2005). The
mutation (Krüger et al., 1998). Although this single most common mutation, G2019S, is respon­
family is relatively small and therefore this conclu­ sible for familial PD in up to 7% of familial cases
sion rests only on very few cases, there appear to be in different Caucasian populations (Di Fonzo
mutation specific differences in disease et al., 2005; Kachergus et al., 2005; Nichols et al.,
presentation. 2005). This mutation has also been found in about
Pathologically, fibrillar aSYN aggregates (i.e. 1–2% of sporadic patients of European descent
Lewy pathology) were found in all patients with (Gilks et al., 2005), indicating that the mutation
SNCA mutations, both point mutations and multi­ has a reduced penetrance, which was estimated
plications, not only in the substantia nigra but also between 35 and 70% (Goldwurm et al., 2005;
in other brain stem nuclei and widespread in Ozelius et al., 2006) and which must be taken
mesocortical and neocortical neurons, again com­ into account in genetic counselling. Even higher
patible with a diagnosis of DLB (Spira et al., G2019S prevalence rates of up to 40% were found
2001). Interestingly, in mutation-positive cases, in genetically more isolated populations, such as
aSYN pathology was also found in oligodendro­ the Ashkenazi Jewish or the North African Arab
glial cells, a feature thought to be typical for multi­ populations, both in sporadic and in familial cases
ple system atrophy (MSA) (Dickson et al., 1999). (Lesage et al., 2006; Ozelius et al., 2006) due to
This large overlap of pathologic features of PD, genetic founder effects.
DLB and MSA in cases with SNCA mutations Despite its reduced penetrance, the G2019S
strongly supports the close aetiologic link between mutation is usually thought of as a true ‘disease­
these disease entities. causing mutation’, because it is very rare in all
control populations studied so far. Other more
common variants in the LRRK2 gene, on the
PARK8 (LRRK2) other hand, appear to act more like risk factors
of modest effect sizes. The G2385R (Farrer et al.,
Another locus for a dominant form of PD was first 2007) variant, for example, was found not only in
mapped in a large Japanese family to the pericen­ approximately 9% of Chinese patients with PD
tromeric region of chromosome 12 and named but also in about 3% of controls. The same role
PARK8, again by a classic linkage analysis of a risk allele has been suggested for the R1628P
approach. Affected members in this family exchange (Lu et al., 2008; Ross et al., 2008).
8

Clinical signs and symptoms of LRRK2-related PARK2 (Parkin)

disease closely resemble typical sporadic PD.
This is also true for age of onset, which is on It was in autosomal-recessive families with very
average in the late fifties and only slightly below early-onset parkinsonism that the first recessive
that in non-mutation-carrying PD patients (Healy PD gene was mapped to the long arm of chromo­
et al., 2008). However, age at onset as well as some 6 in the vicinity of the gene for superoxide
severity of the disease may be highly variable, dismutase 2 (SOD2) (Matsumine et al., 1997).
even within families. Because of the suspected role of toxic oxygen
Pathological changes in patients with LRRK2 radicals in the pathogenesis of PD, SOD2 was a
mutations are consistent with typical Lewy body plausible candidate gene. However, no sequence
PD in most cases reported so far and also include variants in this gene could be identified. Rather, a
diffuse Lewy body disease, nigral degeneration with­ number of different mutations were found in a
out distinctive histopathology and rarely even pro­ very large neighbouring gene which was then
gressive supranuclear palsy-like tau aggregation. called Parkin (PRKN) (Kitada et al., 1998).
LRRK2 mutations may therefore be an upstream PRKN mutations turned out to be a common
event in the cascade leading to neurodegeneration cause of parkinsonism with early onset, particularly
with different pathologies. Although the natural sub­ in individuals with evidence of recessive inheri­
strate and disease-relevant function of LRRK2 is tance. Nearly 50% of sibling pairs with PD were
unknown, cell culture studies suggest that pathogenic found to have PRKN mutations (Lücking et al.,
mutations seem to be associated with increase, rather 2000), if at least one of them had an age of onset
than a loss, of kinase activity (Gloeckner et al., below 45 years. As recessive diseases often appear
2006), raising the interesting possibility that kinase to be sporadic, particularly in societies with rela­
inhibition may be a potential therapeutic strategy. tively small families, because statistically only 25%
of the offspring of two heterozygous mutation car­
riers will be homozygous for the disease allele,
Autosomal-recessive parkinsonism PRKN mutations are also responsible for the major­
ity of sporadic cases with very early onset (before
The strategies of linkage mapping and positional age 20) and are still common (25–40%) when onset
cloning can also be used to identify loci in genes is between 20 and 35 years. PRKN mutations are
responsible for autosomal-recessive monogenic rare in sporadic cases with onset later than 45 years.
diseases. This mode of inheritance is characterized
typically by the occurrence of the disease in sib­
lings while the parents are obligatory heterozygous Other recessive forms of parkinsonism
mutation carriers and usually remain healthy.
Autosomal-recessive PD has clinically been first Mutations in the PINK1 gene (PARK6) have
recognized and characterized in Japan (Ishikawa been identified as another cause for autosomal-
and Tsuji, 1996). Sibling pairs with PD often have recessive early-onset parkinsonism (Valente
much earlier age of onset compared with patients et al., 2004), again following a linkage mapping
with the sporadic disease, which is why the term approach in several multiplex recessive families
‘autosomal-recessive juvenile parkinsonism’, has (Valente et al., 2001). This gene is particularly
been coined. Since families with a recessive disease interesting within the context of the findings link­
are usually much smaller than multigenerational- ing PD to mitochondrial dysfunction and oxidative
dominant pedigrees, linkage analysis is only stress, as PINK1 encodes a mitochondrially
successful if several families mapping to the same located protein. Mutations in the PINK1 gene
locus are included into a study. are less common than PRKN mutations in most
 9

populations studied and probably account for only While some mutations such as R1441C or Y1699C
1–2% of early-onset cases (Hatano et al., 2004; have so far only been found in families with clear
Rogaeva et al., 2004; Rohe et al., 2004; Valente autosomal-dominant inheritance and thus are con­
et al., 2004). From work with animal models it has sidered to be high-penetrance disease genes, the
become quite clear that PINK1 acts in a common G2019S variant is clearly a disease gene with mark­
pathway with PRKN (Dodson and Guo, 2007). edly reduced penetrance, which is still very rare in
However, the natural substrate of the kinase unaffected individuals, while the variants G2385R
activity of PINK1 is still unknown. or R1628P are found in >1% of the healthy
Mutations in the DJ-1 gene (PARK7) are yet Chinese population and thus must be considered
another rare cause of autosomal-recessive parkin­ relatively ‘common’ genetic variants, which are
sonism (Bonifati et al., 2002; Healy et al., 2004; associated with an approximately threefold
Hedrich et al., 2004). The clinical picture with increased risk to develop PD.
early onset and slow progression is similar to the In addition to the concept that rare mutations
other recessive parkinsonian syndromes. Follow­ cause rare genetic diseases and – on the other
ing the initial discovery of two mutations in an hand – common variants are associated with a mod­
Italian and a Dutch family (Bonifati et al., 2002), erate increase of relative risk for common disorders,
only a few additional bona fide pathogenic muta­ another concept has emerged over the last several
tions [one homozygous (Hering et al., 2004) and years: the rare variant-common disease hypothesis.
one compound heterozygous (Abou-Sleiman It states that multiple rare variants in a potentially
et al., 2003)] have been identified. large number of genes may each be significant (par­
While mutations in the genes named above all tial) causes in a relatively small proportion of
cause a ‘pure’ form of early-onset parkinsonism, patients with a common disease such as PD. This
an increasing number of genes have been found to hypothesis is exemplified in the role of mutations in
cause more complex phenotypes which includes, the gene for glucocerebrosidase (GBA) in PD.
in addition to parkinsonism, often dystonia, spas­ About 10 years ago, astute clinical observation
ticity, and dementia (Klein et al., 2009). suggested that patients with Gaucher’s disease, an
The common denominator for all those cases is autosomal-recessive, usually childhood-onset lysoso­
that they represent Mendelian forms of the dis­ mal storage disease associated with a wide variety of
ease, which are relatively rare, can be tackled by organ manifestations, had a conspicuous tendency to
classic approaches of linkage analysis and posi­ develop PD in later life (Machaczka et al., 1999;
tional cloning, and can be modelled in different Tayebi et al., 2001). Gaucher’s disease is caused by
cellular and animal model systems, which is mutations in the gene for glucocerebrosidase, GBA,
invaluable for a better understanding of the mole­ which is located in chromosome 1q21. GBA is an
cular pathways leading to dopaminergic neurode­ enzyme of the ceramide pathway, and its deficiency
generation (Gasser, 2009b). leads to the excessive storage of its substrate,
glucosylceramide, within lysosomes of many differ­
ent cell types, including neurons and macrophages
Rare genetic variants causing or pre-disposing to PD (Grabowski, 2008). Following the initial reports of a
clinical association of Gaucher’s disease with PD it
While the genes described above are generally was observed that heterozygous, otherwise healthy,
thought of as high-penetrance disease-causing carriers of GBA mutations, that is the relatives of
genes, the genetic epidemiology of LRRK2­ Gaucher patients, also have an increased risk to
associated PD has already made clear that the develop PD. This association, which was initially
boundary between disease genes and risk factors observed in Ashkenazi Jewish families in whom the
is more a matter of semantics than of biology. carrier frequency for GBA mutations is particularly
10

high, was later confirmed in a number of larger develop a disease. Over the years, literally hun­
patient series from different Jewish and non-Jewish dreds of studies have been published, but unfortu­
populations (Aharon-Peretz et al., 2004; De Marco nately, only very few of them have produced
et al., 2008; Mata et al., 2008), in a study comprising robust and reproducible results. The reasons for
autopsy-proven cases (Goker-Alpan et al., 2006) the failure of this approach are manifold:
and, most recently, in a large meta-analysis
(Sidransky et al., 2009). This association is now 1. Most studies were greatly underpowered in
firmly established. For example, a recent study relation to the small increases in the relative risk
showed that the prevalence of GBA mutations in that today are known to be conferred by common
British patients with sporadic PD is 3.7%, while genetic variants (usually the odds ratios are in
the frequency of these variants is less than 1% in the range of 1.2–2, with some exceptions, for
the general population. Mutations in the GBA gene example apolipoprotein E for AD).
therefore are the most common risk factor for 2. The choice of candidate genes was often based
development of PD in this population detected so on rather arbitrary rationales with very weak
far (Neumann et al., 2009). The relative risk experimental or epidemiologic evidence. As the
conferred by heterozygous carrier status for the gene-mapping studies in monogenic diseases
development of PD varies, for different mutations have shown, newly identified genes most often
and in different studies, from about 4 to 20, with the could not have been predicted based on the
large meta-analysis suggesting a relative risk of current knowledge of pathogenesis.
about 4 for the N370S mutation, the most common 3. In most studies only arbitrarily chosen
variant allele in Ashkenazi Jews, and about 6 for individual genetic variants were interrogated;
the most common mutation in non-Jews, L444P thus it was a priori unlikely that the causative
(Sidransky et al., 2009). variant or a variant in high linkage
This estimation of modest but significant rela­ disequilibrium, tagging the risk-conferring
tive risks to develop the disease explains why so variant, would be among those studied.
far no large ‘GBA-families’ have been identified.
As far as it is known today there is no way to Finally, it is not easy to match patient and con­
distinguish PD patients with GBA mutations from trol cohorts with respect to their genetic back­
those without, both clinically and pathologically. ground. Often, due to different recruiting
As a group, GBA-positive patients have a slightly strategies, these cohorts differ in their genetic
earlier age of onset (55 vs. 59 years) and a some­ composition (a problem called undetected popula­
what higher prevalence of dementia (Sidransky tion stratification, which today can be easily
et al., 2009). Neuropathologic examination of 17 resolved in GWAS, see below). Due to different
GBA mutation carriers showed typical PD allele frequencies in different populations, spur­
changes, with widespread and abundant SNCA ious associations can be detected.
pathology, and most also had neocortical Lewy Due to these shortcomings in study design, so far
body pathology (Neumann et al., 2009). not a single association study result concerning PD
truly withstood the test of time and replication, with
the notable exceptions of candidate gene associa­
Common risk variants for PD: candidate gene and tion studies of the SNCA and MAPT genes, two
whole genome association studies genes that initially were identified by mapping
disease genes in monogenic families (see below).
Association studies have been widely used in an Technical advances in sequencing technologies
attempt to identify common genetic variations including a vast increase in speed and accuracy of
that carry a mild to moderately increased risk to genotyping individual sequence changes (SNPs)
 11

together with a rapid increase in knowledge about certain degree of linkage disequilibrium between
the haplotype structure of the human genome those regions, it was not possible to definitively
paved the way for a more systematic study of separate the signals. On the other hand, as a num­
genetic variability and its relationship to common ber of different regulatory mechanisms are likely
diseases such as PD. This is nicely exemplified to determine SNCA expression, it would not be
in the way that variability in SNCA and in the surprising if multiple regions of the gene were
gene for the microtubule-associated protein tau found to be involved.
(MAPT) is now recognized as the major genetic Since then a number of additional association
risk factors in sporadic PD. studies were able to replicate these results in dif­
Since aSYN aggregates are widely accepted to ferent populations (Mizuta et al., 2006; Winkler
be the hallmark neuropathologic change in PD et al., 2007). Most of them found 30 -variants to be
and the role of SCNA point mutations and gene most significantly associated with the disease.
multiplications is clearly established in familial Further evidence for a role of genetic variants in
PD, it was only obvious to search for a possible the SNCA gene came from GWAS (see below).
association of genetic polymorphisms in the The history of the association of MAPT, the
SNCA gene with sporadic PD. SNCA variability, gene encoding the microtubule-associated protein
putatively influencing the level of SNCA gene tau (MAPTau), is less straightforward. The gene
expression, became a particularly plausible candi­ has been studied as a candidate gene for neuro­
date after the discovery that multiplications of the degenerative diseases for many years. Initially,
SNCA locus and thus a gene dosage effect of the MAPTau was identified as the main component
wt aSYN were the cause for dominant PD. of paired helical filaments, the intracellular
Early studies had produced somewhat conflict­ hallmark neuropathology of AD (Goedert et al.,
ing results, because again, in hindsight, those stu­ 1988). Tau aggregates also form the major pathol­
dies were underpowered given the relatively small ogy in several other diseases, the so-called tauo­
odds ratios that are known today. Nevertheless, pathies, which include a subset of patients with a
the majority of association studies, including a dementia syndrome called frontotemporal lobar
large meta-analysis of studies interrogating the degeneration (FTLD), and also atypical parkinso­
‘NACP-REP1’ polymorphism, a complex repeat nian syndromes, such as progressive supranuclear
element located about 10 kb upstream of the palsy (PSP) and corticobasal degeneration (CBD)
SNCA coding region (Maraganore et al., 2006), (Williams, 2006). Point mutations in MAPT,
supported the assumption of a role of SNCA var­ which either affect its amino acid sequence or
iants in sporadic PD. Müller et al. performed the the splicing of its isoforms, were then identified
first systematic study of SNPs in the SNCA gene, to cause a monogenic form of FTLD, called
analyzing more than 50 variants distributed over FTLD-17 or, today, FTLD-Tau (Hutton et al.,
the entire gene in more than 600 patients and a 1998). Given the prominent tau-pathology in
similar number of controls, thereby capturing PSP and CBD, it was not surprising when it was
nearly all of the genetic variability of the gene. reported that genetic variability in MAPT, initi­
They found that the SNCA gene consists of two ally a dinucleotide repeat, was found to be asso­
major haplotype blocks: one comprising the pro­ ciated with these disorders (Conrad et al., 1997).
motor and exon 1–4 and another one spanning A closer study of this genetic region on chromo­
exons 5 and 6 and the 30 -untranslated region of some 17 revealed that this variant was part of an
the gene. The strongest association signal was extended haplotype, spanning about 1.5 Mb. Two
detected with SNPs in 30 -haplotype block, major forms of this haplotype exist, called H1 and
although a second, somewhat weaker signal in H2, with H1, which occurs on about 80% of Cau­
the promotor region was also detected. Due to a casian chromosomes, conferring the higher risk.
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