European Human Genetics Conference 2007 June 16 – 19, 2007 ...

Molecular and biochemical basis of disease
aberrant expression of Connexin40 and Connexin43 and the transcription
factor Nkx2.5 in vivo specifically within the sinoatrial nodal region,
and show that Shox2 deficiency interferes with pacemaking function
in Zebrafish embryos. Conclusion: From these results, we postulate a
critical function of Shox2 in the recruitment of sinus venosus myocardium
comprising the sinoatrial nodal region.
P0806. Multiple sulfatase deficiency is due to hypomorphic
mutations of the SUMF1 gene
I. Annunziata1 , V. Bouchè1 , L. Schlotawa2 , T. Dierks2 , A. Ballabio1 ;
1 2 TIGEM, Naples, Italy, Universitaet Bielefeld Universitaetsstr, Bielefeld, Germany.
Sulfatases catalyze the hydrolysis of sulfate esters bonds from a wide
variety of substrates. Several human inherited diseases are caused
by the deficiency of individual sulfatases. Multiple sulfatase deficiency
(MSD) is a rare autosomal recessive disorder characterized by the
simultaneous deficiency of all known sulfatases. MSD patients carry
mutations of the Sulfatase Modifying Factor 1 (SUMF1) gene encoding
the α-formylglycin generating enzyme (FGE) which is required for
the post-translational modification of sulfatases. Residual sulfatase activities
are detectable, at variable levels, in all MSD patients. We have
used a recently developed Sumf1 KO mouse line which completely
devoid of all sulfatase activities to investigate on the nature of the residual
sulfatase activities detected in MSD patients. Four mutations
(i.e. S155P, R224W, R345C, R349W) found in homozygosis in MSD
patients were over-expressed, using viral-mediated gene delivery, in
Sumf1-/- mouse embryonic fibroblasts (MEFs). The results obtained
indicate that mutant SUMF1 cDNAs encode stable SUMF1 proteins
which are of the appropriate molecular weight and are properly localized
in the endoplasmic reticulum. Expression of these cDNAs in
Sumf1-/- MEFs results in low, albeit significant, FGE activity, and in
partial rescue of sulfatase activities. These data indicate that MSD is
due to hypomorphic SUMF1 mutations and suggest that a complete
loss of SUMF1 function is likely to be lethal in humans.
P0807. Frequency of LHON mutations in mitochondrial disease
phenotypes from Centre Portugal
M. M. Grazina 1,2 , J. M. Pratas 2 , S. Oliveira 2 , M. Oliveira 2 , L. Diogo 3 , P. Garcia 3 ,
C. Macário 4 , C. R. Oliveira 1,2 ;
1 Faculty of Medicine, Coimbra, Portugal, 2 Centre for Neuroscience and Cell
Biology, Coimbra, Portugal, 3 Paediatric Hospital, Coimbra, Portugal, 4 University
Hospitals, Coimbra, Portugal.
INTRODUCTION: Leber’s hereditary optic neuropathy (LHON) is a
maternally mitochondrial disorder, characterized by bilateral visual
loss, most frequently found in young males. Classical LHON is mainly
associated to mitochondrial DNA (mtDNA) mutations G11778A,
G3460A and T14484C, localized in the coding regions for ND4, ND1
and ND6 (complex I subunits of mitochondrial respiratory chain), respectively.
Other mutations (secondary) have also been described in
LHON cases.
METHODS: We have studied a total of 648 subjects, counting with
46 healthy family members. The techniques employed included PCR,
RFLP and sequencing analysis. We investigated 7 point mutations,
described as primary (G3460A, G11778A, T14484C) or secondary
(T4216C, A4917G, G13708A, G15257A), associated previously to
LHON.
RESULTS: We have found 40 positive cases (6.2%), corresponding to
the analysis of 52 tissues. Considering the total number of cases studied,
the frequencies found were: 0.002; 0.012; 0.003; 0.034; 0.026;
0.014 and 0.012, for mutations G3460A, G11778A, T14484C; T4216C,
A4917G, G13708A and G15257A, respectively.
DISCUSSION: The most frequent mtDNA alterations found, among
primary and secondary mutations studied, were G11778A and
T4216C, respectively. The primary mutations were found only in LHON
or LHON-plus cases. Secondary mutations were observed in a wide
variety of phenotypes, as expected. On the other hand, the primary
mutations were found isolated, whereas secondary mutations were
frequently encountered in combinations.
21
P0808. Mutations in mtDNA in Children with manifestations of
mitochondrial encephalomyopaties
J. Pilch 1 , M. Asman 2 , E. Jamroz 1 , M. Kajor 3 , E. Kotrys-Puchalska 4 , M. Goss 4 ,
M. Krzak 2 , M. Tarnowski 2 , J. Witecka 2 , J. Gmiński 4 , A. L. Sieroń 2 , E. Marszał 1 ;
1 Department of Child Neurology, Medical University of Silesia, Katowice, Poland,
2 Department of General & Molecular Biology & Genetics, Medical University
of Silesia, Katowice, Poland, 3 Department of Anatomopathology, Medical
University of Silesia, Katowice, Poland, 4 Department of Biochemistry, Medical
University of Silesia, Katowice, Poland.
Mitochondrial encephalomyopaties are composed disorders with wide
range of clinical manifestations. Lack of specific correlation between
genotype and phenotype makes the genetic diagnostic even more difficult.
Particularly tedious is identification of mutations in mitochondrial
DNA. In years 2004 to 2006 in Department of Child Neurology muscle
biopsies were performed in a group of sick children with manifestations
of mitochondrial encephalomyopaties. In the biopsies the biochemical,
histopathological, and ultrastructural analyses have been conducted.
The cellular respiratory chain enzymes activities were assayed in muscles
and skin fibroblasts. Based on clinical manifestations and results
of enzymatic assays a group of 21 sick children was selected for further
molecular studies.
Aim of the study: The goal was to identify mutations in mitochondrial
DNA isolated from muscle biopsies of affected children.
Material and methods: Molecular analysis performed in two steps for
entire mitochondrial DNA. First step was to screen the mitochondrial
DNA for any mismatches by heteroduplex analysis using endonuclease
SURVEYOR kit (Trangenomic, Inc., Omaha, NE, USA). DNA fragments
showing mismatches were subjected to DNA sequencing using
ABI PRISM 310 DNA analyzer.
Results: The most common changes found in all analyzed fragments
of mitochondrial DNA were polymorphisms. Mutations were detected
in few genes e.g.: a) 16S rRNA (in 8 children); b) subunits ND2, ND4L,
ND5, and ND6 of complex I (in 6); c) tRNA for leucine (in 1); and d)
tRNA for proline (in 1). Further analysis is necessary for comparing
genotypes with clinical phenotypes to final confirmation of pathogenic
character of detected mutations.
P0809. Mutational analysis of the mitochondrial 12SrRNA and
tRNASer(UCN) genes in patients with aminoglycoside-induced
and nonsyndromic hearing loss from Volga-Ural region and
Siberia (Russia)
L. U. Dzhemileva 1 , O. L. Posukh 2 , N. A. Barashkov 3 , A. M. Tazetdinov 1 , S. A.
Fedorova 3 , N. R. Maximova 3 , E. E. Fedotova 3 , V. N. Tadinova 4 , E. K. Khusnutdinova
1 ;
1 Institute of biochemistry and genetics, Ufa, Russian Federation, 2 Institute of
Cytology and Genetics, Siberian Branch of Russian Academy of Sciences,
Novosibirsk, Russian Federation, 3 Yakutsk Research Centre, Siberian Branch
of Russian Academy of Medical Sciences, Yakutsk, Russian Federation, 4 Republican
Altai Children’s Hospital, Gorno-Altaisk, Russian Federation.
Several mtDNA mutations have been found to be associated with
nonsyndromic sensorineural hearing loss (NSHL). The m.1555G>A
(12SrRNA) was confirmed as the main cause of aminoglycoside induced
deafness in different populations. The m.7445A>G, m.7472_
7473insC, m.7510T>C, m.7511T>C (tRNASer(UCN)) were reported to
be associated with NSHL. Pathogenicity of some other variations in
12SrRNA and tRNASer(UCN) genes associated with NSHL has not
yet been confirmed (http://www.mitomap.org). We report here the results
of mutational screening for 12S rRNA and tRNASer(UCN) genes
among Cx26- and Cx30-negative deaf individuals of different ethnicity
from some regions of Russia. Previously, 301 unrelated patients
(70 with aminoglycoside-induced deafness, and 231 with NSHL) from
Volga-Ural region, 78 unrelated patients with NSHL from the Republic
Sakha (Yakutia, northeastern Siberia), and 119 deaf patients (75 unrelated
families) from the Republic Altai (south Siberia) were analyzed
for Cx26 and Cx30 mutations. Different variations at the 961 position in
12S rRNA gene have been found among deaf individuals from Volga-
Ural region. Four patients of Tatar ethnicity, one with m.961delTinsCn,
and three with m.961delTinsC, two Russian patients, one with
m.961T>G, and another with m.961T>A, were detected. Moreover, the
m.7444G>A (tRNASer(UCN)) was found in one Russian patient with
NSHL. The m.1555G>A (12S rRNA) was detected in one Yakut patient
(Republic Sakha). Finally, m.7445G>C (tRNASer(UCN)) was found in
two sibs of one Kazakh family (Altai region) in whom moderate sen-