Genomics and health/disease
Part II: Genetic variation, inheritance and genetic
disorders
Assoc Prof Benjamin Kumwenda and Team
Adapted and presented by Samuel Gwayi,MSc
OBJECTIVES
Describe mutagenesis and explain how the cell tries to protect itself against it
Describe the different types of mutation/variation
Describe and explain the molecular basis of Mendel’s laws
Understand and explain different modes of inheritance
Explain the difference between monogenic and multifactorial disorders
Describe examples of monogenic disorders and multifactorial disorders
Refs:
WHO Report of the Advisory Committee on Health Research. Genomics and World
Health. WHO (2002).
Turner P, McLennan A, Bates A, White M. Bios Instant Notes: Molecular Biology.
Taylor & Francis Group (2005).
www.wikipedia.org
Genetic variation and mutation
• Only < 1% DNA sequence difference between individuals
• Accounts for phenotypic variability
• Variation in occurrence and susceptibility for diseases → genetic
predisposition
• Mutation is event that creates genetic variation
• Changes in DNA, introduced by errors during replication or cell
division, viruses, chemical agents, radiation
• ‘Heritable’ change in DNA
• In somatic cells, only the daughter cells (in that tissue) will
inherit mutation
• In germline (egg or sperm) cells, mutation will be inherited by
offspring
Mutagenesis
• Process by which genetic material is altered/changed in a stable manner, resulting
in a mutation
• may occur spontaneously in nature, or due to exposure to mutagens
• may also be done through experimental laboratory procedures
• Replication errors: mismatch, insertion, deletion
• 1/ 100 000 mistakes by DNA-polymerase
• Would be 60 000 errors per cell division
• BUT
►
Proofreading
 5’ → 3’ addition of nucleotides by DNA
polymerase has errors
 Exonuclease activity associated with
some DNA-polymerases
 3’ → 5’ removes wrong residue (base)
http://www.virology.ws/wp-content/uploads/2009/05/dna-polymerase-3.jpg
Mutagenesis
mismatches
2.Mismatch repair
 system for recognizing and repairing
erroneous insertion, deletion, and misincorporation of bases that can arise
during DNA replication and recombination,
as well as repairing some forms of DNA
damage.
 Mismatch repair enzymes recognize these
errors and cut wrong bases out
 Gap filled again with correct nucleotides
by DNA polymerase I and DNA ligase forms
bonds
• So in the end, error rate is low: high
fidelity of replication
 Mutation rate in human DNA
~ 1 /10
000 000, but differs per genomic region
 Still ~ 600 errors per cell division
• If repair mechanisms are damaged, this
will most likely lead to cancer
Mutagenesis
Physical mutagens
Ionizing radiation e.g. X-rays
UV-light
Chemical mutagens
Reactive oxygen species
Nitrous acid
Alkylating agents e.g. ethylnitrosourea (ENU)
Viruses
DNA by itself is chemically quite unstable and lesions can be generated
spontaneously
Base changes, strand breaks and other lesions can become fixed mutations
after replication or can block replication and cause cell death
Most chemical and physical mutagens are carcinogenic
E.g. UV-light – skin cancer
Mutation
• Remember base pairing
A,T,G,C
Mutation/variation
1 basepair change = point mutation
Two types:
Transitions: purine to purine or pyrimidine to pyrimidine
Transversions: purine to pyrimidine or pyrimidine to purine
= Single Nucleotide Polymorphism (SNP)
CCTAAACCCACCACTGGCCACATCCCA
CCTAAACCCACCAATGGCCACATCCCA
In protein coding sequence: can affect protein
Mutation
Silent mutation (synonymous substitution)
Does not result in a new amino acid in the protein sequence, eg. GCA or GCG codons in
the translated mRNA both mean (code for) arginine
3rd base of codon often no effect
Nonsense mutation (nonsynonymous substitution)
Alters the amino acid sequence of a protein
Forms a terminating codon and results in premature shortened protein
Effects are variable depending upon how much of the truncated protein is present and
is required for its function
Missense mutation:
a type of nonsynonymous substitution in which a single nucleotide change results in a
codon that codes for a different amino acid.
Sometimes ‘neutral’: new amino acid has almost same structure as normal one and no
effect on protein function
But often with serious effect when new amino acid changes
STOP GAIN vs STOP LOSS
Mutation
• E.g. Sickle-cell anemia:
• beta-globin gene on chromosome 11p15
• GAG in mRNA codes for glutamate residue
• if this is altered to GUG in mRNA, it results in a
valine residue in the protein chain
• Missense mutations may thus have very
serious consequences
Mutation/variation
• Insertions and deletions
CCTAAACCCACCATGGCCACATCCCA
CCTAAACCCACCATGGCCACATCCCA
CCTAAACCCACCAACTGGCCACATCCCA
CCTAAACCCACGGCCACATCCCA
• Often in areas with repeats: ‘strand-slipping’ → repeat length var
CCTACTACTACTACTACTACTACTCCA
 E.g. Huntington’s disease
► neurodegenerative
CCTACTACTACTACTACTACTACTACTACTCCA
CCTACTACTACTACTACTACTCCA
disorder causing uncontrolled
movement, loss of cognition and emotional problems
► HTT gene (huntingtin) on chromosome 4
► trinucleotide repeats (CAGCAGCAG...) in normal
individual 10-35, in individual with Huntington 40->120
► CAG codes for glutamin in protein chain
► longer protein is cut into smaller toxic fragments that
accumulate in neurons
► cause dysfunction and cell death
► Disease
starts around age 30-40
Mutation
• Frameshift mutation
 Disrupts the ‘reading frame’ = gene sequence containing the codons that will be
translated into protein
 Insertion or deletion of number of bases in coding sequence that is not divisible by 3
 Causes dysfunctional protein
Structural DNA-variation
Chromosomal rearrangement
Copy-number Variation (CNV)
Deletions and duplications of long stretches of DNA
CCTAAACCCACCACTGGCCACATCCCA
CCTAAACCCACCACTGGCCACATCCCA
CCTAAACCCACCACTGGCCACATCCCA
CCTAAACCCACCACTGGCCACATCCCA
CCTAAACCCACCACTGGCCACATCCCA
CCTAAACCCACCACTGGCCACATCCCA
X
Translocation t(9:22)
Acute mylogenous leukemia
Duplication of repeats at Xq27.3
Fragile X syndrome (mental
retardation)
http://www.biology.iupui.edu/biocourses/n100/2k2humancsomaldisorders.html
Aneuploidy
failure of homologous chromosomes or sister chromatids to separate
properly during cell division.
Remarkably common, causing termination of at least 25% of human
conceptions
Aneuploidy
Cause of chromosomal disorders
E.g. Down Syndrome, Turner Syndrome
Driving force in cancer progression, virtually all cancer cells are aneuploid
Inheritance
Gregor Mendel
Quantitative breeding experiments
with pea plants
Genes and inheritance
• From his experimental data, Mendel deduced that an organism has two ‘genes’
for each inherited trait/characteristic
• So, each trait in the offspring is defined by two ‘genes’ inherited from both
parents
• 1 form of the gene, inherited on the maternal chromosome = maternal allele
• 1 form of the gene, inherited on the paternal chromosome = paternal allele
• Locus = location, certain spot or position on the chromosome
• E.g.: 7p15, bp 120 000-120 500
• Alleles = alternative forms of a gene through differences in DNA-sequence,
found at the same locus on a certain chromosome (homologous chromosomes
in 1 individual)
Genes and inheritance
►Phenotype = trait/characteristic
►Genotype = genetic make-up defining this trait
GENE LOCI
P
P
a
a
B
DOMINANT
allele
b
RECESSIVE
allele
GENOTYPE:
PP
aa
HOMOZYGOUS
for the
dominant allele
HOMOZYGOUS
for the
recessive allele
Bb
HETEROZYGOUS
First law
• The Law of Segregation states
that when an individual produces
gametes, the two copies of a gene
separate such that each gamete
receives only one copy of the gene
= one allele of the pair
• Molecular basis of the principle: 2
divisions in meiosis
• separation of homologous
chromosomes in division 1
• separation of chromatids in division
2
Second law
• The Law of Independent Assortment, also
known as "Inheritance Law", states that
alleles of different genes assort
independently of one another during
gamete formation
• Molecular basis: independent alignment of
pairs of homologous chromosomes
• Maternal and paternal chromosomes will be
distributed randomly into gametes
• So gamete has random mixture of genetic info
from ‘grandmother’ and ‘grandfather’
Genes and inheritance
Inheritance follows the rules of
probability
F1 GENOTYPES
Bb female
Bb male
Formation of eggs
Formation of sperm
Recessive allele: b
1/
B
1/
2
Only bb has this phenotype
With heterozygous parents: ¼ of
offspring
B
2
B
B
1/
b
1/
1/
2
b
4
B
B
1/
b
1/
4
b
b
F2 GENOTYPES
1/
4
4
2
b
Dominant allele: B
BB and Bb have the phenotype
With heterozygous parents: ¾ of
offspring (¼ BB, ½ Bb)
Codominance: b&B
Both alleles contribute to
phenotype
With heterozygous parents: ½ Bb
‘mixed’ phenotype
In Blood grouping
A and B are dominant, while O is recessive
• Consider a Mother with blood
group A and a father with blood B.
what are the possible offspring
genotype if the both parents are
1. Homozygous
2. heterozygous
• If a family has a child with blood
group genotype BO. What are
possible parents genotype
combinations?
Genetic disorders
Change/mutation in a gene causes a ‘disease phenotype’
1 gene ~ 1 disorder:
monogenic = single-gene = Mendelian disorder
Mendelian inheritance patterns:
Autosomal dominant - one copy of an abnormal gene (from at least one
parent) required to develop the genetic disorder
Autosomal recessive - 2 copies of an abnormal gene must be present in
order for the disease or trait to develop.
X-linked dominant - gene responsible for the genetic disorder is located
on the X chromosome, and only one copy of the allele is sufficient to
cause the disorder if one parent has the disorder.
X-linked recessive - an abnormal gene on X chromosome from each
parent is required for females to develop disorder since a female has two
X chromosomes.
Autosomal dominant inheritance
50% chance of disease (half of children) if 1 parent affected
Every generation
Equal chance of occurrence in male and female
Father-son transmission is possible
Examples: Huntington’s disease, polycystic kidney disease, familial
hypercholesterolaemia, myotonic dystrophy,…
Autosomal recessive inheritance
Carriers that do not express disease
Healthy parents can have diseased child if both are carriers
Skips generations
Equal chance of occurrence in male and female
Father-son transmission is possible
Examples: sickle cell anemia, thalassemia, cystic fibrosis, spinal muscular atrophy,
phenylketonuria, …
X(sex)-linked inheritance
• Never father to son transmission
X(sex)-linked inheritance
Recessive
Dominant
• Most sex-linked human disorders due to recessive alleles
• A male receives a single X-linked allele from his mother, and will
have the disorder, while a female has to receive the allele from
both parents to be affected
• Examples: haemophilia, red-green color blindness, Duchenne
muscular dystrophy,…
Mitochondrial inheritance
• Mitochondria have own circular DNA in multiple copies
• Only inherited from mother’s egg
• Trait can only be transmitted from mother to child
• Passes on to all offspring
• Primary function of mitochondria is energy production → diseases will
affect organs with high-energy use such as heart, skeletal muscle, liver,
and kidneys
‘Mendelian’ monogenic disorders
• Single-gene disorders with simple Mendelian inheritance
• About 5000 different known
• But each one quite rare
• Examples: inborn errors of metabolism (AR or XR) (PDH deficiency, G6PD deficiency,
glycogen storage diseases,…), hemoglobin disorders (AR) (sickle cell anemia,
thalassemias,..), cystic fibrosis (AR), oculocutanuous albinism (AR), haemophilia (XR),
Huntington’s disease (AD), osteogenesis imperfecta (AD), neurofibromatosis (AD)
• Global prevalence of all together = 10/1000 births (1%)
Thalassemias
• Most common inherited single gene disorders in the world
• Absence or decreased production of hemoglobin leading to microcytic
anemia
• Cause: mutations in or deletion of globin gene(s)
• Alpha thalassemia: 2 genes HBA1 & HBA2 (4 alleles)
• Beta thalassemias: 1 gene HBB (2 alleles)
• Disease severity varies
• From no symptoms to blood transfusions needed throughout life (every 4 to
6 weeks: lot of donated blood needed)
• Depends on effect of mutation (no or decreased chain production) and on
number of alleles affected
• Each mutation inherited in simple Mendelian recessive manner
• Carriers disease free but gives protection against malaria
Beta-thalassemia
http://wiki.medpedia.com/Image:Thalassemia_beta.jpg?filetimestamp=20080805214700#file
Alpha-thalassemia
http://medpediamedia.com/u/Thalassemia_alpha.jpg/Thalassemia_alpha.jpg
Thalassemia
diagnosis
Complete blood count, blood smear: microscopic inspection of RBCs,
hemoglobin electrophoresis
Genetic testing & counselling
Carrier detection
Population screening
Neonatal screening
Prenatal diagnosis, preimplantation diagnosis
Beta-thalassemia: HBB gene on 11p
sequencing of ‘whole’ gene: only most variable parts where all mutations
until now have been found will be sequenced
Many different mutations possible
Also detects sickle cell anemia (point mutation in codon6 of exon1)
Alpha-thalassemia: HBA genes on 16p
PCR to detect deletions
Sequencing of parts of genes to detect mutations
Preimplantation
diagnostics
http://www.scq.ubc.ca/wp-content/uploads/2006/07/prenatalGD.gif
Osteogenesis imperfecta
Autosomal dominant disorder
Prevalence: 1/10 000
Defects in the formation of bone caused by
defects in collagen type I
 Collagen type I is the most abundant protein in bone, skin, and
other connective tissues that provide structure and strength to the
body
 Patients have bones that break easily and irregular connective
tissue
 Other symptoms may include short stature, hearing loss, abnormal
tooth development
Type I collagen
 3 subunits that form a triple helix: 2 alpha1-chains and 1 alpha2chain
 mature collagen molecules arrange themselves into long, thin
collagen fibrils
Osteogenesis imperfecta
Genetic defect: mutations in COL1A1 or COL1A2 genes
 COL1A1 on chromosome 17q encodes the pro-alpha1 chain
 COL1A2 on chromosome 7q encodes the pro-alpha2 chain
Hundreds of different disease-causing mutations have been
detected in these genes, mainly point mutations and small
deletions
Mutations can be inherited from a parent or be sporadic
(new mutation during meiosis)
Genetic diagnosis:
 PCR amplification of COL1A1 and COL1A2 genes
 Sequencing of the genes for mutation analysis
P GENERATION
Complex genetic
inheritance
aabbcc
AABBCC
(very light) (very dark)
F1 GENERATION
Eggs
F2 GENERATION
Sperm
• Single characteristic can be
influenced by many genes
• ‘Polygenic trait’
Fraction of population
AaBbCc AaBbCc
Skin pigmentation
Multifactorial or complex disorders
• Interplay between different genetic (polygenic) and environmental
factors
• Different ‘risk factors’ contribute to disease development
• Genetic predisposition + environmental factors → disease
Viral/bacterial infections
Cholesterol metabolism
genes
Cigarette smoking
BRCA genes
Radiation exposure
Atherosclerosis
Diet
Breast cancer
Platelet aggregation
factor genes
Age
Tumor supressor genes
38 genetic variants associated
Little physical
exercise
Increased fat intake
Obesity
Cardiovascular disease
Type 2 diabetes
Age
Asante Sana