Genetic MutationsSDK

October 8, 2013

OBJECTIVES 

By the end of this session the student should be able to:Define MutationFrequency of mutations in normal individualsClassify different types of mutationExplain the mechanism of mutationExplain the role of mutation in biodiversityExplain how mutations can cause severe diseasesGive examples of deletions, duplications, and insertions in genesDefine trinucleotide repeat expansions and how they cause neurological diseases

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What is a gene mutation?

Replacement or change of a nucleotide base with another, in one or both strands, or addition or deletion of a base pair in a DNA molecule .

Mutations are changes in genetic material(Nitrogenous bases) – changes in DNA code – thus a change in a gene(s)

In gene mutations, the DNA code will have a base (or more) missing, added, or exchanged in a codon.

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• Mutations can lead to missing or malformed proteins, and that can lead to disease.

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Gene mutation out come

Types of Mutations Germ-line mutations .Mutations that are inherited

from parents are called germ-line mutations. Acquired mutations. Mutations that are acquired

during your lifetime are called acquired mutations Some mutations happen during cell division, when

DNA gets duplicated. Still other mutations are caused when DNA gets

damaged by environmental factors, including UV radiation, chemicals, and viruses.

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How common are mutations?

Mutations occurs at a frequency of about 1 in every 1 billion base pairs

Everybody has about 5-10 potentially deadly mutations in our genes- in each cell of our body!

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When everyone has mutations, Why they are not always seen

They are not always seen because the mutation may have occurred in a section of DNA that

doesn’t make a protein.

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• Most inherited genetic diseases are recessive, which means that a person must inherit two copies of the mutated gene to inherit a disorder.

• This is one reason that marriage between close relatives is discouraged; two genetically similar adults are more likely to give a child two copies of a defective gene.

Diseases caused by just one copy of a defective gene are not manifested with the exception of Huntington's disease, which is rare and afflicted carriers are more likely to die before reproducing.

Mutations Outcome

The affected gene may still function.

Mutations may be harmful.

Mutations may be beneficial.

Mutations may have no effect on the organism. Mutations are a major source of genetic variation in a

population increasing biodiversity.

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Mutations a cause of Biodiversity

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Only mutations in gametes (egg & sperm) are

passed onto offspring(Germline Mutation).

Mutations in somatic cells (body cells) only

affect the body in which they occur and are not

passed onto offspring.

Does all mutations passed on to next generation?

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NO

Brain Work 1 A mutation may happen in any gene.

TRUE OR FALSE?

TRUETRUE

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Spontaneous and Induced MutationsSpontaneous: Occur spontaneously without

obvious reason.Induced mutations: caused by mutagens.Mutagens are the agent that causes the DNA

code to change (mutate) X-Ray, Chemicals, UV light, Radiation, etc

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Which of the following may cause mutations?A. Coffee

B. UV light (sun light)

C. Hair gel

D. Vaccines

Brain Work 2

UV Light (Sun Light)

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Types of Mutations1. Point mutations. A point mutation is a simple change in

one base of the gene sequence.

2. Frame shift mutations. one or more bases are inserted or deleted

• Original The fat cat ate the wee rat.• Point Mutation The fat cat ate the wet rat.

• Original The fat cat ate the wee rat.• Point Mutation The fat cat ate the wet rat.

Original. The fat cat ate the wee rat

Frame Shift The fat caa tet hew eer at

Original. The fat cat ate the wee rat

Frame Shift The fat caa tet hew eer at

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Morphological Types of Point mutations

1. Transitions. Transitions occur when a Purine is converted to a purine (A to G or G to A) Pyrimide is converted to a pyrimidine (T to C or C to T)

2. Transversion. A transversion results when Purine is converted to a pyrimidine (A to C or G to T) Pyrimidine is converted to a purine. (T to A or C to G)

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Types of Mutations according to their effects on the protein (or mRNA).

Silent Mutations. Mutation in a codons that produce same amino acid. These mutations affect the DNA but not the protein. Therefore they have no effect on the organism’s phenotype.

CUU CUCMissense Mutations. Missense mutations substitute one amino acid for another. Example. HbS, Sickle Cell Hemoglobin, is a change in the beta-globin gene, where a GAG codon is converted to GUG. GAG GUGNonsense mutations. convert an amino acid into a stop codon. The effect is to shorten the resulting protein. Sometimes this has only a little effect, however, often nonsense mutations result in completely non-functional proteins. UUU UAA\ UGA

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Frame-shift

In a frameshift mutation one or more bases are inserted, or deleted.

Because our cells read our DNA in three letter words, adding or removing one letter changes each subsequent word.

This type of mutation can make the DNA sequence meaningless .

For example: Original= THE FAT CAT ATE THE WEE RAT FRAMESHIFT= THE FAT CAA TET HEW EER AT.

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Mutations are a natural part of the cellular process reproduction. The cell has tools that catch and repair 99.9% of mutations.

TRUE OR FALSE?

TRUETRUE

Brain Work 3

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Most mutations are caught and repaired in the cell.

TRUE or FALSE?

Brain Work 4

TRUETRUE

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Classical Types of Point mutation Mutations

Point mutation occurs when the base sequence of a codon is changed. (ex. GCA is changed to GAA)

There are 3 types:

Also called frameshift mutations

•Substitution

•Deletion•Insertion

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Substitution

A substitution is a mutation that exchanges one base for another (i.e. a change in a single “chemical letter” such as switching an A to G.

For example:CTGGAGCTGGGG

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Normal DNA: CGA – TGC – ATC

Substitution Mutations

Mutated DNA: CGA – TGC – TTCAlanine – Threonine - stop

Alanine – Threonine - Lysine

What has happened to the DNA?The adenine was replaced with thymine

What will happen to the amino acids?

This is a substitution mutation

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The cat ate the rat

The hat ate the rat

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Clinical Examples

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Sickle Cell Anemia• Sickle cell anemia is the result of a (substitution) point

mutation in codon 6 of the -globin gene resulting in the substitution of amino acid glutamic acid by valine

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Under conditions of low oxygen tension, such as Following exercise or In an atmosphere containing a low oxygen level,

The following changes occur: The haemoglobin agglutinates to form insoluble rod-shaped polymers Red blood cells become distorted and sickle-shaped The sickle-shaped cells rupture easily causing haemolytic anaemia The sickle shaped cells tend to block capillaries interfering with the blood flow

to various organs.

Sickle Cell Anemia

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Thalassaemia

Substitution of C by U in mRNA that is coding globin chain of 146 amino acid.

Resulted in the formation of a stop signal UAG in place CAG of glutamate in codon number 39.

This result in a shortened globin chain containing only 39 instead of the normal 146 amino acids in the -globin protein chain.

This protein is functionally useless and is equivalent to absence of -globin gives clinical symptoms of thalassaemia,

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Thalassaemia

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Beta thalassaemia is a genetic disorder in which there is lack of beta globin.

It may be the result of: Deletion of the whole gene so that beta globin cannot not produced

(designated o ) Deletion of the promoter region so that transcription cannot occur

(designated o ) Deletion of a large part of the gene resulting in a grossly abnormal or

reduced synthesis functional protein (designated + )

Thalassemias

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Clinical Features of -Thalassaemia Haemoglobin A (α2 2) cannot be produced Hb F (α2 g2) is produced even in adults Hb A2 (α 2 d2) formation is increased Eerythrocytes are microcytic (small) due to lack of normal

haemoglobin Erythrocytes rupture easily causing severe haemolytic anaemia,

requiring repeated blood transfusions The bone marrow expands trying to compensate by increasing

haemopoiesis.

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The bones of the face and skull are thickened causing a characteristic facial appearance

The spleen and liver enlarge because haemopoietic tissue forms in them

Excess iron accumulates in the blood and is deposited in the heart, liver, pancreas and other organs (this is because of repeated transfusions while no blood is actually lost from the body)

Children have delayed growth and development and are prone to repeated infections

Clinical Features of -Thalassaemia

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Point Mutation In alpha-Globin Gene “Elongated α Globin Chain, Haemoglobin Constant spring \Wayne

Hb”

Here the stop codon UAA at position 142 in the alpha (-) globin gene was substituted by the codon for glutamine.

Translation of the protein thus continued until a stop codon was encountered at codon 173.

The -globin was considerably elongated, resulting in a variant of haemoglobin termed Haemoglobin Constant spring\ Wayne Hb.

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Elongated α Globin Chain Or Haemoglobin Wayne

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Insertion

Insertions are mutations in which extra base pairs are inserted into a new place in the DNA.

CTGGAG

CTGGCCTAG

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This is an insertion mutation, also a type

of frameshift mutation.

Normal DNA: CGA – TGC – ATC

Insertion Mutations

Mutated DNA: CGA – TAG – CAT – C

Alanine – Threonine – stop

Alanine – Isoleucine – Valine

What will happen to the amino acids?

An adenine was inserted thereby pushing all the other bases over a frame.

What has happenedto the DNA?

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The cca tat eth era t

The cat ate the rat

Insertion Mutations

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Haemophilia A An X-linked recessive disorder in which blood clotting does not

occur due to deficiency of clotting factor VIII. In most cases the mutation is the result of insertion of a large

segment, consisting of about 3800 bp, in the coding region of the factor VIII.

This results in total inactivation of the protein.

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Inherited as a sex linked recessive trait with bleeding manifestations only in males.

Genes which control factor VIII and IX production are located on the x chromosome

Affected male marries a normal female: none of sons will be affected, all daughters will be carriers

Female carrier marries normal male: 50% chance sons will be affected and 50% chance daughters will be carriers

Haemophilia A

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Deletion

CTGGAG CT AG

Deletions involve removal of one or more base pairs. They vary greatly in size from deletion of a single base

to deletion of a whole gene. The clinical effects often depend on the size and

location of the deleted part of the gene.  

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What will happen to the amino acids?

Mutated DNA: CGA – TCA- TC

A guanine was deleted, thereby pushing all the bases down a frame.

Alanine – Threonine – stop

Alanine – Serine

Deletion MutationsNormal DNA: CGA – TGC – ATC

This is called a deletion mutation, also a type of frameshift mutation.

What has happenedto the DNA?

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Muscular Dystrophy

Deletions of Dystrophin Gene

Dystrophin is a protein that is an important component of skeletal muscle.

The dystrophin gene is located on the p arm of the X chromosome (Xp21.2).

It is a very large gene spanning 2.5 million bp of genomic DNA and consists of 79 exons coding for a protein of approximately 3600 amino acids (11kb).

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Muscular Dystrophy

Deletions of Dystrophin Gene

Deletion of the whole or most of the dystrophin geneDystrophin may not be produced at all Or produce in in abnormal forms,

Resulting in Duchenne muscular dystrophy This is a severe X-linked recessive disorder that affects boys and is

transmitted by carrier females. X linked Recessive disorder. In affected boys there is almost complete lack of dystrophin, muscle

weakness beginning in childhood and increasing progressively in severity so that the individual is wheel-chair bound at the age of about 15 years.

Death usually ensues in the early twenties due to respiratory muscle involvement.

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X linked Recessive disorder

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Becker Muscular Dystrophy(BMD)Deletions involving a small non-critical part of the gene result

in altered dystrophin.This causes the clinical condition of Becker muscular

dystrophy

in BMD muscle weakness begins in adolescence and is very slowly progressive, and affected individuals may lead an almost normal life.

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Cystic Fibrosis.

Cystic fibrosis (CF) is a genetic condition that affects many organs in the body: especially the lungs, pancreas and sweat glands.

Cystic fibrosis is caused by a mutation in the Cystic Fibrosis Trans-membrane Regulator (CFTR) gene, that is located on chromosome 7.

This gene Produces a trans-membrane protein that regulates the flow of chloride ions into the cells.

The most common mutation is termed the ∆508 mutation, which is a deletion of a single codon at position number 508 in exon 10 of the CFTR gene.

Homozygotes(both parents need to be the carriers of the defective gene) for a ∆508 mutation have cystic fibrosis disease.

Gene That Encodes CFTR The gene that encodes the CFTR protein is found on

the human chromosome 7, on the long arm at position q31.2.

Mutations consist of replacements, duplications, deletions or shortenings in the CFTR gene.

This may result in proteins that may not function, work less effectively, are more quickly degraded, or are present in inadequate numbers

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Cystic Fibrosis.

The defective gene produces a defective protein leading to a blockage in the transportation of the salt, thus leading to production of thick, sticky mucus.

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Presentation

The secretion of very thick, sticky mucus causes Obstruction of the bronchi and predisposing to pulmonary infections, Pancreatic duct obstruction leads to problems with digestion. Intestinal and liver problems and When it blocks the sweat glands, it leads to loss of excessive salt

through sweat. This leads to imbalance of minerals within the body.

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• Paroxysmal nocturnal hemoglobinuria is a disorder of blood cells in which absence of specific molecule(GPI anchor protein, CD55 [Decay Accelerating Factor (DAF)], and CD59 [Membrane Inhibitor of Complement Lysis (MIRL)] on the surface of the cells (particularly RBC) leads to premature destruction of the cells by the complement system.

• This destruction is intermittent (paroxysmal).

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Paroxysmal Nocturnal Hemoglobinurea

Paroxysmal Nocturnal Hemoglobinurea

GPI anchor protein on the surface of red blood cells produced by the bone marrow stem cells

This is caused by a mutation of PIG-A gene, PIG-A gene is present on the X chromosome important in

making GPI protein anchors. Defect makes the red cells in susceptible to destruction

by the complement system.

The PIG-A mutation occurs in a bone marrow stem cell.

All the blood cells made by this defective stem cell are deficient in GPI-anchored proteins. (glycosyl-phosphatidylinositol GPI).

The GPI-anchored proteins are present on the surface of red blood cells that protect red cells from the activity of the complement system.

When they are absent , no protection from complement and this lead to intravascular haemolysis.

Paroxysmal Nocturnal Hemoglobinurea

Genetics• PIGA gene(phosphatidylinositol glycan class A) is

present in in the X chromosome and can have several mutations, from deletions to point mutations.

• The genetic mutation leading to the inability to synthesize the glycosyl-phosphatidylinositol (GPI) anchor proteine.

RBC Lysis

Normal RBCsNormal RBCs PNH RBCPNH RBC

Intact RBCIntact RBC

ComplementComplementActivationActivation

Lysed PNH RBCs and free hemoglobin in the plasmaLysed PNH RBCs and free hemoglobin in the plasma

Chronic HemolysisChronic Hemolysis

CD59CD59

Lectin Pathway Classical Pathway Alternative Pathway Carbohydrate structure Antibody bound to antigen Microbial membranes Pathogen/Damaged cell Immune complexes Bacterial LPS C3 convertase C4b2a, C3bBb C3 C3a Weak anaphylatoxin

C3b CD55 C5 convertases C4b2a3b, C3bBb3b CD55 C5 C5a Inflammation Cell Activation C6,C7,C8,C9 CD59

C5b-9 Microorganism Destruction Red Blood Cell Lysis Platelet Activation

Role of CD55 &CD 59

• The term "nocturnal" refers to the belief that hemolysis is triggered by acidosis during sleep and activates complement to hemolyze an unprotected and abnormal RBC membrane.

• However, this observation was later disproved. Hemolysis has been shown to occur throughout the day and is not actually paroxysmal, but the urine concentrated overnight produces the dramatic change in color.

Paroxysmal Nocturnal Hemoglobinurea

Deletion of 6 codons in the -globin gene resulting in a variant Hemoglobin

The codons 92 to 97 of the -globin gene are deleted. This results in a shortened -globin protein that produces

a haemoglobin variant termed Haemoglobin Gun Hill. In homozygotes it produces mild clinical; symptoms.

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Codon no Codon Amino acid

91 CUG Leucine

92 CAC Histidine

93 UGU Cysteine

94 GAC Aspartic acid

95 AAG Lysine

96 CUG Leucine

97 CAC Histidine

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Frame shift mutationsFrame shift mutations involve a deletion or insertion of

one or two base pairs within a coding sequence of a gene.

As the coding message is read in triplets codons and deletions will altered the the reading frame of mRNA

This results in a non-sense sequence of amino acids till stop codon. Original= THE FAT CAT ATE THE WEE RAT

Frame-shift= THE FAT CAA TET HEW EER AT

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Frame Shift Mutations(Deletion)

An example occurs in the -globin gene in which one nucleotide of codon 39 is deleted leads to altered sequence.

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Frame Shift Mutations(Insertion)Insertion of a sequence of bases into a coding sequence of

a gene. Sometimes a whole gene sequence may be duplicated.

1. Hereditary motor and sensory neuropathy type I

A DNA segment at locus 17p11 is duplicated.

2. Tay-Sachs Disease

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Tay-Sachs Disease Tay-sachs disease is an autosomal recessive disorder This genetic defect is located in the HEXA (hexosaminidase)

gene, which is found on chromosome 15. The hexa gene makes part of an enzyme called beta-

hexosaminidase A This enzyme helps break down a fatty substance called GM2

ganglioside in nerve cells. Mutations in the HEXA gene disrupt the activity of beta-

hexosaminidase A, preventing the breakdown of the fatty substances.

As a result, the fatty substances accumulate to deadly levels in the brain and spinal cord.

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Tay-Sachs Disease

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Tay-Sachs Disease

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Tay-Sachs Disease

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Loss of hearingPhysical and mental retardationSeizuresDementiaAnd most noticeably detected by the red dots it causes

on the retina of an individuals eye

Tay-Sachs Disease S\S

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Trinucleotide Repeat Expansions  Trinucleotides are triplets of nucleotides that are repeated

The number of repeats varies in different individuals. Trinucleotide repeats is - - - CAG CAG CAG CAG CAG - - - - Trinucleotide repeats are widespread in the genome, and may

occur in exons, introns, promoter sequences or non-coding regions.

They are perfectly normal and occur in all individuals However a mutation arises when the repeats become unstable and

undergo expansion, namely an increase in the number of repeats as they are transmitted from one generation to the next.

When the number of repeats exceeds a certain limit, clinical symptoms occur.

An example is the huntingtin gene, which, when mutated, causes Huntington's disease.

Normal range of (CAG)n is 11 to 34 Huntington's disease appear in individuals in whom the number

of repeats is greater than 37.

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Huntington's Disease

Autosomal Dominant InheritanceDue to an excess of C-A-G nucleotide repeats(>37) in

the HTT gene on the short arm of chromosome 4 forms (4p 16.3)Huntington's protein.

Huntington's protein has increase number of glutamine (polyglutamine).

Excess of repeats causes the protein to form aggregates that are deposited within the neurons causing neuronal degeneration

Affects brain and spinal cord, especially the basal ganglia.

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Trinucleotide Repeat Expansions

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Clinical Manifestations Most commonly appear in mid-forties If appear at a younger age, more severe Manifestations occur because of wasting away of brain cells.

1. Sudden jerky, involuntary movements throughout body2. Difficulties with balance and coordination3. Dysphasia 4. Hesitant/slurred speech5. Progressive dysfunction of intellectual and thought

processes (dementia)6. Cognitive deficits

a. Working memory lossb. Reduced capacity to plan, organize, and sequence

7. Restlessness, irritability 8. Depression or Euphoria

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What happens to a person who has a mutation?

Most of the time the mutation is harmless because 95% sections of DNA do not code for protein (junk DNA).

But sometimes the mutations can cause disorders such as Huntington’s disease and sickle cell anemia etc.

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Brain Work 5

Which of these is NOT a type of mutation.a) Point mutation

b) Flyaway mutation

c) Frameshift mutation

d) Nonsense mutation

Flyaway Mutation

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Brain Work 61. THE CAT SAW THE FAT RAT

2. THE CAT SAW THE RAT The change in Statement 1 to form Statement 2 is most

similar to what type of mutation?A.InsertionB.DeletionC. SubstitutionD. Frameshift

The correct answer is B deletion. Because the sentence is missing the word fat

which occurs in deletion as it is the removal of a section of DNA.

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Brain Work 7

5 AGAUCGAGU3 → 5’ACAUCGAGU3′ ′ ′The chain above represents three codons.

Which of the following changes would be expected in the amino acid chain if the mutation shown

above occurred?

A. The amino acid sequence would be shorter than expected.

B. The identity of one amino acid would change.C. The amino acid sequence would remain

unchanged.D. The identities of more than one amino acid

would change. The correct answer is B because according to the codon chart if G is switched by a C then one amino acid is affected because now instead of coding for Arginine it now codes for Threonine.

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Thank You

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