Parkin dosage mutations have greater
pathogenicity in familial PD than simple
N. Pankratz, PhD
D.K. Kissell, BS
M.W. Pauciulo, MBA
C.A. Halter, MS
A. Rudolph, PhD
R.F. Pfeiffer, MD
K.S. Marder, MD
T. Foroud, PhD
W.C. Nichols, PhD
For the Parkinson Study
Objective: Mutations in both alleles of parkin have been shown to result in Parkinson disease (PD).
However, it is unclear whether haploinsufficiency (presence of a mutation in only 1 of the 2 parkin
alleles) increases the risk for PD.
Methods: We performed comprehensive dosage and sequence analysis of all 12 exons of parkin in
a sample of 520 independent patients with familial PD and 263 controls. We evaluated whether
presence of a single parkin mutation, either a sequence (point mutation or small insertion/deletion)
or dosage (whole exon deletion or duplication) mutation, was found at increased frequency in
cases as compared with controls. We then compared the clinical characteristics of cases with 0,
1, or 2 parkin mutations.
Results: We identified 55 independent patients with PD with at least 1 parkin mutation and 9
controls with a single sequence mutation. Cases and controls had a similar frequency of single
sequence mutations (3.1% vs 3.4%, p ? 0.83); however, the cases had a significantly higher rate
of dosage mutations (2.6% vs 0%, p ? 0.009). Cases with a single dosage mutation were more
likely to have an earlier age at onset (50% with onset at ?45 years) compared with those with no
parkin mutations (10%, p ? 0.00002); this was not true for cases with only a single sequence
mutation (25% with onset at ?45 years, p ? 0.06).
Conclusions: Parkin haploinsufficiency, specifically for a dosage mutation rather than a point mu-
tation or small insertion/deletion, is a risk factor for familial PD and may be associated with earlier
age at onset. Neurology®2009;73:279–286
ADL ? Activities of Daily Living; GDS ? Geriatric Depression Scale; MLPA ? multiplex ligation-dependent probe amplifica-
tion; MMSE ? Mini-Mental State Examination; PD ? Parkinson disease; UPDRS ? Unified Parkinson’s Disease Rating Scale.
Parkinson disease (PD; MIM 168600) is the second most common neurodegenerative disease.
Mutations in 5 genes have been identified to segregate with PD in a mendelian fashion.
Alterations in 3, PRKN (parkin), DJ-1, and PINK1, are typically transmitted with autosomal
recessive inheritance, whereas alterations in the other 2, SNCA and LRRK2, are inherited in an
autosomal dominant fashion. Mutations in these 5 genes seem to be causative in less than 5%
More than 100 different parkin mutations have been reported and are found in all 12 exons
of the gene.2Point mutations as well as copy number variation (i.e., deletions and duplications
of entire exons, hereafter referred to as “dosage” changes) have been identified as pathogenic
mutations.3-11Harboring 2 parkin mutations typically leads to juvenile-onset (ages 15–25
years) or early-onset (?45 years) forms of PD.5,6Several previous studies have hypothesized
that haploinsufficiency, or the presence of a mutation in only 1 of 2 parkin alleles, may lead to
*Parkinson Study Group–PROGENI Investigators are listed in the appendix.
From the Indiana University Medical Center (N.P., C.A.H., T.F.), Indianapolis, IN; Cincinnati Children’s Hospital Medical Center (D.K.K.,
M.W.P., W.C.N.), OH; University of Rochester (A.R.), Rochester, NY; University of Tennessee Health Science Center (R.F.P.), Memphis, TN;
Columbia University and The Taub Institute (K.S.M.), New York, NY; and University of Cincinnati School of Medicine (W.C.N.), Cincinnati, OH.
Supported by NIH R01 NS37167 (N.P., D.K.K., M.W.P., C.A.H., A.R., R.F.P., K.S.M., T.F., W.C.N.) and M01 RR-00750.
Disclosure: Author disclosures are provided at the end of the article.
Address correspondence and
reprint requests to Dr. Tatiana
Foroud, Medical and Molecular
Genetics, Indiana University,
School of Medicine, Hereditary
Genomics Division, 410 W. 10th
St., MI-4000, Indianapolis, IN
Copyright © 2009 by AAN Enterprises, Inc.
an increased risk of PD. If true, this would
suggest that parkin may be both a causative
and a susceptibility gene.11-14
We have performed both sequencing and
dosage analysis of all parkin exons in a sample
of controls and sequentially recruited familial
PD cases, allowing us to obtain unbiased esti-
mates of the frequency of parkin mutations in
controls as well as cases with a family history
METHODS Subjects screened (n ? 520). PD families
were ascertained as part of an ongoing genetic study through a
pair of siblings, both of whom were reported to have PD. The
study was approved at each site’s institutional review board. The
first 520 families out of a total of 735 recruited were included in
the present study, and a single affected individual from each
family, typically the proband, was selected to be comprehen-
sively screened for parkin mutations. Results of parkin screening
for 127 of these individuals have been previously reported.9,11All
participants were seen by a movement disorder specialist at 1 of
59 Parkinson Study Group sites located throughout North
America. Each participant completed a uniform clinical assess-
ment that included the Unified Parkinson’s Disease Rating Scale
(UPDRS) Parts II (Activities of Daily Living) and III (Motor
Exam),15Schwab and England score,16Hoehn and Yahr stage,17
the Mini-Mental State Examination (MMSE),18the Geriatric
Depression Scale (GDS),19and the Blessed Functional Activity
Scale (Blessed).20In addition, a diagnostic checklist with inclu-
sion criteria consisting of clinical features highly associated with
autopsy-confirmed PD and exclusion criteria consisting of clini-
cal features highly associated with non-PD pathologic diagnoses
was used to classify individuals as having either verified PD
(90%) or nonverified PD (10%).21DNA was obtained from pe-
Cases used in association analyses (n ? 420). To avoid
bias from population stratification, association analyses were
limited to self-reported white, non-Hispanic individuals who
met criteria for verified PD and did not harbor a known LRRK2
mutation.22-24Of the 520 individuals screened, 29 were not of
white, non-Hispanic descent, 50 did not meet criteria for veri-
fied PD, and 33 had previously been identified to harbor an
LRRK2 mutation.22-24Some PD cases fulfilled more than 1 of
these exclusion criteria. The final analyzed data set included 420
Controls screened (n ? 263). The control sample consisted
of 270 neurologically normal individuals obtained from the Na-
tional Institute of Neurological Disorders and Stroke Human
Genetics DNA and Cell Line Repository (Camden, NJ; DNA
panels NDPT002, NDPT006, NDPT009). Data from a prior
genome-wide association study verified that these individuals
were unrelated.25Individuals not of white, non-Hispanic descent
(n ? 2) and controls reporting a family history of PD in a first or
second degree relative (n ? 5) were excluded. All controls pro-
vided appropriate written informed consent for storage and fu-
ture use of their samples. The final analyzed data set included
263 independent controls.
Parkin sequencing. PCR and sequencing primers were de-
signed using the chromosome 6 genomic contig sequence
NT_007422.13 enabling PCR/sequencing of all 12 coding ex-
ons and corresponding intron/exon boundaries of parkin. Typi-
cally, 80 ng of each genomic DNA was PCR amplified in a
40-?L reaction using conditions empirically determined for each
primer pair as previously described.26The resulting PCR prod-
ucts were purified using the QIAquick 96 PCR purification kit
(QIAGEN, Santa Clara, CA) and sequenced on an ABI 3730
DNA analyzer using the Applied Biosystems (Foster City, CA)
BigDye Terminator Kit, version 1.1. DNA sequences were
aligned and analyzed using Sequencher 4.5 (Gene Codes Corpo-
ration, Ann Arbor, MI). All missense mutations were analyzed
using SIFT (http://blocks.fhcrc.org/sift/SIFT.html) to predict in
silico whether the amino acid change would be deleterious or
Parkin dosage analysis using MLPA. Multiplex ligation-
dependent probe amplification (MLPA) was performed with
100 ng of genomic DNA according to manufacturer’s instruc-
tions using the P051 and P052 Salsa MLPA Parkinson probe sets
(MRC-Holland, Amsterdam, The Netherlands). This probe set
includes probes for all parkin exons, and 10 of the 12 exons have
a duplicate probe. Probe amplification products were run on an
ABI 3730xl DNA Analyzer using GS500 size standard (Applied
Biosystems). MLPA peak plots were visualized using Genemap-
per Software version 3.7 (Applied Biosystems), and nonnormal-
ized values for peak height and peak area were then exported to
an Excel template. Normalization of data and calculation of dos-
age ratios was performed as described at www.mrc-holland.com/
MLPA%20analysis.htm. A dosage ratio value of ?0.7 was used
as the boundary for deletions, and ?1.35 was used as the bound-
ary for duplications.
Parkin dosage analysis using real-time PCR. All dele-
tions and duplications identified using MLPA were verified us-
ing real-time PCR. Applied Biosystems’ Assay by Design service
was used to design fam-labeled TaqMan gene expression assays
for each exon of parkin. Genomic DNA samples were quanti-
tated by Pico Green fluorescence in triplicate with the Quant-iT
PicoGreen dsDNA Kit (Molecular Probes, Eugene, OR). After
quantitation, 50 ng of genomic DNA was used in a real-time
absolute quantitation assay for the parkin exon in question, per-
formed on the 7300 Real Time PCR System (Applied Biosys-
tems). Assays were performed as 25-?L reactions in triplicate,
with each genomic DNA sample being done in duplicate for the
parkin exon in question. After real-time PCR, data were analyzed
with the ABI Sequence Detection Software, RQ Study upgrade,
version 1.2.3 (Applied Biosystems). Quantitation of the target
amount in DNA samples was accomplished by measuring the Ct
value and comparing it with a standard curve. Quantitation for
study samples was then compared with those of control samples
known not to have exonic deletions/duplications.
Statistical analysis. The common variants S167N, V380L,
and D394N, previously shown to be benign polymor-
phisms,5,27,28were not coded as mutations in these analyses.
The Fisher exact test was used to evaluate the first hypothe-
sis: the presence of a single parkin mutation increased the risk of
PD. Analyses were also performed classifying the single parkin
mutation as either a dosage or sequence mutation and tested
whether the presence of each of these different types of muta-
tions individually increased the risk of PD.
The second hypothesis tested was that individuals with
parkin mutations (either 1 or 2) had different clinical charac-
teristics (i.e., age at onset, UPDRS subscores, MMSE, GDS,
Hoehn and Yahr stage) compared with those cases without a
Neurology 73July 28, 2009
parkin mutation. The groups with 2 parkin mutations and 1
parkin mutation were compared independently with the con-
trol group. Linear and logistic regression models were used,
RESULTS A total of 520 consecutive, independent,
familial PD cases were screened for parkin mutations
using both direct sequencing and MLPA analysis (ta-
bles 1 and 2). Mutations, defined as any variant
found in ?1% of all control chromosomes and that
change the amino acid sequence, were identified in
55 patients (10.6% of familial PD cases) (table 3).
The percentage of individuals with parkin mutations
was higher when symptoms began in earlier decades:
33% for those with onset between ages 11 and 20
years, 47% for onset between 21 and 30 years, 52%
for onset between 31 and 40 years, 14% for onset
between 41 and 50 years, 2% for onset between 51
and 60 years, 7% for onset between 61 and 70 years,
and 5% for onset at age ?71 years (tables e-1 and e-2
on the Neurology®Web site at www.neurology.org).
Of those harboring at least 1 parkin mutation, 27
had a mutation in both alleles (11 homozygous, 16
compound heterozygous), and 28 had a mutation de-
tected in only 1 of the 2 alleles (12 dosage mutations,
16 sequence mutations). The most common muta-
tion detected in our PD cohort was the cis deletion of
exons 3–4 (13% of observed mutations), followed by
P437L (11%) and R275W (10%). All other muta-
tions each represented less than 10% of the total
number of mutations observed (tables e-1, e-2, 2,
Nine of the 263 controls were heterozygous carri-
ers of a single parkin sequence mutation (table 2).
The most common mutations detected in the control
sample were A82E, R275W, and P437L, with 2 in-
stances of each. MLPA analysis yielded no parkin
dosage changes in the controls (tables e-1 and e-2).
Controls carrying a parkin variant did not have a sig-
Table 1Sample demographics
Mean age at onset for
cases or examination
for controls (range) % Male
PROGENI (all samples screened for
Cases520 59.3 (18–88)58.1
PROGENI (white non-Hispanic
with verified PD and no LRRK2
Cases 42060.3 (21–84)59.5
NINDS Human Genetics Resource
Controls263 69.5 (55–88)48.3
PD ? Parkinson disease; NINDS ? National Institute of Neurological Disorders and Stroke.
Table 2Sequence mutations identified*
No. alleles/No. chromosomes (frequency)
Exon Nucleotide changeProtein change SIFT predictionCases Controls
c.98G?A R33QBenign 1/1,040 (0.001)0/526 (0)
c.101-102delAGQ34fs (?4 amino acids) 4/1,040 (0.004)1/526 (0.002)
c. 155delAN52fs (?28 amino acids) 3/1,040 (0.003)0/526 (0)
c. 235G?T E79X1/1,040 (0.001)0/526 (0)
c.245C?AA82E Benign 1/1,040 (0.001)2/526 (0.004)
c.247A?GT83A† Benign 0/1,040 (0)1/526 (0.002)
c.337-376delP113fs (?51 amino acids)6/1,040 (0.006)0/526 (0)
c.400insCCAP133dup† 1/1,040 (0.001)0/526 (0)
c.518C?T T173M Deleterious 1/1,040 (0.001) 0/526 (0)
IVS4 535G -9T?A† 2/1,040 (0.002)0/526 (0)
c.574A?G M192V Benign 1/1,040 (0.001)0/526 (0)
c.633A?T K211NDeleterious 1/1,040 (0.001)0/526 (0)
c.718C?T T240MBenign 2/1,040 (0.002)0/526 (0)
c.766C?TR256C Benign1/1,040 (0.001) 1/526 (0.002)
c.823C?TR275W Deleterious8/1,040 (0.008) 2/526 (0.004)
c.930G?CE310D Benign1/1,040 (0.001) 0/526 (0)
IVS9 1083?1delG† 1/1,040 (0.001)0/526 (0)
IVS9 1083?5G?T 1/1,040 (0.001) 0/526 (0)
c.1289G?A G430D Deleterious1/1,040 (0.001) 0/526 (0)
c.1310C?T P437L Benign9/1,040 (0.009)2/526 (0.004)
*Among 520 familial Parkinson disease (PD) cases and 263 controls screened.
†A novel, not previously reported mutation.
Neurology 73 July 28, 2009
nificantly earlier age at examination (69.4 years vs
69.5 years for those not harboring a parkin muta-
tion); however, those carrying a variant predicted to
be deleterious by the program SIFT (table 2) tended
to be younger at examination (57, 60, 73 years)
Within the 420 unrelated white, non-Hispanic,
familial PD cases, the frequency of mutations in only
1 of the 2 parkin alleles was 5.7% (24 PD cases), with
3.1% having a single change detected by sequence
analysis and 2.6% having a single dosage change
identified by MLPA (table 4). In comparison, only 9
controls carried a parkin mutation (3.4% of con-
trols), with none being a dosage mutation. Overall,
cases were not more likely than controls to carry a
mutation in only 1 of 2 parkin alleles (5.7% vs 3.4%,
p ? 0.20). However, when the type of parkin muta-
tion was compared, there was a higher frequency of a
single parkin dosage mutation in the cases than in the
controls (2.6% vs 0%, p ? 0.009). The frequency of
each individual sequence and dosage mutation was
too low to provide sufficient power to test for associ-
ation with each mutation separately.
The clinical features of the parkin mutation-
positive and mutation-negative PD individuals were
remarkably similar (table 5). The core symptoms of
PD (bradykinesia, rest tremor, muscular rigidity, and
postural instability) were all found at comparable
rates. There was no difference in the proportion of
individuals classified as verified PD (87.3% of muta-
tion carriers vs 90.8% of noncarriers, p ? 0.46). PD
cases with a single dosage mutation were more likely
to have an earlier age at onset (?45 years) compared
with those with no parkin mutations (p ? 0.00002),
but the same could not be said for those with only a
single sequence mutation (p ? 0.06). Individuals
with 2 mutations had a significantly earlier onset
than those with only 1 mutation (p ? 0.0008),
though this too was different for sequence (p ?
0.001) and dosage mutations (p ? 0.02). No other
clinical feature (other than ethnicity) differed be-
tween those that carried 2 mutations vs 1 mutation.
As shown previously,29there was no association be-
tween harboring 1 or 2 parkin mutations and depres-
sive symptomatology (GDS score ? 10) when
individuals showing signs of cognitive impairment
(MMSE ? education-specific threshold) were ex-
cluded from analysis (p ? 0.19).
DISCUSSION Interpreting the clinical significance
of parkin mutations has been problematic because of
the large number of different mutations that have
been identified in the gene.1Truncating or frame-
shift mutations likely lead to a loss of enzyme func-
tion, whereas the effect of missense, synonymous and
variants near splice sites may not have as obvious an
effect on enzyme function. It has not been conclusively
shown whether a mutation on only 1 of the 2 alleles
affects the risk for PD.11-14,27Here we provide evidence
are more likely to increase risk than others. Specifically,
a single copy of a deletion or duplication was found in
have onset before age 45 years (50% compared with
no more likely to have onset before age 45 years than
those without a mutation (25% vs 10%, p ? 0.06).
These data suggest that dosage changes in parkin play a
larger role in disease susceptibility than do sequence
Table 3Dosage mutations identified*
No. alleles/No. chromosomes (frequency)
ExonAlteration Cases Controls
Deletion 2 4/1,040 (0.004)0/526 (0)
Deletion 2–3 4/1,040 (0.004)0/526 (0)
Duplication 2 1/1,040 (0.001)0/526 (0)
Duplication 2–4 2/1,040 (0.002)0/526 (0)
Triplication 2–41/1,040 (0.001)0/526 (0)
Deletion 3–411/1,040 (0.011)0/526 (0)
Deletion 3–61/1,040 (0.001)0/526 (0)
Deletion 4 1/1,040 (0.001)0/526 (0)
Deletion 4–51/1,040 (0.001)0/526 (0)
Deletion 4–6 1/1,040 (0.001)0/526 (0)
Duplication 4 2/1,040 (0.002)0/526 (0)
Deletion 51/1,040 (0.001)0/526 (0)
Deletion 5–6 2/1,040 (0.002) 0/526 (0)
Duplication 5–8 1/1,040 (0.001)0/526 (0)
Deletion 62/1,040 (0.002) 0/526 (0)
Duplication 71/1,040 (0.001)0/526 (0)
Deletion 8–9 1/1,040 (0.001)0/526 (0)
*Among 520 familial Parkinson disease cases and 263 controls screened.
Table 4 Association between parkin carrier status and Parkinson disease
(n ? 840
(n ? 526
Two parkin mutations
17 (4.0%)0 (0.0%) 0.0005
One parkin mutation
24 (5.7%)9 (3.4%) 0.20
13 (3.1%) 9 (3.4%)0.83
11 (2.6%)0 (0.0%)0.009
*Analyses were limited to 420 white non-Hispanic Parkinson disease cases without a
known LRRK2 mutation.
Neurology 73July 28, 2009
There is also heterogeneity of effect with regard to
missense mutations. At one end of the spectrum are
several common variants previously shown to be be-
nign polymorphisms (S167N, V380L, D394N)5,27,28
that were therefore excluded from analysis in the cur-
rent study. Other observed variants included in our
analyses may also be benign, but they are found at
such a low frequency that it is difficult to draw firm
conclusions about their pathogenicity. One such
variant, P437L, is predicted by the program SIFT to
be benign. This variant is found in 6 independent
cases within this familial cohort, and the 3 PD cases
that were homozygous for the variant tended to have
a later age at onset (ages 45, 70, 81 years; table 2). If
the allele is pathogenic, it would seem to result in
later onset PD as compared with other missense mu-
tations. When P437L and all other variants catego-
rized as “benign” by the SIFT algorithm were
excluded from analyses, the same conclusions are
reached—namely, single sequence mutations are
equally common in cases and controls (p ? 0.75),
single sequence mutations are not associated with
early age at onset (22% with onset at ?45 years com-
pared with 10% among those without a parkin muta-
tion; p ? 0.26), and single dosage changes are more
common in cases than controls (p ? 0.009). How-
ever, with benign mutations excluded and the 2 types
of mutations (sequence and dosage) combined, cases
had a higher frequency of a single parkin mutation
than controls (p ? 0.02). These results suggest that
the variants predicted by SIFT as being benign are
likely to be correct; however, additional studies are
required to confirm the in silico predictions.
Individuals with 2 parkin mutations tended to
have earlier onset and consequently tended to have a
longer duration of disease. Significant differences in
other clinical characteristics were not seen in this
study. Hispanics made up a disproportionately
higher percentage of the individuals with 2 parkin
mutations as compared with their representation in
Table 5 Comparison of the clinical characteristics of the parkin mutation groups with the no parkin
27 1216 465
Age at onset
40.51 ? 10?7
50.9 0.0456.2 0.3260.7
Early onset (<45 y)
77.8%1 ? 10?22
50.0% 0.0000225.0% 0.06 10.3%
20.9 0.000114.8 0.2110.40.67 9.5
81.5%0.11 100.0%0.27 87.5% 0.6690.8%
77.8%0.0001 100.0%0.44 93.8%0.7895.3%
18.5%0.00001 0.0%0.57 6.2% 0.372.6%
At least 1 parent reported to have
25.9%0.24 25.0%0.3931.2% 0.6337.2%
LRRK2 G2019S carrier
0.0% 0.308.3% 0.520.0%0.394.4%
Any LRRK2 mutation carrier
0.0% 0.25 8.3% 0.680.0%0.335.6%
Hoehn and Yahr stage
2.6 0.40 2.4 0.642.3 0.542.5
Blessed Functionality score
3.30.25 2.70.16 184.108.40.206
UPDRS Part II (ADL)
14.00.74 12.30.57 12.8 0.7313.5
UPDRS Part III (motor)
27.0 0.9226.5 0.96 26.60.97 26.7
28.20.009 27.50.17 28.0 0.10 26.7
7.1% 0.04 7.5% 0.348.1% 0.569.0%
Hallucination with drugs†
11.1% 0.0333.3%0.84 37.5%0.56 30.6%
Hallucinations without drugs†
11.1% 0.508.3% 0.92 0.0% 0.257.5%
100.0%0.38 100.0%0.54 93.8% 0.4797.0%
100.0%0.16 100.0% 0.3393.8% 0.8792.6%
76.0%0.17 58.3%0.78 68.8%0.60 62.3%
80.0%0.90 66.7%0.31 81.2%0.82 78.9%
*Among 520 familial Parkinson disease cases.
PD ? Parkinson disease; UPDRS ? Unified Parkinson’s Disease Rating Scale; ADL ? Activities of Daily Living; MMSE ?
Mini-Mental State Examination; GDS ? Geriatric Depression Scale.
Neurology 73 July 28, 2009
the rest of the sample (12.1% of cases with 2 parkin
mutations vs 2.4% of cases with no parkin muta-
tions, p ? 0.00001). All 6 Hispanic cases harboring
parkin mutations were from Puerto Rico. Particular
alleles were more common in the Puerto Rican cases; of
the 12 parkin alleles observed, 4 were cis deletions of
2–3, and 1 was the wild-type allele. If parkin mutations
are in fact more common in Puerto Rico or in the
greater Hispanic population at large, it is especially im-
portant that any genetic association analyses be per-
formed within a single population to avoid bias due to
population stratification. Overrepresentation of parkin
mutations in Hispanic populations has been noted pre-
viously,30and the cis deletion of exons 3 and 4 was
found to be common among Puerto Rican cases.2
One subject with PD harbored both a heterozy-
gous deletion of exon 2 and an LRRK2 G2019S allele
(table 5). This individual met criteria for verified PD
and had onset at age 41 years. One sibling of this
individual had onset at 68 years and had the same 2
mutant alleles at LRRK2 and parkin, whereas the
other sibling had onset at 83 years and inherited only
the G2019S allele, suggesting that the mutant parkin
allele contributed to earlier disease onset.
In this study, we have used MLPA to screen for
whole exon deletions and duplications and con-
firmed every dosage change using real-time PCR.
Previously, we used semiquantitative fluorescent
multiplex PCR to identify parkin dosage muta-
tions.9,11Several dosage changes reported in our pre-
vious studies9,11did not replicate using the newer
more robust methods, including what seemed to be a
common deletion in exon 8. We have now deter-
mined that one of the primers used to amplify this
region annealed to a region containing a previously
unknown SNP, which resulted in the erroneously
observed exon deletion. In contrast, only 2 muta-
tions flagged by MLPA (deletion of exon 4, duplica-
tion of exon 9) did not validate using real-time PCR.
Phase could be determined conclusively for 89%
of individuals (including all heterozygous rearrange-
ments involving consecutive exons), and identity by
descent estimates from prior linkage data provided
supportive evidence for an additional 8% of individ-
uals (tables e-1 and e-2). Should any of the inconclu-
sive compound heterozygotes with dosage mutations
prove to have both variants on a single allele, this
would only strengthen the evidence that heterozy-
gous dosage mutations are associated with disease.
In the past, most studies of parkin have focused
on either sporadic or early-onset cases. Our study is
relatively unique in its ascertainment of familial PD,
and it is therefore likely to be enriched for a genetic
etiology. In addition, we have not limited our screen-
ing to cases below an age at onset threshold. For this
reason, we were able to show that parkin mutations
are present in individuals with an older age at onset
(tables e-1 and e-2). Therefore, a mutation in the
parkin gene should not be ruled out when evaluating
patients with a later age at onset, especially if they
have a family history of PD.
We have completed the largest study to perform
both sequencing and dosage analysis in cases and
controls. Given the high frequency of exon rear-
rangements in our sample, it is apparent that thor-
ough screening of the parkin gene must include
dosage studies. In this study, we were able to show
similar between cases and controls (even when variants
predicted to be benign were excluded), cases are much
more likely to harbor dosage changes, indicating that
this type of mutation may be more pathogenic.
All statistical analyses were performed by N. Pankratz.
Control samples and clinical data were provided from the National Insti-
tute of Neurological Disorders and Stroke Human Genetics DNA and
Cell Line Repository (http://ccr.coriell.org/ninds). The authors thank Dr.
Lorraine Clark for her helpful suggestions in drafting the manuscript. The
authors thank all subjects for their participation in this research study.
Dr. Marder serves as an editorial board member of Neurology®. Dr.
Pfeiffer serves on the Center Review Board of the National Parkinson
Foundation; serves as Co-editor-in-Chief of Parkinsonism and Related
Disorders; receives royalties from publishing Neurogastroenterology [But-
terworth Heinemann(Elsevier) 2008], Parkinson’s Disease [CRC Press
(Taylor & Francis) 2008], and Parkinson’s Disease and Nonmotor Dys-
function (Humana Press, 2008); has served on an external advisory com-
mittee for the Udall Center; has received speaker honoraria from the
Parkinson Association of Southwest Florida, the Black Hills Neurology
Clinic, Mayo Clinic Jacksonville, Jackson-Madison County Memorial
Hospital, Indian Physicians Medical Group (Memphis, TN), and Yale
University; serves on scientific advisory boards for Kyowa, Solvay, and
Ipsen; serves on speakers’ bureaus of GlaxoSmithKline, Boehringer-
Ingelheim, Novartis, Teva, and UCB/Schwarz; receives research support
from Boehringer-Ingelheim, Kyowa, Novartis, Eisai, UCB/ Schwarz, and
Santhera; and has served as an expert consultant to law firms Spriggs &
Hollingsworth and Davis Graham & Stubbs.
Parkinson Study Group Investigators: PROGENI Steering Committee:
University of Tennessee Health Science Center: R.F. Pfeiffer (Chair);
University of Rochester: F. Marshall, D. Oakes, A. Rudolph, A. Shina-
man; Columbia University Medical Center: K. Marder; Indiana Univer-
sity School of Medicine: P.M. Conneally, T. Foroud, C. Halter;
University of Kansas Medical Center: K. Lyons; Eli Lilly and Company:
E. Siemers; Medical College of Ohio: L. Elmers; University of California,
Irvine: N. Hermanowicz.
PSG-PROGENI Investigators and Coordinators: Albany Medical College:
S. Factor, D. Higgins, S. Evans; Barrow Neurological Institute: H. Shill,
M. Stacy, J. Danielson, L. Marlor, K. Williamson; Baylor College of Med-
icine: J. Jankovic, C. Hunter; Beth Israel Deaconess Medical Center: D.
Simon, P. Ryan, L. Scollins; Beth Israel Medical Center: R. Saunders-
Pullman, K. Boyar, C. Costan-Toth, E. Ohmann; Brigham and Women’s
Neurology 73July 28, 2009
Hospital: L. Sudarsky, C. Joubert; Brown University (Memorial Hospital
of Rhode Island): J. Friedman, K. Chou, H. Fernandez, M. Lannon;
Cleveland Clinic Florida-Weston: N. Galvez-Jimenez, A. Podichetty, K.
Thompson; Clinical Neuroscience Center: P. Lewitt, M. DeAngelis; Col-
orado Neurological Institute: C. O’Brien, L. Seeberger, C. Dingmann, D.
Judd; Columbia University Medical Center: K. Marder, J. Fraser, J. Har-
ris; Creighton University: J. Bertoni, C. Peterson; Evanston Northwestern
Healthcare: M. Rezak, G. Medalle; Hotel-Dieu Hospital-Chum: S.
Chouinard, M. Panisset, J. Hall, H. Poiffaut; Hunter Homes McGuire
Veterans Medical Center: V. Calabrese, P. Roberge; Indiana University
School of Medicine: J. Wojcieszek, J. Belden; Institute for Neurodegen-
erative Disorders: D. Jennings, K. Marek, S. Mendick; Johns Hopkins
University: S. Reich, B. Dunlop; London Health Sciences Centre: M. Jog,
C. Horn; Mayo Clinic Jacksonville: R. Uitti, M. Turk; McFarland Neu-
rosciences: T. Ajax, J. Mannetter; Medical College of Georgia: K. Sethi,
J. Carpenter, B. Dill, L. Hatch, K. Ligon, S. Narayan; Medical College of
Wisconsin: K. Blindauer, K. Abou-Samra, J. Petit; Medical University of
Ohio: L. Elmer, E. Aiken, K. Davis, C. Schell, S. Wilson; Mount Sinai
School of Medicine: M. Velickovic, W. Koller (deceased), S. Phipps;
North Shore-LIJ Health System: A. Feigin, M. Gordon, J. Hamann, E.
Licari, M. Marotta-Kollarus, B. Shannon, R. Winnick; Northwestern
University: T. Simuni, A. Videnovic, A. Kaczmarek, K. Williams, M.
Wolff; Ochsner Clinic Foundation: J. Rao, M. Cook; Ohio State Univer-
sity: M. Fernandez, S. Kostyk, J. Hubble, A. Campbell, C. Reider, A.
Seward; Oregon Health and Science University: R. Camicioli, J. Carter, J.
Nutt, P. Andrews, S. Morehouse, C. Stone; Ottawa Hospital Civic Site:
T. Mendis, D. Grimes, C. Alcorn-Costa, P. Gray, K. Haas, J. Vendette;
Pacific Neuroscience Medical Group: J. Sutton, B. Hutchinson, J. Young;
Saskatoon District Health Board Royal University Hospital: A. Rajput, A.
Rajput, L. Klassen, T. Shirley; Scott and White Hospital/Texas A&M
University: B. Manyam, P. Simpson, J. Whetteckey, B. Wulbrecht; The
Parkinson’s and Movement Disorder Institute: D. Truong, M. Pathak, K.
Frei, N. Luong, T. Tra, A. Tran, J. Vo; Toronto Western Hospital, Uni-
versity Health: A. Lang, G. Kleiner-Fisman, A. Nieves, L. Johnston, J. So;
UMDNJ-School of Osteopathic Medicine: G. Podskalny, L. Giffin; Uni-
versity of Alabama at Birmingham: P. Atchison, C. Allen; University of
Alberta: W. Martin, M. Wieler; University of Calgary: O. Suchowersky,
M. Klimek; University of California Irvine: N. Hermanowicz, S. Nis-
wonger; University of California San Diego: C. Shults (deceased), D.
Fontaine; University of California San Francisco: M. Aminoff, C. Chris-
tine, M. Diminno, J. Hevezi; University of Chicago: A. Dalvi, U. Kang, J.
Richman, S. Uy, J. Young; University of Cincinnati: A. Dalvi, A. Sahay,
M. Gartner, D. Schwieterman; University of Colorado Health Sciences
Center: D. Hall, M. Leehey, S. Culver, T. Derian; University of Connect-
icut: T. Demarcaida, S. Thurlow; University of Iowa: R. Rodnitzky, J.
Dobson; University of Kansas Medical Center: K. Lyons, R. Pahwa, T.
Gales, S. Thomas; University of Maryland School of Medicine: L. Shul-
man, S. Reich, W. Weiner, K. Dustin; University of Miami: K. Lyons, C.
Singer, W. Koller (deceased), W. Weiner, L. Zelaya; University of Minne-
sota: P. Tuite, V. Hagen, S. Rolandelli, R. Schacherer, J. Kosowicz; Uni-
versity of New Mexico: P. Gordon, J. Werner; University of Puerto Rico
School of Medicine: C. Serrano, S. Roque; University of Rochester: R.
Kurlan, D. Berry, I. Gardiner; University of South Florida: R. Hauser, J.
Sanchez-Ramos, T. Zesiewicz, H. Delgado, K. Price, P. Rodriguez, S.
Wolfrath; University of Tennessee Health Science Center: R. Pfeiffer,
L. Davis, B. Pfeiffer; University of Texas Southwestern Medical Center:
R. Dewey, B. Hayward, A. Johnson, M. Meacham, B. Estes; Wake Forest
University School of Medicine: F. Walker, V. Hunt, C. O’Neill; Wash-
ington University: B. Racette, L. Good, M. Rundle.
Biostatistics and Clinical Trials Coordination Centers Staff: A. Watts, A.
Wang, T. Ross, S. Bennett, D. Kamp, E. Julian-Baros, S. Daigneault, R.
Received January 14, 2009. Accepted in final form April 17, 2009.
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