Organization of the human synphilin-1 gene, a candidate for
* Tracy Wanner,
John J. Kleiderlein,
Russell L. Margolis,
Christopher A. Ross
Department of Psychiatry, Johns Hopkins University School of Medicine, 720 Rutland Avenue, Ross Research Building, Room 618,
Baltimore, Maryland 21205-2196, USA
Brain Research Institute, Niigata University, Niigata, Japan
Department of Neuroscience, Johns Hopkins University, Baltimore, Maryland 21205, USA
Program in Cellular and Molecular Medicine, Johns Hopkins University, Baltimore, Maryland 21205, USA
Received: 21 September 1999 / Accepted: 16 March 2000
Abstract. We have recently identified a protein we called syn-
philin-1, which interacts in vivo with alpha-synuclein. Mutations in
alpha-synuclein cause familial Parkinson’s disease (PD). Alpha-
synuclein protein is present in the pathologic lesions of familial
and sporadic PD, and diffuse Lewy body disease, indicating an
important pathogenic role for alpha-synuclein. Here we describe
the structure of the human synphilin-1 gene (SNCAIP). The open
reading frame of this gene is contained within ten exons. We have
designed primers to amplify each SNCAIP exon, so these primers
can now be used to screen for mutations or polymorphisms in
patients with Parkinson’s disease or related diseases. We found a
highly polymorphic GT repeat within intron 5 of SNCAIP, suitable
for linkage analysis of families with PD. We have mapped
SNCAIP locus to Chromosome (Chr) 5q23.1-23.3 near markers
WI-4673 and AFMB352XH5. In addition, using immunohisto-
chemistry in human postmortem brain tissue, we found that syn-
philin-1 protein is present in neuropil, similar to alpha-synuclein
protein. Because of its association with alpha-synuclein, syn-
philin-1 may be a candidate for involvement in Parkinson’s disease
or other related disorders.
Parkinson’s disease (PD) is a neurodegenerative disorder charac-
terized by bradykinesia, rigidity, and tremor. At post-mortem ex-
amination, brains from PD patients have loss of neurons in the
dopamine-synthesizing neurons of the substantia nigra, and sur-
viving neurons develop a characteristic inclusion body called the
Lewy body (Forno 1996; Hardy and Gwinn-Hardy 1998; Olanow
and Tatton 1999; Dunnett and Bjo¨rklund 1999). While most cases
of Parkinson’s disease are sporadic, a subset of cases is familial.
Several genes have been identified whose mutations cause familial
Parkinson’s disease, including alpha-synuclein, parkin, and UCH-
L1 (Polymeropoulos et al. 1997; Kru¨ger et al. 1998; Leroy et.
1998; Kitada et al. 1998; Hattori et al. 1998; Abbas et al. 1999).
However, other families with Parkinson’s disease do not have
mutations in any of these genes (Scott et al. 1997, Gasser et al.
1998; Farrer et al. 1998; Dunnett and Bjo¨rklund 1999).
Study of pathologic material from PD patients has indicated
that alpha-synuclein is a prominent component of Lewy bodies in
both familial and sporadic cases of PD and also in diffuse Lewy
body disease (DLB) (Spillantini et al. 1997, 1998; Wakabayashi et
al. 1997, 1998; Mezey et al. 1998). In a previous study we have
characterized synphilin-1 as a protein, which associates with al-
pha-synuclein (Engelender et al. 1999). Synphilin-1 is a cytoplas-
mic protein with several ankyrin-like repeats, a coil-coiled domain,
and an ATP, GTP-binding domain. Co-transfection of cells in
culture with synuclein and synphilin-1 yielded cytoplasmic inclu-
sion bodies similar to Lewy bodies (Engelender et al. 1999).
We have now defined the intron-exon structure of the human
synphilin-1 (SNCAIP) gene and identified primers spanning each
exon, which can be used to screen for polymorphisms in patient
material. We have further refined the chromosomal localization of
the SNCAIP gene. We have also found that synphilin-1 protein is
present in neuropil in normal human brain, a distribution very
similar to that of alpha-synuclein. These data provide the rationale
and the tools for a search for mutations in SNCAIP in PD and
Materials and methods
PAC genomic library screening.
A human PAC genomic library was
screened with clone 1 DNA, nucleotides 181–1732 of the human SNCAIP
cDNA (Engelender et al. 1999). Screenings were done by plaque hybrid-
ization by using high-stringency conditions. PAC DNAs were isolated with
the QIAfilter Plasmid Maxi Kit (Qiagen) according to manufacturer’s in-
PAC DNA sequences were determined by the dideoxy ter-
mination method with an ABI automated sequencing apparatus (Johns
Hopkins University Core Genetics Facility).
PCR reactions were carried out in a volume of 50 l with an initial
denaturation at 96°C for 2 min, followed by 30 cycles at 96°C for 1 min,
annealing at 55°C for 1 min, extension at 72°C for 1 min and 30 s, and a
final extension of 10 min at 72°C.
Radiation hybrid screening.
SNCAIP human-specific primers (sense:
5Ј-CATCAGGGGGACGCAGGTTT-3Ј; antisense: 5Ј-TGTGGAGG-
TCCGCTGGAGAG-3Ј) were used to amplify by PCR human-hamster
chromosome-specific hybrid cell lines from the Genebridge4 panel (Re-
search Genetics). PCR reactions were performed at an annealing tempera-
ture of 55°C for 30 cycles. Chromosomal location of SNCAIP gene was
estimated according to manufacturer’s instructions.
To amplify the GT repeat region of the
SNCAIP locus, PCR reactions were performed at an annealing temperature
GenBank accession number: Synphilin-1 gene: AF 167301–AF 167306.
* Present address: Departamento de Anatomia, Universidade Federal do
Rio de Janeiro, Ilha do Fundao, Brazil. Fax: (+5521) 270-8647; E-mail:
Correspondence to: C.A. Ross; E-mail: firstname.lastname@example.org
Mammalian Genome 11, 763–766 (2000).
© Springer-Verlag New York Inc. 2000