Hanny Kremer about the project

HANNIE KREMER
KNO & ANTROPOGENETICA
ANTROPOGENETICA – HUMAN GENETICS
Things we do
1. Map diseases to chromosomes
(position) - monogenic and complex
disorders
2. Interpret DNA variation – monogenic
and complex disorders
3. Understand the function of genes pathogenesis
99,8% identical
4. Therapy
98% Identical
Things we do
1. Map diseases to chromosomes
(position) - monogenic disorders
2. Interpret DNA variation – monogenic
and complex disorders
3. Understand the function of genes pathogenesis
4. Therapy
MAP DISEASES TO CHROMOSOMES
MONOGENIC DISORDERS
A
*
A
*
B
Linkage:
If a gene and a marker are
on the same chromosome
they will segregate
together
n
B
n
UNLESS
They are separated by
recombination
A
B
A
B
*
n
*
n
A
B
B
A
*
n
*
n
Robinow syndrome
Short stature
Wide-spaced eyes
Short nose
Small penis
Linkage interval Robinow syndroom
cM
2.8
D9S1842
Chromosoom 9q21-q22.3
D9S1781
ROR2
2.1
D9S197
0.6
0.1
D9S1816
D9S280
max = 6.47
=0
D9S1842
1.4
D9S1851
0
D9S287
1.6
D9S176
Human Genetics Nijmegen
Robinow syndrome Ror2 null mouse
From DeChiara et al. Nature Genetics March 2000
Human Genetics Nijmegen
MAP DISEASES TO CHROMOSOMES
COMPLEX DISORDERS
Genotyping Single Nucleotide Polymorphisms (SNPs)
> 1 SNP per >3 kb
Person A:
Person B:
Allel 1 …cctcctagggttgcaaagcctccttggctatg…
…cctcctagggttgcatagcctccttggctatg…
Allel 2 …cctcctagggttgcatagcctccttggctatg…
…cctcctagggttgcatagcctccttggctatg…
~ 1,000,000 SNPs
500,000 SNPs
Whole genome association studies
Diabetes type 1
Obesity
ADHD
arrays
Case control design
2000 cases
4000 controls
SNPs indicate genes involved
Gene 1
Gene 2
Gene 3
Gene 4
……
Gene 30.000
500,000 SNPs
Whole genome association study
Obesitas
arrays
Case control design
9000 cases
30000 controls
BMI > 30
FTO gene
Frayling et al. A common variant in the FTO gene is associated with
body mass index and predisposes to childhood and adult obesity.
Science 316: 889-894, 2007.
500.000 SNPs
Whole genome association study
Obesitas
arrays
9000 cases
30000 controls
BMI > 30
FTO gene
FTO gene
35%
+0 kg
50%
15%
+1.5 kg
+3.0 kg
INTERPRET GENETIC VARIATION
• Sequence variation at a specific nucleotide
• Copy number variations (CNV)
p63 gene mutations in EEC syndrome
TA-p63
TA
DNA binding
L162P
Y163C
Y192C (3)
V202M
C269Y
S272N
L248C
R279C (3)
R279H (12)
R204W (10)
R279Q
R204Q (7)
R204L
R227Q (8)
29 Mutations in 90 families
28 missense
1 frameshift
Iso
A315E
R313G
D312H
D312N
P309S
C308S
C308Y
C306R
R304W (8)
R304Q (14)
R304P
R280C (6)
R280H (2)
R280S
SAM
Ins A
Structure model of p63 DNA binding domain
276 copy number abnormalities in 100 patients with
Mental Retardation
25
2525
23
23 23
21
21 21
20
20 20
20
2020
Percentage of
of patients
patients
Percentage
Percentage of patients
23
23 23
How do we differentiate
normal variation from causal
changes?
15
1515
10
1010
5 55
5 5
3 3
3 33
1 1
00 0
1 11
0 00
0
0 0
11 1
22 2
333
4 44
5 55
6 6
7 77
DNA
copy
number
per
patient
DNA
copy
numberalterations
alterationsidentified
per
patient
DNA
copy
number
alterations
identified
per
patient
8 88
9 99
Genomic profile obtained 250K SNP array
Patient 1
de novo
inherited variation
Paient 1
Chromosome 1
Mother
Chromosome 1
Chromosome 15
Chromosome 1
Chromosome 15
Father
Chromosome 15
Alex Hoischen
Christian Gillisen
Next Generation sequencing
1953
The complete genome of an individual by massively parallel DNA sequencing.
Wheeler et al. Nature, April 2008
Here we report the DNA sequence of a diploid genome of a single individual, James
Watson, sequenced to 7.4-fold redundancy in two months using massively parallel
sequencing
D.
Nature genetics
Question of the year 2007
The sequencing of the equivalent of an entire
human genome for $1,000 has been announced
as a goal for the genetics community
What would you do if this sequencing
capacity were available immediately?
Can we look at the all EXons of the genOME?
EXOME sequencing!
1.) Sequence Capture
2.) Sequencing
mapped reads
formed contigs
targeted exon(s)
3.) Mapping
4.) Mutation detection
ABI SOLID
600 million map-able
50bp reads  30Gb
Roche 454
1 million map-able
500bp reads  500Mb
* Per individual genome
~3,000,000 SNP variants*
~1,000,000 CNVs*
To understand human
health and disease we
have to understand all
types of genomic
variation:
~4,000,000 variants
~10,000 non-synonymous
coding variants*
Focus on de novo disease
• 4 DNAs from patients with Schinzel-Giedion
syndrome
• patient samples n=14
• 4 human exomes: 2.5Gb output per sample
Alex Hoischen
Bregje van Bon
Christian Gillisen
De novo mutations of SETBP1 cause Schinzel-Giedion syndrome in 13
patients
Alexander Hoischen*, Bregje WM van Bon*, Christian Gilissen*, Peer Arts, Bart van Lier,
Marloes Steehouwer, Petra de Vries, Rick de Reuver, Geert Mortier, Koen Devriendt, Marta
Z Amorim, Nicole Revencu, Alexa Kidd, Mafalda Barbosa, Anne Turner, Janine Smith,
Christina Oley, Alex Henderson, Ian M Hayes, Elizabeth M Thompson, Han G Brunner,
Bert BA de Vries, Joris A Veltman
Nature Genetics
Things we do
1. Map diseases to chromosomes
(position) - monogenic disorders
2. Interpret DNA variation – monogenic
and complex disorders
3. Understand the function of genes pathogenesis
4. Therapy
Photoreceptor cilium protein complex
Retinitis Pigmentosa
RPGR
Photoreceptor cilium protein complex
Arl3
RP2
β-tubulin
inversin
NPHP2
lebercilin
*
PDE-δ
nephrocystin-3
IQCB1
NPHP5
CC2D2A
NPHP3
nephrocystin-1
NPHP1
nephrocystin-4
Dynein
RPGR
NPHP4
RPGRIP1L
CEP290
RPGRIP1
Photoreceptor cilium protein complex
Arl3
LCA
lebercilin
Joubert
CC2D2A
β-tubulin
inversin
Nephron
*
PDE-δ
- ophthisis
IQCB1
RP
Dynein
RP2
NPHP5
Senior
Loken
NPHP2
Joubert
nephrocystin-3
NPHP3
nephrocystin-1
NPHP1
nephrocystin-4
RPGR
NPHP4
RPGRIP1L
CEP290
LCA / Joubert /
Meckel
RPGRIP1
LCA
Joubert /
Meckel
GENETICA VAN GEHOORVERLIES
ROL VAN BIOINFORMATICA
AANGEBOREN GEHOORVERLIES
~ 1 in 900 children has congenital hearing impairment >20 dB in one or more
frequencies
50% environmental
50 % inherited
70% Nonsyndromic
~%22
AD
~%77
AR
21
66
16
Known Genes
~%1
X-linked
30% Syndromic
<%1
Mitochondrial
•Ototoxic drugs
•Acustic trauma
•Infections
•Usher
•Alport
•Pendred
•Norrie
•Waardenburg
•Branchio-Oto-Renal
•Jervell and Lange-Nielsen
WAAROM IS HET OPHELDEREN VAN OORZAKEN
VAN ERFELIJKE ZIEKTEN BELANGRIJK?
 Vraag van patiënt naar de oorzaak beantwoorden:
is het erfelijk - erfelijkheidsadvies
 Vroege diagnostiek van familieleden – goede
begeleiding
 Inzicht in genen/eiwitten die essentieel zijn voor
ontwikkeling en functie van het binnenoor
 Handvaten voor therapie
FAMILIE TR57
DFNB63 LOCUS
15.5
15.4
DFNA32
TR57
D11S987
D11S1337
MYO7A
FGF3
D11S1314
D11S2371
D11S1291
DFNB24
TECTA
DFNB20
D11S4179
DFNB63
D11S4139
D11S916
23.3
24.1
24.2
24.3
25
DFNB63
DFNB63
D11S4136
DFNB63
D11S4113
11.2
11.12
11.11
11
12.1
12.2
12.3
13.1
13.2
13.3
13.4
13.5
14.1
14.2
14.3
21
22.1
22.2
22.3
23.1
23.2
26 bekende of
voorspelde genen
DFNB51
DFNB63
12
FT2 PKDF702
USH1C
14.3
14.2
14.1
13
FT1A-G
~5.29 Mb
15.3
15.2
15.1
1.03 Mb
LRTOMT KARAKTERISATIE
Genome browser build 36.1
LRTOMT1
LRTOMT2
EFFECT VAN MUTATIES
3’ UTR
A215A
G163VfsX4 (c.358+4G>A)
R81Q
W105R
A29SfsX54 (c.358+4G>A)
3’ UTR
E110K
Catechol-O-methyltransferase domein
MOLECULAR MODELING
EFFECT VAN MISSENSE MUTATIES
HET BINNENOOR
SAMENVATTING
 Bioinformatica is essentieel voor verschillende
stappen in studies naar ziektegenen
 De structuur en functie van het humane genoom
en genen zijn nog lang niet in kaart gebracht
 De oorzaak van DFNB63 is gelegen in defecten in
het LRTOMT gen. Het precieze effect van
mutaties in dit gen op de functie van het
binnenoor is nog niet duidelijk.