The spectrum of human diseases
Cystic fibrosis
thalassemia
<5%
cancer
Huntington’s
Mapping complex loci
PAF – population attributable factor:
Fraction of the disease that would be eliminated if the risk
factor were removed
High PAF for single gene conditions (>50% for CF)
Low PAF for complex disease (< 5% for Alzheimer’s)
Identifying genes involved in complex
diseases
Steps
Perform family, twin or adoption studies
- check for genetic component
Segregation analysis
- estimate type and frequency of susceptibility alleles
Linkage analysis
- map susceptibility loci
Population association
- identify candidate region
Identify DNA sequence variants conferring
susceptibility
Linkage versus Assocciation
Association studies compare the
allele frequency of a
polymorphic marker, or a set of
markers, in unrelated patients
(cases) and healthy controls to
identify markers that differ
significantly between the two
groups.
Used to identify common modestrisk disease variants
Higher density of markers needed
e.g. HapMap uses association
data
Linkage analyses search for
regions of the genome with a
higher-than-expected number of
shared alleles among affected
individuals within a family.
Used to identify rare high-risk
disease alleles
<500 markers needed for initial
genome scan
Haplotype analysis
• specific combination of 2 or more DNA marker
alleles situated close together on the same
chromosome (cis markers)
• SNPs most commonly used markers in haplotypes.
• series of closely linked mutations accumulate
over time in the surviving generation derived
from a common ancestor.
• powerful genetic tool for identifying ancient
genetic relationships.
• Alleles at separate loci that are associated with
each other at a frequency that is significantly
higher than that expected by chance, are said to
be in linkage disequilibrium
Direct versus indirect association analysis.
a, In direct association analysis,all functional variants (red arrows) are catalogued and tested for association with disease. A
GeneSNPs image of the CSF2 gene is shown. Genomic features are shown as boxes along the horizontal axis (for example,
blue boxes indicate exons). Polymorphisms are shown as vertical bars below the axis, with the length of the line indicating allele
frequency and colour indicating context (for example, red indicates coding SNPs that change amino acids).
b, For indirect association analysis, all common SNPs are tested for function by assaying a subset of tagSNPs in each gene
(yellow arrows), such that all unassayed SNPs (green arrows) are correlated with one or more tagSNPs. Effects at unassayed
SNPs (green arrows) would be detected through linkage disequilibrium with tagSNPs. Images adapted from GeneSNPs
(http://www.genome.utah.edu/genesnps).
Formation of haplotypes over time
Ancient disease loci are
associated with haplotypes
• Start with population genetically isolated for a long
time such as Icelanders or Amish
• Collect DNA samples from subgroup with disease
• Also collect from equal number of people without
disease
• Genotype each individual in subgroups for
haplotypes throughout entire genome
• Look for association between haplotype and disease
phenotype
• Association represents linkage disequilibrium
• If successful, provides high resolution to narrow
parts of chromosomes
Haplotype analysis provides high
resolution gene mapping
Why is it still so difficult?
Genetic heterogeneity
Mutations at more than one locus cause same
phenotype
e.g. thalassemias
– Caused by mutations in
either the a or b-globin
genes.
– Linkage analysis studies
therefore always
combine data from
multiple families
Variable expressivity - Expression of a mutant
trait differs from person to person
• Phenocopy
– Disease phenotype is not caused by any
inherited predisposing mutation
– e.g. BRCA1 mutations
• 33% of women who do not carry BRCA1 mutation
develop breast cancer by age 55
Incomplete penetrance
– when a mutant genotype does not
always cause a mutant phenotype
• No environmental factor associated with
likelihood of breast cancer
• Positional cloning identified BRCA1 as one
gene causing breast cancer.
– Only 66% of women who carry BRCA1 mutation
develop breast cancer by age 55
• Incomplete penetrance hampers linkage
mapping and positional cloning
– Solution – exclude all nondisease individuals form
analysis
– Requires many more families for study
• Polygenic inheritance
– Two or more genes interact in the expression of
phenotype
• QTLs, or quantitative trait loci
– Unlimited number of transmission patterns for QTLs
» Discrete traits – penetrance may increase with number
of mutant loci
» Expressivity may vary with number of loci
– Many other factors complicate analysis
» Some mutant genes may have large effect
» Mutations at some loci may be recessive while others
are dominant or codominant
Polygenic inheritance
E.g heart attacks or cholesterol levels
Sudden cardiac death (SCD)
Breast cancer
Common condition – familial or sporadic forms
Although a genetic basis for familial BC identified, the
causes of sporadic disease still unknown
Mutations in 2 loci account for 20-25% of early onset
(<45 years) breast cancer cases due to inherited
factors
– BRCA1: mutations found in 80-90% of families
with both breast and ovarian cancer
– BRCA2: mutations mainly in male breast cancer
familiesSudden cardiac death (SCD)
Alzheimer’s disease
Affects 5% of people >65 years and 20% of people over 80
has familial (early-onset) or sporadic (late-onset) forms, although
pathologically both are similar
Aetiology of sporadic forms unknown
familial AD – mutations in APP, presenilin-1 and 2
Sporadic AD – strong association with APOe4, Apolipoprotein e4,
which affects age of onset rather than susceptibility
3 major alleles (APO E2, E3, and E4)
Position
112
158
ApoE2
Cys
Cys
ApoE3
Arg
Cys
Sudden
cardiac
death
(SCD)
ApoE4
Arg
Arg
Genetic conditions that are
independent of the DNA sequence
Epigenetics – differential
imprinting
Epigenetics – differential imprinting
Prader-Willi syndrome
failure to thrive
during infancy,
hyperphagia and
obesity during early
childhood, mental
retardation, and
behavioural
problems
molecular defect involves a ~2
Mb imprinted domain at
15q11–q13 that contains
both paternally and
maternally expressed
genes
Angelman syndrome
characteristics
include mental
retardation,
speech
impairment and
behavioural
abnormalities
defect lies within the
imprinted domain at
15q11–q13
Genetic causes
Prader-Willi syndrome
Angelman syndrome
70% have a deletion of the
PWS/AS region on their
paternal chromosome 15
70% have a deletion of the PWS/AS
region on their maternal
chromosome 15
25% have maternal
uniparental disomy for
chromosome 15 (the
individual inherited both
chromosomes from the
mother, and none from the
father)
7% have paternal uniparental
disomy for chromosome 15 (the
individual inherited both
chromosomes from the father, and
none from the mother)
5% have an imprinting
defect
<1% have a chromosome
abnormality including the
PWS/AS region
3% have an imprinting defect
11% have a mutation in UBE3A
1% have a chromosome
rearrangement
11% have a unknown genetic cause
Molecular pathology
Nomenclature
Effect of mutant allele and not the sequence
Loss of function
Gain of function
Gene to disease
Disease to gene
Chromosomal disorders
The Haemoglobinopathies
Thalassemias
-Anaemias associated with impaired synthesis of Hb subunits
Thalassaemias can arise from different mutations causing a
disease of varying severity.
a0/b0 thalassaemias – globin chain absent
a+/b+ thalassaemias – normal globin chain in reduced amounts
Evolution of globin superfamily
Fig. 21.16
Organisation of globin genes
Fig. 21.16
Developmental variation in gene
expression
a-like chains - z & a
b-like
chains - e, g, d, b
Fig. 21.16
Adult human made of
a2b2 – 97%;
a2d2 - ~2%;
a2g2-~1% (fetal persistence)
Gene expression controlled by location
Fig. 21.16
e – embryonic yolk sac
g – yolk sac & fetal liver
b & d – adult bone marrow
a- thalassemias
a- thalassemias
deletion of one or both a globins in an a gene cluster
Severity depends on whether the individual has 1,2,3, or 4
missing a globin genes.
GENOTYPE
a+ a+ a+a+
a+a
a+a+
PHENOTYPE
Normal
Silent carrier
a+ a
a-thalassaemia trait
minor anaemic
conditions
HbH
Hydrops foetalis
mild – moderate anaemia
foetus survives until
around birth
a+a
a+a+ a a
a+a
aa
aa
aa
asymptomatic condition.
a-thalassaemia – 2
b- thalassemias
b- thalassemias
5’
Mutations in b globin cluster
are of different types
gene deletion
transcriptional mutation
RNA processing
mutations
RNA cleavage signal
mutations
Nonsense & frameshift
mutations
3’
Non coding regulatory
regions
Exons
Introns (InterVening
Sequences)
3’ cleavage mutant
deletion
RNA splicing mutant
transcription mutant
nonsense mutation
frameshift insertion
b- thalassemias
Main genetic mechanisms that contribute to the
phenotypic diversity of the b-thalassaemias.
Reading
HMG3 by T Strachan & AP Read : Chapter 14
AND/OR
Genetics by Hartwell (2e) chapter 11
References on Cystic fibrosis:
Science (1989) vol 245 pg 1059 by JM Rommens et al (CF mapping)
J. Biol Chem (2000) vol 275 No 6 pp 3729 by MH Akabas (CFTR)
Optional Reading on Molecular medicine
Nature (May2004) Vol 429 Insight series
•
human genomics and medicine pp439 (editorial)
•
Mapping complex disease loci in whole genome
studies by CS Carlson et al pp446-452