Lecture 9 Mutagensis and Mutation- Dr. Faisal Al-Allaf-md-y1-2011

/ /١٤٤٤ ٠٩ ٢٤ Dr. Faisal Al-Allaf, [email protected] 1

Dr. Faisal Al-AllafAssistant Professor of Genetics and Molecular Medicine

Umm Al-Qura University Faculty of Medicine, Makkah, Saudi Arabia

[email protected]/Fax: 5270000 Ext: 4198

Mutagenesis and Mutations


Mutagenesis and Mutations Where do mutants come from? How do mutations occur naturally? Numerical aberrations

Aneuploidy, Polyploidy, Monosomy, Trisomy

Structural aberrations Deletions, ring chromosome,

duplications, inversions, and translocations

Mosaicism, Chimarism, Lyonization

Mutations at the DNA level Base substitutions

Insertions Duplication Deletions

Functional effects of mutations on the proteins


Mutations can be inherited, spontaneous or induced

Mutation is a permanent change in nucleotide sequence.

Inherited

Induced mutations are due to exposure to environmental agents which causes or increases the rate of mutation (mutagens). Mutagens can be:

Physical Chemical Biological

Spontaneous or Naturally occurring mutations are thought to arise through:

Errors in chromosomal division (Non disjunction)

Errors in DNA replication Faulty DNA repair

Assault from environmental mutagens

DNA is damaged approximately 10 000 times per cell per day.

DNA damage will lead to disease.

Smoke

Pyrolysed(burnt )food

Radiation UV lightChemicals

DNA Damage

Disease (e.g. cancer)

DNA is susceptible to damage from environmental mutagens


Categories of environmental agent that cause damage to living cells


Chromosomal disorders It can be at chromosomal or DNA levels.

Gross lesions (chromosomal accounts for less than 8%)

Micro-lesions (molecular accounts for more than 92%)

At the chromosomal level: Numerical abnormalities include

Aneuploidy (Monosomy, Trisomy and Tetrasomy) and Polyploidy (Triploidy and Tetraploidy)

Structural abnormalities includes translocations, deletions, insertions, inversions and rings formation

Cell fusion, Chimerism, and mosaicism (Mixoploidy). Different chromosome constitution in two or more cell line


Numerical aberrations Euploidy means that the chromosome number

per body cell is an integral multiple of the haploid number, N=23, aneuploidy that it is other than an integral multiple.

Diploidy describes the normal situation, 2N = 46 chromosomes.

1. Extra haploid sets )Polyploidy( refers to multiples of haploid number (e.g. triploidy, 3N =69).

2. Extra single chromosome )Trisomy( (2N+1) is presence of three copies of one chromosome, (2N-1) is called monosomy.

3. Missing single chromosome )monosomis( are nearly always incompatible with survival (lethal) except for XO.


Mosaicism and Chimerism )Mixoploidy(

Mosaicism: presence within an individual of 2 or more genetically different cells issued from a single zygote (following a single fertilization event)

A mosaic is an individual with multiple cell lines (which exhibit different genotypes) that arise from a single zygote

Chimerism: presence within an individual of 2 or more genetically different cells issued from different zygotes (result from multiple fertilizations )

Both yield incidence patterns that depart from general rules


Mechanisms leading to numerical chromosomal abnormalities

Polyploidy arises as a result of:

1. Fertilization by two sperms

2. A diploid sperm due to failure in meiosis

3. A diploid ovum due to a failure in meiosis

Monosomies and Trisomies may result from

1. Non-disjunction or

2. Anaphase lag


Non-disjunctionNullisomic, monosomic and disomic gametes


Structural aberrations Structural aberrations include:

Translocations Deletions Ring chromosomes Duplications Inversions Isochromosomes Fragile sites

Most structural aberrations result from Chromosomal breakage occurs close together and

enzymatic repair mechanisms link the wrong ends

Unequal exchange between homologous repeated sequences on the same or different chromosomes


Ring chromosomes )code: r( If two breaks occur in the same

chromosome the broken ends can fuse as a ring

Acentric are lost, but if the ring contains a centromere it can survive subsequent cell division

46,XY,r(21)


Duplications )code: dup( Duplication is the presence of two

adjacent copies of chromosomal segment and can be either ‘direct’ (or ‘tandem’ ), or ‘inverted’.

Duplications are more common, but generally less harmful than deletions.


Inversions )code: inv( Inversions arise from two chromosomal

breaks with end– to –end switching of the intervening segment.

If inversion includes the centromere it is pericentric )A(, if not, the inversion is paracentric )B(.

46,XX, inv)9(, )p12q12(


Translocations )code: ‘rob’ or ‘t’ or ‘ins’(

A translocation involves transposition of chromosome material usually between chromosomes (non homologous).

Three types are recognized: 1. Centric fusion or ‘Robertsonian’ 2. Reciprocal 3. Insertional

Centric fusion or ‘Robertsonian translocations’ )code: ‘rob’( arises from breaks at or near the centromeres of two chromosomes, followed by their fusion

46,XY,t)2;4()p23;q25(, male with receprocal translocation involving the short arm of chromosome 2 at region 2 band 3 and the long arm of chromosome 4 at region 2 band 5


Mutations at the molecular level

At the molecular level: range from single base substitutions, through insertions and deletions of single or multiple bases.

It can occur in somatic or germline cells. Mutations which occur in germline

cells (gametes) are inherited and therefore pass to future generations unless it affects fertility or survival.

Mutations which occur in somatic cells may cause adult-onset disease such as cancer but cannot be transmitted to offspring.


Mutations Mutations drive evolution but can

also be pathogenic.

A conservative estimate suggests that each individual carries up to six lethal or semi-lethal recessive mutant alleles that in homozygous state would have very serious effects.

Mutations can occur either in non-coding or coding sequences, it is only when they occur in the latter that they are recognized as an inherited disorder or disease.


Types of mutations Base substitutions

Synonymous or Silent mutations Nonsense mutation Missense mutation Readthrough mutation

Insertions may cause frameshift

Duplication duplications of part of a gene or DNA sequence

Deletions may cause frameshift

Splicing mutations may cause truncation


Base substitutions Substitution is the replacement of a

single nucleotide by another with no net gain or loss of chromosomal material.

Base substitutions are most prevalent and missense mutations accounts for nearly half of all mutations.

If the substitution involves replacement by the same type of nucleotide, i.e. a pyrimidine for a pyrimidine (C for T or vice versa) or a purine for a purine (A for G or vice versa), this is termed a transition.

Substitution of a pyrimidine by a purine or vice versa is termed a transversion.


Base substitutions Point mutations may arise as a result

of mistakes in DNA replication, mistakes in repair following DNA damage, or most commonly as the result of the spontaneous deamination of methylated cytosine to thymine.

Point mutations may be silent or deleterious depending upon the site.

Rarely, a mutation may be advantageous and favored by natural selection.


Synonymous or silent mutations Based on the structural effects of the

mutations on the polypeptide sequence of the encoded protein, mutations can also be subdivided into two main groups, being either synonymous or non- synonymous mutations.

Synonymous or Silent mutations: when a mutation does not alter the polypeptide product of the gene

Because of the degeneracy of the genetic code, a single base pair substitution, particularly if it occurs in the third position of a codon, will often result in another triplet which codes for the same amino acid with no alteration in the properties of the resulting protein.


Non synonymous mutation Non synonymous mutation occurs

when the mutation leads to an alteration in the encoded polypeptide.

Alteration of the amino acid sequence of the protein product of a gene is likely to result in abnormal function. Occur less frequent than

synonymous mutation

Non-synonymous mutations can occur in one of two main ways. Nonsense mutation Missense mutation

Conservative Non conservative Readthrough


Missense mutations A single bp substitution can

result in coding for a different amino acid and the synthesis of an altered protein, a so-called missense mutation.

Examples: Cystic fibrosis: CFTR G551D Sickle cell anaemia: defective

haemoglobin due to mutation in β-globin gene


Nonsense mutations A substitution which leads to the

generation of one of the stop codons will result in premature termination of translation of a peptide chain or what is termed a nonsense mutation.

Premature termination of translation produces truncated (shortened) protein

Almost always pathogenic due to loss of large amount of the normal protein


Deletions and insertions Deletions and insertions/duplications

involves the loss or gain of one or more nucleotides.

The addition or deletion of a single nucleotide will cause the amino acids to be grouped incorrectly.

The effects of deletion and insertion on protein depend on:

The amount of material lost Whether the reading frame is affected


Deletions and insertions of multiples of 3 nucleotides

Small insertions and deletions in coding sequences of multiples of 3 nucleotides will retain the frame (NOT cause frameshift but extra amino acid(s) will be encoded)


Deletions and insertions of multiples of 3 nucleotides

ك ع غ ف ت ر س ك م م ل ج ك ا ع م ر ب م ل ق م ر ف مل ي س ه ط ح ر خ ص د ي و ل ب ع س ز ر ك ف ر ط س

سيل مكر حطه صخر مود كجل معا مبر مقل مفرعلي

مبر مكر مقل سيل ...مفر حطه صخر مود كجلعلي

صخر مكر مود كجل معا مبر مقل صلبمفرسيل عليحطه


Deletions and insertions of NOT multiples of 3 nucleotides

Small deletions and insertions in coding sequences of one, two or more nucleotides which are not a multiple of three will destroys the reading frame causing frameshift and may be premature terminations or readthrough.


Deletions and insertions of NOT multiples of 3 nucleotides

ك ع غ ف ت ر س ك م م ل ج ك ا ع م ر ب م ل ق م ر ف مل ي س ه ط ح ر خ ص د ي و ل ب ع س ز ر ك ف ر ط س

سيل مكر حطه صخر مود كجل معا مبر مقل مفرعلي

مق مكر يلع ممفر طهس خرح ودص جلم عاك برمليس

ص مكر مود كجل معا مبر مقل رحط بمفر خي لعل هسي


Mutations in non-coding DNA Mutations in non-coding DNA are

less likely to have a phenotypic effect. Exceptions include mutations in promoter sequences, or other regulatory regions. Mutations in in the splicing of introns, e.g. the highly conserved GT and AG dinucleotides at the end of introns.

Mutations in regulatory elements can affect the level of gene expression while mutations in splice junctions can result in coding sequences being lost or intronic sequences being added to the mRNA molecule.


Mutations in promoter and regulatory elements

It may lead to up-regulation or to down-regulation of transcription which may be detrimental


Stable and unstable mutations Mutations which transmitted unaltered are termed fixed or stable

mutations and mutations which undergo further alteration as they are transmitted in families are termed dynamic or unstable mutations.

Fixed/stable mutations are point mutation (single base pair substitutions, insertions, deletions or duplications of part of a gene or DNA sequence).

Unstable or dynamic mutations consist of triplet repeat expansion which, in affected persons, occur in increased copy number when compared to the general population.

Triplet repeat expansions are a type of unstable insertion that have been associated with a variety of neurological disorders.


Examples of unstable trinucleotide repeat expansions


Functional effects of mutations on the protein

The mutations effect can appear either through Loss-of-function or Gain-of-function

Loss-of-function mutations can result in either reduced activity or complete loss of the gene product. In the heterozygous state would be associated with half normal levels of the protein product. They may be dominantly or recessively inherited.

Gain-of-function mutations, are inherited dominantly and result in either increased levels of gene expression (simple gain of function) or the development of new protein function (dominant negative mutations).

Rarely, a mutation may be advantageous and favored by natural selection.


Summary of mutation types

Mutations and their effects on protein products

Class Group Type Effect on protein product

Stable/ fixedSynonymous Substitution Silent – same amino acid

Non-synonymousSubstitution 1.Missense Non-conservative Conservative

Altered amino acidAltered activity, function or stabilityNo effect

2. NonsenseStop codon – premature termination with loss of function/activity/stability

Deletions/insertion1.Multiple of 3 (codon)

2.Not multiple of 3

In frame deletion/insertion of one or more amino acid(s) in protein – may altered activity/ function/stability

Altered reading frame (frameshift), premature termination of protein – altered amino acid sequence, loss of function/activity/stability

Dynamic/unstable

Triplet repeatExpansion Altered gene expression, reduced transcription or translation, altered transcript – altered activity/ function/stability


References and Private Reading1. Emery’s Elements of Medical Genetics, 13th edition 2007, by Peter TURNPENNY and

Sian ELLARD. Churchill Livingstone ELSEVIER. ISBN: 978-0-7020-2917-2

2. Medical Genetics at a Glance, 2nd edition 2008, by Dorian PRITCHARD and Bruce KORF. Blackwell Publishing. ISBN: 978-1-4051-4846-7

3. Elsevier's Integrated Genetics, 2007, by Linda Adkison and Michael Brown. MOSBY ELSEVIER. ISBN: 978-0-323-04329-8

4. Genetics for Dummies, 2005, by Tara Robinson, Wiley Publishing, Inc. ISBN: 978-0-7645-9554-7

5. New Clinical Genetics, 2007, by Andrew Read and Dian Donnai. Scion. ISBN: 978-1-904842-31-6

6. Cell Biology and Genetics, Crash Course, 2nd edition 2006, by Manson, Jones, Morris, Michael STEEL and Dan HORTON-SZAR. MOSBY ELSEVIER. ISBN: 0-7234-3248-1

7. Human Molecular Genetics, 3rd edition, 2003, by STRACHAN T. and A. READ. Garland science/Taylor and Francis group. ISBN: 978-0-8153-4182-6

8. Human Genetics: Concepts and Applications, 7th edition 2007, by Ricki LEWIS. McGraw Hill international. ISBN: 978-0-07-110779-2

9. Genomes, 3rd edition 2006, by T.A. BROWN. Garland science, ISBN: 978-0-8153-4138-3

10. Colour Atlas of Genetics, 2007, by Eberhard Passarge. Thieme. ISBN: 978-1-58890-3365


Acknowledgments For the providers of all the educational materials

(video clips, pictures, diagrams and charts) including publishers, pharmaceutical companies or unknown internet users who made their material available for use, in this and other presentations, I offer heartfelt thanks and deep appreciation.

I feel particularly grateful to faculty, staff, and our brilliant students who provided a unique intellectual and wonderful environment for work.

Lecture 9 Mutagensis and Mutation- Dr. Faisal Al-Allaf-md-y1-2011

Health & Medicine

chromosome number

single chromosome trisomy

ring chromosome

inherited induced mutations

chromosomal division

chromosomal level

chromosomal disordersit

multiple cell lines