Part 2

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.

2SDK 2012

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

3SDK 2012

Deletion

CTGGAG

C T A G

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.  

4SDK 2012

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?

5SDK 2012

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).

7SDK 2012

8SDK 2012

9SDK 2012

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.

10SDK 2012

X linked Recessive disorder

11SDK 2012

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.

12SDK 2012

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

SDK 2012 14

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.

15SDK 2012

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.

SDK 2012 16

• 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).

SDK 2012 17

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 RBCs PNH RBC

Intact RBC

ComplementActivation

Lysed PNH RBCs and free hemoglobin in the plasma

Chronic Hemolysis

CD59

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.

25SDK 2012

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

26SDK 2012

Frame shift mutations

Frame 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= T H E FAT C A T AT E T H E W E E R AT

Frame-shift= T H E FAT C A A T E T H E W E E R AT

27SDK 2012

Frame Shift Mutations(Deletion)

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

28SDK 2012

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

29SDK 2012

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.

The buildup of GM2 ganglioside causes progressive damage to the nerve cells.

30SDK 2012

Tay-Sachs Disease

31SDK 2012

Tay-Sachs Disease

32SDK 2012

Tay-Sachs Disease

33SDK 2012

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

34SDK 2012

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.

35SDK 2012

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.

36SDK 2012

Trinucleotide Repeat Expansions

37SDK 2012

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 body

2. Difficulties with balance and coordination

3. Dysphasia

4. Hesitant/slurred speech

5. Progressive dysfunction of intellectual and thought processes (dementia)

6. Cognitive deficits

a. Working memory loss

b. Reduced capacity to plan, organize, and sequence

7. Restlessness, irritability

8. Depression or Euphoria 38SDK 2012

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.

39SDK 2012

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

40SDK 2012

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. Insertion

B. Deletion

C. Substitution

D. 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.

41SDK 2012

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.

42SDK 2012

Brain Work 7

5 AGAUCGAGU3 → 5′ ′ ’AGAUGAAGU3′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 A because according to the codon chart if C G are switched by a G A then one codon will be affected because now instead of coding for Serine it now codes for Stop codon.

43SDK 2012

Thank You

44SDK 2012