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

4. Therapy98% Identical 99,8% 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

n*

B

n

B

*

Linkage:

If a gene and a marker are on the same chromosome they will segregate together

UNLESS

They are separated by recombination

A

A

n*

B A

n*

B

A

* n

B B

n*

A

Robinow syndrome

Short stature

Wide-spaced eyes

Short nose

Small penis

Human Genetics Nijmegen

max = 6.47 = 0D9S1842

Chromosoom 9q21-q22.3D9S1842

D9S1781

D9S197

D9S1816

D9S280

D9S1851

D9S287

D9S176

2.8

ROR2

1.6

0

1.4

0.1

0.6

2.1

cM

Linkage interval Robinow syndroom

Human Genetics Nijmegen

Robinow syndrome Ror2 null mouse

From DeChiara et al. Nature Genetics March 2000

COMPLEX DISORDERS

MAP DISEASES TO CHROMOSOMES

Genotyping Single Nucleotide Polymorphisms (SNPs)

…cctcctagggttgcaaagcctccttggctatg……cctcctagggttgcatagcctccttggctatg…

Person B:Person A: …cctcctagggttgcatagcctccttggctatg……cctcctagggttgcatagcctccttggctatg…

Allel 1

Allel 2

~ 1,000,000 SNPs

> 1 SNP per >3 kb

500,000 SNPs

arrays

Whole genome association studies

Diabetes type 1

Obesity

ADHD

2000 cases 4000 controls

SNPs indicate genes involved

Gene 1Gene 2Gene 3Gene 4

……Gene 30.000

Case control design

500,000 SNPs

arrays

Whole genome association study

Obesitas

9000 cases 30000 controls

BMI > 30

FTO gene

Case control design

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.

35% +0 kg50% +1.5 kg

15% +3.0 kg

FTO gene

500.000 SNPs

arrays

Whole genome association study

Obesitas9000 cases 30000 controls

BMI > 30

FTO gene

INTERPRET GENETIC VARIATION

• Sequence variation at a specific nucleotide• Copy number variations (CNV)

Ins A

p63 gene mutations in EEC syndrome

V202M S272N

R279C (3)R279H (12)R279Q

C306RR304W (8)R304Q (14)R304P

R204W (10)R204Q (7)R204L R280C (6)

R280H (2)R280S

R227Q (8)

C308SC308Y

P309S

D312HD312N

C269Y

TA SAMDNA binding IsoTA-p63

Y192C (3)

L162PY163C

29 Mutations in 90 families28 missense1 frameshift

A315ER313G

L248C

Structure model of p63 DNA binding domain

276 copy number abnormalities in 100 patients with Mental Retardation

3

20

23 23

21

5

1

3

01

0

5

10

15

20

25

Perc

enta

ge o

f pat

ient

s

0 1 2 3 4 5 6 7 8 90

5

10

15

20

25

0 1 2 3 4 5 6 7 8 90

5

10

15

20

25

DNA copy number alterations identified per patient

0 1 2 3 4 5 6 7 8 9

How do we differentiate normal variation from causal changes?

Patient 1

Genomic profile obtained 250K SNP array

Log

2 Pa

tient

/ Con

t rol

Chromosome 15

Chromosome 15

Mother

Father Chromosome 1

Chromosome 1

Chromosome 1

Paient 1 Chromosome 15

de novo inherited variation

Alex Hoischen Christian Gillisen

Next Generation sequencing

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 D. Watson, sequenced to 7.4-fold redundancy in two months using massively parallel sequencing

1953

Question of the year 2007

Nature genetics

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?

1.) Sequence Capture

2.) Sequencing

3.) Mapping

mapped reads

formed contigstargeted exon(s)

4.) Mutation detection

Can we look at the all EXons of the genOME?EXOME sequencing!

ABI SOLID 600 million map-able 50bp reads 30Gb

Roche 454 1 million map-able 500bp reads 500Mb

To understand human health and disease we have to understand all

types of genomic variation:

~4,000,000 variants

~3,000,000 SNP variants*

~10,000 non-synonymous coding variants*

~1,000,000 CNVs*

* Per individual genome

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

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, MartaZ 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 VeltmanNature Genetics

Alex Hoischen Christian GillisenBregje van Bon

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

RPGR

Photoreceptor cilium protein complex

Retinitis Pigmentosa

RPGRIP1

NPHP2inversin

NPHP5IQCB1

nephrocystin-3

NPHP3

RPGRIP1L

NPHP4nephrocystin-4

PDE-δ

Arl3 RP2 β-tubulin

NPHP1nephrocystin-1

RPGR

Photoreceptor cilium protein complex

CEP290

Dynein

lebercilin *

CC2D2A

RPGRIP1

NPHP2inversin

NPHP5IQCB1

nephrocystin-3

NPHP3

RPGRIP1L

NPHP4nephrocystin-4

PDE-δ

Arl3 RP2 β-tubulin

NPHP1nephrocystin-1

RPGR

Photoreceptor cilium protein complex

Senior Loken

RP

LCA / Joubert / Meckel

Joubert / Meckel

CEP290

Dynein

lebercilin *LCA

LCA

Nephron- ophthisis

CC2D2A

Joubert

Joubert

GENETICA VAN GEHOORVERLIES

ROL VAN BIOINFORMATICA

AANGEBOREN GEHOORVERLIES

~ 1 in 900 children has congenital hearing impairment >20 dB in one or more frequencies

50 % inherited 50% environmental

70% Nonsyndromic 30% Syndromic

~%77AR

~%22 AD

~%1 X-linked

<%1 Mitochondrial

• Usher• Alport• Pendred• Norrie• Waardenburg• Branchio-Oto-Renal• Jervell and Lange-

Nielsen

• Ototoxic drugs

• Acustic trauma

• Infections~%77AR

~%22 AD

Known Genes

21 6 166

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

TECTA

MYO7A

USH1C

DFNA32

DFNB20

DFNB24

DFNB51

DFNB63

D11S2371

D11S1337

D11S4179

D11S1291

D11S916

D11S1314

D11S4139

D11S4136

D11S4113

D11S987

~5.2

9 M

b

15.515.415.315.215.114.314.214.1131211.211.1211.111112.112.212.313.113.213.313.413.514.114.214.32122.122.222.323.123.2

23.3

24.124.224.325

FGF3DFN

B63

TR57

26 bekende of voorspelde genen

FT1A-G PKDF702

DFN

B63

DFN

B63

DFN

B63

FT2

1.03 Mb

LRTOMT KARAKTERISATIE

Genome browser build 36.1

LRTOMT1

LRTOMT2

EFFECT VAN MUTATIES

E110KA29SfsX54 (c.358+4G>A)

W105RR81Q

A215AG163VfsX4 (c.358+4G>A)3’ UTR3’ UTR

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.