2009 Vienna - European Society of Human Genetics

Molecular basis of Mendelian disorders
identification of the c.1374+654C>G variant, which was located deep
within intron 13. In silico analyses suggested that this mutation leads
to the creation of a strong donor splice site. Analyses of mRNA derived
from patient cell lines indicated that 2 mutant isoforms, both containing
parts of intron 13, are synthesized from the PRPF31 allele carrying
this mutation. These mRNAs harbour premature termination codons,
and were shown to be present in reduced levels in patient cell lines
due to their degradation by the nonsense-mediated mRNA decay. Protein
analysis revealed a decrease in the amount of full length PRPF31
protein and the lack of mutant proteins in patient cell lines. Our results
indicate that this mutation is pathogenic and, as with the vast majority
of PRPF31 mutations described so far, leads to the reduction of functional
PRPF31 protein and, consequently, that haploinsufficiency is the
cause of retinitis pigmentosa in the studied family.
P12.142
Novel mutation in RP2 gene in Russian family with X-linked
retinitis pigmentosa type 2
A. V. Polyakov 1 , O. V. Khlebnikova 1,2 , S. V. Gudzenko 1 , N. A. Beklemitcheva 1 ;
1 1- Research Centre for Medical Genetics, Russian Academy of Medical Sciences,
Moscow, Russian Federation, 2 2- Moscow Research Institute of Eye
Diseases, Moscow, Russian Federation.
X-linked retinitis pigmentosa type 2 (RP2) - is a severe form of congenital
X-linked retinal degeneration, characterized by constriction of
the visual fields, night blindness and fundus changes, including ‘bone
corpuscle’ lumps of pigment with a severely reduced visual acuity outcome.
RP2 gene, responsible for X-linked retinitis pigmentosa type 2, is located
on Xp11.3, consist of five exons and encodes a protein of the
same name RP2 which links the cell membrane with the cytoskeleton
in photoreceptors of the eye.
The purpose of our study was searching for a disease-causing mutation
in Russian family with RP2. The penetrance among female carriers
is incomplete in the family. Among 28 persons of a four-generation
family, in which 6 individuals were affected (5 males and a female),
26 individuals, including 6 affected were involved in DNA-study. Genotyping
analysis with polymorphic microsatellite markers NDPCA,
DXS1055 and DXS1003 from Xp11.3 showed the evidence of linkage
the disease with RP2 locus on chromosome X. Sequencing analysis
of all exons and intron-exon junctions of RP2 in affected man revealed
a novel mutation- small deletion c.10-12delTTC in exon 1 of RP2. The
mutation c.10-12delTTC was identified in all affected men in hemizygous
and in all women-carriers in heterozygous state in the family.
The results of the DNA-study revealed a novel disease-causing mutation
c.10-12delTTC in RP2 gene, leading to X-linked retinitis pigmentosa
type 2.
P12.143
Expression and siRNA interference of Rhodopsin cis-acting
splicing mutants associated with autosomal dominant Retinitis
Pigmentosa
I. Hernan, M. Gamundi, E. Borràs, M. Carballo;
Hospital de Terrassa, Terrassa, Spain.
Retinitis Pigmentosa (RP), a clinically and genetically heterogeneous
group of retinal degeneration disorders affecting the photoreceptor
cells, is one of the leading causes of genetic blindness. Mutation in the
rhodopsin gene (RHO) is the most prevalent cause of adRP (autosomal
dominant RP).
Two cis-acting mutations (c.531-2A>G and c.937-1G>T), that lead to
a deficient pre-mRNA splicing, affect the splice sites of RHO and are
linked to adRP while a similar cis-acting mutation (c.936+1G>T) has
been linked to autosomal recessive RP (arRP). Transcriptional expression
analysis shows that cis-acting splicing mutations causing adRP
use intronic and/or exonic alternative splice sites while arRP mutation
results in a total exclusion of exon 4. Although protein expression analysis
confirms the translation of three RHO mutants, if some of these
mutants (carrying a premature termination codon) are targeted by a
NMD mechanism is being studied.
Since most mutations causing adRP have a dominant-negative effect,
three siRNA molecules have been designed to interfere the mutant
transcripts detected in adRP families. Two of them specifically eliminate
the desired product.
P12.144
Identification of mutations in the intracellular Ca2+ release
channels caused cardiac and skeletal muscle disorders
I. Valášková1,2 , E. Flodrová1,2 , E. Švandová2 , Š. Prášilová1,2 , R. Gaillyová1,2 , P.
Kuglík1,2 , T. Novotný1 ;
1 2 University Hospital, Brno, Czech Republic, Masaryk University, Brno, Czech
Republic.
Rapid mobilization of calcium from the sarcoplasmic reticulum (SR)
into cytosol triggers activation of contractile elements, and it is therefore
a fundamental process in the physiology of heart and muscles.
The channels that regulate the duration and amplitude of calcium efflux
from the SR are the ryanodine receptors (RyRs).Three subtypes
of these proteins exist: RyR1 is mainly expressed in skeletal muscle,
RyR2 is highly represented in cardiac tissue, and RyR3 is preferentially
expressed in the brain. Mutations of RyR1 muscle isoform have been
associated with predisposition to 2 diseases, malignant hyperthermia
and central core disease. Mutations in the RyR2 gene have been identified
in families and in sporadic patients affected by catecholaminergic
polymorphic ventricular tachycardia (CPVT) and arrhythmogenic right
ventricular dysplasia type 2 (ARVD2). The distribution of mutations
is identical for RyR1 and RyR2. Most RyRs mutations are clustered
in the amino terminus, in the FKBP12.6-binding domains, and in the
transmembrane domains of the proteins. We performed RYRs mutation
screening based on clustering of known mutations along the RyR1
and RyR2 genes. A multi-step approach is proposed: melting curve
and HRM analysis of critical exons, DNA and cDNA sequencing of MH
critical regions. We are able to identify pathogenic mutations in the
clinically affected proband. DNA analysis can be extended to all family
members to identify those are asymptomatic but genetically affected.
Detection of RyRs mutations is important because a preventing of fatal
cardiac and skeletal muscle disorders in genetically affected patients
has been shown to be effective.
P12.145
two heterozygous itPR1 deletions in German families with
dominant ataxia
P. Bauer 1 , C. Bauer 1 , M. Synofzik 2 , T. Schmitz-Hübsch 3 , U. Wüllner 3 , M. Bonin 1 ,
O. Riess 1 , L. Schöls 2 ;
1 Department of Medical Genetics, Tübingen, Germany, 2 Hertie-Insitute for Clinical
Brain Research, Neurodegeneration, Tübingen, Germany, 3 Department of
Neurology, Bonn, Germany.
At least 28 loci have been linked to autosomal dominant spinocerebellar
ataxia (ADCA). Causative genes have been cloned for nine nucleotide
repeat expansions (SCA1,2,3,6,7,8,10,12&17) and eight genes
with missense mutations (SCA4,5,11,13,14,15(16),27&28). Recently,
heterozygous genomic deletion comprising the ITPR1 gene on human
chromosome 3p24 have been identified as the molecular defect underlying
SCA15 in Australian and Japanese ataxia families. In order to assess
the prevalence and clinical phenotypes of SCA15, we screened
69 patients with autosomal-dominant ataxias for genomic deletions in
exons 1 and 4 of the ITPR1-gene.
Two index patients showed relative gene dosage reduction for both
exons after qPCR indicating a heterozygous genomic deletion for at
least exon 1 and exon 4 of the ITPR1-gene. To validate these findings
we performed a high density SNP genotyping array (Affymetrix 6.0).
Copy number analysis validated both heterozygous genomic deletions
deleting approximativly 200kb and 500kb, respectively. Both patients
had phenotypes compatible with rather pure cerebellar ataxia.
In our entire ADCA cohort (n=274), SCA15 is a rare cause of spinocerebellar
ataxia in Caucasians accounting for approximatively 1% of
dominant ataxias. Noteworthy this prevalence is comparable to SCA14
and higher than SCA11 and SCA27.
P12.146
scA15/16: Phenotype in 4 families with deletions of the itPR-1
gene
A. Durr 1,2 , C. Marelli 1,2 , J. Johnson 3 , J. van de Leemput 4 , E. Ollagnon-Roman 5 ,
F. Tison 6 , F. Picard 7 , S. Sangla 8 , C. Thauvin-Robinet 9 , H. Dollfus 10 , J. Hardy 11 ,
G. Stevanin 1,2 , A. Brice 1,2 , A. Singleton 4 ;
1 CRicm UMRS975/NEB, Paris, France, 2 APHP, Département de Génétique et
Cytogénétique, Groupe Hospitalier Pitié-Salpêtrière, Paris, France, 3 Neurogenetics,
Department of Molecular Neuroscience, Institute of Neurology, London,
United Kingdom, 4 Molecular Genetics Unit, National Institute in Aging, NIH,