2008 Barcelona - European Society of Human Genetics

Molecular and biochemical basis of disease
entire RYR1 complementary DNA (cDNA) and melting point analysis
of fluorscently labelled probes after high speed PCR amplification on
real-time enabled us to confirm MH diagnosis on molecular level in 50
MHS individuals . Recently we establish High-resolution melting (HRM)
as a method that allows RYR1 mutation scanning and genotyping . We
expect that our methodical process of identification of MH causal mutations
in MHS patients and subsequent performing noninvasive predictive
genetic testing in their family members decrease number of
individuals which would have to undergo the invasive IVCT .
P05.118
mutation screening of FBN1 and tGFBR2 genes in patients with
marfan and marfan-like syndromes from Russia: 4 mutations
and 11 polymorphisms have been found
R. Valiev, R. Khusainova, E. Khusnutdinova;
Institute of biochemistry and genetics, Ufa, Russian Federation.
Marfan syndrome (MFS) is an inherited autosomal dominant connective
tissue disorder . Abnormalities appear in skeletal, ocular and
cardiovascular systems . The main cause of MFS is mutations in the
fibrillin1 gene (FBN1). Recently, the transforming growth factor beta
receptor 2 gene (TGFBR2) has been shown to be associated with a
second type of this disorder with typically mild or absent ocular involvement
(MFS type 2) as well as with classical MFS . We analyzed
30 exons of FBN1 gene and 4 exons of TGFBR2 gene in 80 patients
with MS and marfan-like syndromes from different regions of Russia .
SSCP analysis revealed different abnormal migrating patterns . We
identified two missense mutations (G1176Y in 28 exon and C2489Y in
60 exon) which affects cbEGF-like motifs of fibrillin-1 protein in two patients
with classical MFS symptoms . We also found 9 polymorphisms
both in coding and non coding regions of FBN1 gene, five of them are
not previously described . One novel mutation (c .670C>T; T223M) has
been found in TGFBR2 gene in two unrelated patients with marfan-like
syndrome who did not fulfill Ghent nosology and who did not have
mutations in FBN1 gene . Mutation T223M affects highly conserved
serine/threonine protein kinases catalytic domain that leads to change
phospho transferring status of TGFBR2 protein . In addition two novel
polymorphisms have been found in intronic regions of TGFBR2 gene .
Mutation screening of FBN1 and TGFBR2 genes continues .
P05.119
Novel mutations of FBN in czech population
E. Prusová, A. Bóday, P. Riedlová, J. Fiser, S. Tavandzis, M. Kučerová, M.
Radina;
JG Mendel Cancer Centre, Nový Jičín, Czech Republic.
Marfan syndrome (MFS) is a heritable autosomal dominant disorder of
connective tissue with prevalence of between 1 in 5-10 000 . Approximately
25% of MFS patients are sporadic cases due to new mutations .
MFS is noteworthy for its clinical variability . Major features of the MFS
include cardiovascular disorders (dilatation and dissection of ascending
aorta), eye disorders - ectopia lentis, defects of skeletal system
- pectus carinatum, pectus axcavatum and/or other diagnostics criteria
as arachnodactyly .
MFS is caused by mutations in fibrillin 1 gene (FBN1) resulting in defective
glycoprotein fibrillin-1. FBN1 is located on chromosome 15 at
locus q15-q21 .1 . Recently, there are showed that two other genes
FBN2 (5q23-q31) and TGFBR2 (3p22) influence MFS.
Since 2006 we have done a molecular analysis for MFS diagnosis in
Czech Republic . The molecular analysis includes mlpa (multiplex ligation-dependent
probe amplification), separation of PCR products by
SSCP (single-strand conformation polymorphism) and sequencing .
We have performed mutation detection on 150 patients with suspected
MFS . There were detected 20 novel mutations .
P05.120
Large deletions account for a significant fraction of mutations in
marfan syndrome
B. M. Rhode, L. Föhse, M. Stuhrmann, J. Schmidtke, M. Arslan-Kirchner;
Institute of Human Genetics, Hannover Medical School, Hannover, Germany.
Marfan syndrome (MFS) is an autosomal dominant hereditary disorder
of connective tissue caused by mutations in the FBN1 gene . Cardinal
manifestations include proximal aortic aneurysm, ectopia lentis, and
involvement of the skeletal system . About 60 to 90% of the cases with
clinically diagnosed Marfan syndrome can be explained by a FBN1
mutation . This rate can be raised by subsequent sequencing analysis
of the genes TGFBR1 and TGFBR2 .
We tested 45 patients without identified mutation in the three genes
by use of MLPA (multiplex ligation-dependent probe amplification) and
investigated whether large deletions in the FBN1 gene would increase
the mutation detection rate sufficiently for incorporation of MLPA in
the routine diagnostics of Marfan syndrome. The patients fulfilled or
partially fulfilled the diagnostic criteria (Ghent nosology).
We identified three large deletions in the 3’ region of the FBN1 gene;
ranging from exon 50 to 54, exon 55 to 58 and exon 58 to 63 (the last
one was previously described by Singh et al ., J . Mol . Cell . Cardiol .
2007) . A fourth deletion comprised the complete FBN1 gene . Breakpoints
of the deletions were determined by long-range PCR techniques
. Implications for the resulting protein product, as well as the
phenotypes of the patients, will be discussed .
We estimated a deletion detection rate of 9% in this pre-screened patient
group, corresponding to an overall rate of 3% among all patients
with MFS . These data imply an inclusion of MLPA analysis in the routine
diagnostics of Marfan syndrome .
P05.121
Alternative splicing variants of mcPH1 with distinct functions
I. Gavvovidis 1 , M. Trimborn 2 , L. Weissflog 1 , H. Hanenberg 3 , H. Neitzel 4 , D.
Schindler 1 ;
1 Department of Human Genetics, University of Wurzburg, Wurzburg, Germany,
2 Institute of Medical Genetics, Charité University Medicine, Berlin, Germany,
3 Department of Pediatric Oncology Hematology and Immunology, Childrens
Hospital, Heinrich Heine University, Dusseldorf, Germany, 4 Institute of Human
Genetics, Charité University Medicine, Berlin, Germany.
Primary microcephaly (MCPH) is an autosomal recessive disorder
characterized by pronounced reduction of brain size and variable mental
retardation without additional neurological deficits. Four underlying
genes have been identified to date. Hallmark of patients with mutations
in the MCPH1 gene (MIM# 606858; MIM# 607117) is a cellular phenotype
of premature chromosome condensation, PCC, in the G2 phase
and delayed decondensation in G1 . MCPH1 is located on human chromosome
8p23 .1, consists of 14 exons, and encodes the protein microcephalin
containing one N-terminal and two C-terminal BRCT-domains
as well as an NLS sequence .
RT-PCR showed alternatively spliced variants of MCPH1-mRNA . In
addition to the full length MCPH1 (MCPH1-L), we detected a variant
skipping exon 8 (MCPH1-S) and another without exons 9-14 (MCPH1-
B) . The resulting polypeptides are lacking the NLS or the two C-terminal
BRCT-domains, respectively .
Expression of any of the three variants as GFP-fusion proteins by
retroviral transfer resulted in complementation of the PCC phenotype
in MCPH1-deficient cells. In contrast, a construct containing MCPH1
without the N-terminal BRCT-domain (MCPH1Δ1-7) did not complement
the PCC phenotype, suggesting a role of this domain in regulating
chromosome condensation, presumably through interaction with
condensin II .
The fluorescent signals confirmed nuclear localization of all three
variants . For MCPH1-S a GFP-signal was detected in the cytoplasm
as well, which appeared to co-localize with the centrosomes, whilst
MCPH1-L and MCPH1-B re-localized to chromatin during anaphase .
In addition, variants containing C-terminal BRCT domains re-localized
to gamma-H2AX foci indicating participation of these variants in DNAdamage
responses .
P05.122
investigation of six mental retardation loci (mcPH1, mcPH2,
mcPH3, mcPH4, mcPH5, and mcPH6) associated with
microcephaly in northeast & southeast of Iran
H. Darvish 1 , G. Bahrami Monajemi 1 , F. Rakhshani 2 , A. Zadeh-Vakili 2 , A. Naghavi
2 , M. Mohseni 1 , S. Ghasemi Firouzabadi 1 , G. Mollashahi Sanatgar 2 , J. Oveisi 2 ,
S. Banihashemi 1 , F. Behjati 1 , K. Kahrizi 1 , H. Najmabadi 1 ;
1 Genetics Research Center, University of Social Welfare and Rehabilitation
Sciences, Tehran, Islamic Republic of Iran, 2 University of medical science,
Zahedan, Islamic Republic of Iran.
Primary autosomal recessive microcephaly is defined as a reduction in
head circumference . Six out of ten Non-Syndromic Autosomal Recessive
Mental Retardation (NS-ARMR) loci associated with microcephaly
(MCPH1-MCPH6) and belong to the family of MCPH (autosomal re-