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

Concurrent Sessions
been replicated in numerous studies in different populations. However,
evidence for the underlying mechanism how USF1 variants relate to
dyslipidemia remains limited.
We here show for the first time, allele-specific differences in USF1
transcript levels in two relevant tissues. In muscle biopsies from 142
twins, following insulin challenge only subjects homozygous for the
non-risk allele responded with significantly increased USF1 transcript
levels, as measured by RT-PCR. Essentially the same phenomenon
was observed in fat-biopsies of FCHL family members as insulin levels
correlated with allelic imbalance. Quantitative sequencing of genomic-
and transcribed RNA from subjects heterozygous for the best associating
SNP, located in the 3’-UTR of USF1 revealed an average ~20%
lower expression of the risk-allele in fat.
In a set of 47 expression arrays of fat-biopsies, differential expression
of numerous USF1 target genes was evident -among them many
genes of lipid metabolism and inflammatory-response. Interestingly,
the neighboring F11R gene recently implicated in development of atherosclerotic
plaques also showed allele-dependent expression, suggesting
the presence of multiple, potentially co-regulated CVD associated
genes in this chromosomal region.
In summary, common allelic variants, defined by non-coding SNPs of
the USF1 transcription factor gene and associated with dyslipidemia
risk, seemingly eradicate the insulin response of transcript levels. Subsequently,
they result in differential expression of numerous relevant
target genes predisposing carriers of risk alleles to dyslipidemia and
CVD.
C66. The cohesion protein NIPBL recruits histone deacetylases
to mediate chromatin remodeling
W. Xu1 , P. Jahnke1 , M. Wülling2 , M. Albrecht1 , G. Gillessen-Kaesbach1 , F. J.
Kaiser1 ;
1 2 Universitätklinikum S-H,, Lübeck, Germany, Universität Duisburg-Essen, Essen,
Germany.
Cornelia de Lange Syndrome (CdLS) is a rare malformation disorder
with multiple congenital anomalies, a characteristic face, growth and
mental retardation as well as gastrointestinal and
limb abnormalities. About 50 % of the patients with CdLS carry mutations
in the NIPBL gene.
NIPBL encodes a homologue of the fungal Scc2-type and Drosophila
Nipped-B protein and is part of the chromatid cohesion complex.
Recent studies show an association of chromatid cohesion with chromatin-remodeling
complexes, either in the recruitment of cohesion to
particular sequences along chromosome arms or in the establishment
of cohesion.
In yeast-two hybrid assays we could identify the chromatin remodeling
factors histone deacetylases 1 and 3 (HDAC1 and 3) as potential NIP-
BL-interacting proteins. Using different fragments of NIPBL in liquid βgalactosidase
assays, we could narrow down the interacting region for
HDAC1 and 3 to a stretch of 162 amino acids (aa) within a highly conserved
region of NIPBL which was predicted to be a HDAC-interacting
domain by in silico studies. In luciferase reporter gene assays we could
show that this HDAC-interacting domain of NIPBL fused to the GAL4-
DNA-binding domain (GAL4-DBD) is able to repress reporter gene
transcription. Moreover, cotransfections of HDAC1 and 3 enhance this
effect significantly. To confirm whether this effect is based on histone
deacetylation we used an antibody recognizing the acetylated histone
3 in chromatin immunoprecipitation assays and could identify specific
deacetylation of histone 3 adjacent to the NIPBL-GAL4-DBD binding
sites. Our data show that NIPBL is able to recruit chromatin remodeling
enzymes mediating histone deacetylation.
C67. The Human Epigenome Project (HEP)
S. Beck;
Wellcome Trust Sanger Institute, Cambridge, United Kingdom.
Epigenetic processes play an essential role in biology with wide-ranging
implications for human health and disease. To understand and harness
these processes we need to read and interpret the epigenetic
code with the same rigour and vigour that made reading the genetic
code one of the greatest scientific achievements. To this end, a number
of efforts have already been initiated of which the EU-funded HEP
was among the first to be set up in 2000 with the aim to map one of the
epigenetic marks, DNA methylation. On behalf of the HEP Consortium,
I will present our findings to date, discuss some of the lessons learnt
and give an outlook on how the data and technology may be used in
an integrated (epi)genetic approach to common disease.
www.epigenome.org
C68. Mechanism of Alu integration into the human genome
J. Chen 1,2 , C. Férec 2,3 , D. N. Cooper 4 ;
1 Etablissement Français du Sang–Bretagne, Brest, France, 2 INSERM, U613,
Brest, France, 3 Université de Bretagne Occidentale, Faculté de Médecine de
Brest et des Sciences de la Santé, Brest, France, 4 Institute of Medical Genetics,
Cardiff University, Cardiff, United Kingdom.
LINE-1 or L1 has driven the generation of at least 10% of the human
genome by mobilising Alu sequences. Although there is no doubt that
Alu insertion is initiated by L1-dependent target site-primed reverse
transcription, the mechanism by which the newly synthesised 3’ end
of a given Alu cDNA attaches to the target genomic DNA is less well
understood. Intrigued by observations made on 28 pathological simple
Alu insertions, we have sought to ascertain whether microhomologies
could have played a role in the integration of shorter Alu sequences
into the human genome. A meta-analysis of the 1624 Alu insertion
polymorphisms deposited in the Database of Retrotransposon Insertion
Polymorphisms in Humans (dbRIP), when considered together
with a re-evaluation of the mechanism underlying how the three previously
annotated large deletion-associated short pathological Alu inserts
were generated, enabled us to present a unifying model for Alu
insertion into the human genome. Since Alu elements are comparatively
short, L1 RT is usually able to complete nascent Alu cDNA strand
synthesis leading to the generation of full-length Alu inserts. However,
the synthesis of the nascent Alu cDNA strand may be terminated prematurely
if its 3’ end anneals to the 3’ terminal of the top strand’s 5’
overhang by means of microhomology-mediated mispairing, an event
which would often lead to the formation of significantly truncated Alu
inserts. Furthermore, the nascent Alu cDNA strand may be ‘hijacked’
to patch existing double strand breaks located in the top-strand’s upstream
regions, leading to the generation of large genomic deletions.
C69. Evolution of mammalian gene expression promoted by
retroposons
N. V. Tomilin;
Russian Academy of Sciences, St.Petersburg, Russian Federation.
Expression of eukaryotic protein-coding genes is strongly affected by
specific sequence motifs which concentrate near transcription start
sites (TSS) and bind transcription factors. These transcription factor
binding motifs (TFBM) may arise in promoters by slow accumulation
of base changes in a random sequence but recent data indicate that
promoter TFBMs can also arise rapidly through insertion of mobile elements
already having functional motifs or having sequences which
may be converted to TFBMs by a small number of mutations. For example,
single C to T transition at specific position can create functional
estrogene-response element in human Alu retroposons. We studied
abundance of retroposon-derived sequences in human and mouse
promoters (-1000 to +200 bp segments relative to TSS) and found that
>15000 of human promoters and >11000 of mouse promoters have
Alu-derived or B1/B2-derived elements, resp. Global distribution of
these retroposons in human and mouse chromosomes strongly correlates
with clusters of CpG islands present in promoters of ~75% genes.
In active genes CpG islands are not methylated but major fraction of
retroposons, especially LINEs and LTRs, are heavily methylated.
It is unknown how promoter CpG islands are protected from spread
of methylation initiated at adjacent LINEs and LTRs. Human Alu and
mouse B1 elements can be transcribed by RNA polymerase III because
their internal promoters bind transcription complex TFIIIC. This
complex can limit spread of repressive histone methylation in S.pombe
and we suggest that Alu- and B1-associated TFIIIC sites have similar
function in mammals.
C70. Prominent use of distal 5’ transcription start sites and
discovery of a large number of additional exons in ENCODE
regions
A. Reymond 1 , F. Denoeud 2 , J. Harrow 3 , C. Ucla 4 , A. Frankish 3 , C. Henrichsen 1 ,
C. Wyss 4 , T. Alioto 2 , J. Drenkow 5 , J. Lagarde 2 , T. Hubbard 3 , S. E. Antonarakis 4 ,
R. Guigo 2 , P. Kapranov 5 , T. R. Gingeras 5 ;
1 Center for Integrative Genomics, University of Lausanne, Lausanne, Switzerland,
2 IMIM, Barcelona, Spain, 3 Wellcome Trust Sanger Institute, Hinxton,
1