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Mai M Abd El-Aziz

Mutation Detection Monday 9 th November 2009. Describe five different genetic mutations and explain the molecular mechanisms that give rise to these mutations. Illustrate your presentation with examples of genetic disorders caused by each of the mutation types?. Mai M Abd El-Aziz.

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Mai M Abd El-Aziz

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  1. Mutation Detection Monday 9th November 2009 Describe five different genetic mutations and explain the molecular mechanisms that give rise to these mutations. Illustrate your presentation with examples of genetic disorders caused by each of the mutation types? Mai M Abd El-Aziz

  2. Genetic Mutation • Mutation is a change in the nucleotide sequence of the DNA (nuclear/mitochondrial) leading to disruption of the involved gene function. • It should be differentiated from a polymorphism which can occur in at least 1% of the control population. • Can be classified according to their effect on structure, function, fitness and by pattern of inheritance.

  3. Classification of genetic mutations • Small scale mutations 1. Point mutations (endogenous mutations) • Silent mutations: code for the same aa • Missense mutations: code for a different aa • Nonsense mutations: code for a stop codon  the protein • Splice site changes: GT/AG sites 2. Insertions: adding one or more extra-nucleotides into the DNA (Repeat disorders) 3. Deletions: removing one or  nucleotide from the DNA

  4. Classification of genetic mutations (Cont.) • Large scale mutations 1. Amplifications/duplications Is any duplication of a region o DNA region that contains a gene 2. Juxtaposition • Chromosomal translocation • Interstitial deletion • Chromosomal inversion 3.Loss of heterozygosity/unipaternal disomy

  5. Mechanisms of point mutations • Spontaneous errors in DNA replication and repair DNA polymerase has an estimated error rate of one base/1000bp 10,000Mbp. These replication errors are kept in check by DNA repair enzymes but this mechanism doesn’t have 100% fidelity. • Depurination • Deamination • Transition A purine  purine, or a pyrimidine  pyrimidine • Transversion A purine  pyrimidine, or vice versa. • Oxidative damage to DNA bases (induced mutations) • DNA replication defect (incorporation of mismatched bases e.g. U instead of T) • Recombination defect

  6. Depurination& Deamination • Depurination: Loss of a purine base (A or G) to form an apurinic site (AP site). • Deamination: hydrolysis changes a normal base to an atypical base containing a keto group in place of the original amine group.

  7. Mechanisms of point mutations • Induced errors in DNA replication and repair • The UV light first causes two adjacent cytosine residues to form a dimer.  • The cytosine dimer could cause adenine (instead of the normal guanine) to be incorporated into the new strand.  • Subsequent DNA replication will produce CC to TT mutation. 

  8. Examples of diseases caused by base substitutions • Missense mutations • Sickle-cell disease  GAG>GTG (E>V) • Hereditary hemochromatosis  TGT/TGC>TAT/TAC (C>Y) • Cystic fibrosis c.G1642C E504Q • Nonsense mutations • Cystic fibrosis  c.G1619>A W496X • Silent mutations • Limb girdle muscular dystrophy patient GGC>GGT (G>G) Activation of a cryptic splice donor sequence aberrant splicing

  9. Examples of diseases caused by base substitutions (cont.) • Mutations affecting mRNA splicing • 1. Exon skippingif the mutation affect the splice acceptor site • DMD & Becker muscular dystrophy • del T in the polythymidine tract upstream of CFTR exon 9 • 2. Intron retentionif the mutation affect the splice donor site • 3. Cryptic splice sitethis is used if the wild type is mutated • NF1 gene intronic AG substit in intron 15 15 intronic nucleotide to be included in the mRNA • 4. splicing enhancers and silencers e.g. silent mutations

  10. Mechanisms of indel mutations • Replication slippage is common in repetitive sequences • Backward slippage insertion • Forward slippage  deletion

  11. Examples of diseases caused by deletions • Small deletions • a) 1 bp del e.g. CFTR • b) 2 bp del e.g. CFTR • c) 3 bp del e.g. FIX • d) 4 bp del e.g. APC • Large deletions e.g. DMD (involving Xp21.2)

  12. Examples of genetic diseases caused by Insertions • Frame shift insertions: Single or 2 bp insertion e.g. • 1.BRCA1: 5382insC • Non-frame shift insertions (Trinucleotide repeats) • Fragile X syndrome CGG repeats (5 UTR) • Friedrich’s ataxia (intonic GAA repeats) • SCA8, Myotonic dystrophy (CTG repeats 3 UTR) • Huntington disease CAG repeats • Blepharophimosis ptosis epicanthus inversus syndrome (BPES) caused by polyalanine expansions in FOXL2

  13. Mechanisms of gene amplification • (a) Unequal crossing over:which results in a recombination event in which the two recombining sites lie at nonidentical locations in the two parental DNA molecules. • (b) Retroposition, which occurs when a message RNA (mRNA) is retrotranscribed to complementary DNA (cDNA) and then inserted into the genome. • Chromosomal duplication • Segemental duplication

  14. Examples of diseases caused by gene duplication • CMT type I: 1.5 Mb 17p11.2-p12 tandem duplication PMP22 gene • Familial hypercholesterolemia 2Kb duplication due to juxtaposition of intron 6 to a partially duplicated exon 6

  15. Mechanisms of Chromosomal translocation • Caused by rearrangement of parts between nonhomologous chromosomes • Can lead to fusion gene (cancer) • Reciprocal • Non-reciprocal

  16. Examples of genetic diseases due to chromosomal translocation Cancer • t(8;14) – Burkitt’s lymphoma (c-myc) • t(8;21)(q22;q22) – AML • t(9;22)(q34;q11) –Philadelphia chromosome, CML • t(14;18)(q32;q21) – Follicular lymphoma (Bcl2) • t(1;11)(q42.1;q14.3)- Schezophrenia • Infertility • Down syndrome

  17. Mechanisms of chromosomal deletion • Losses from translocation • Chromosomal crossovers within a chromosomal inversion • Unequal crossing over • Breaking without rejoining

  18. Examples of genetic diseases due to large deletions • DMD dystrophin gene • Deletion of one of the -globin gene in  thalassemia • Wolf-Hirschhorn syndrome: partial 4p del • Jacobsen syndrome: term 11q deletion disorder

  19. Mechanisms of chromosomal inversion • Clustered inverted repeats with a high degree of sequence identity may be located within or close to a gene • This may predispose pairing of the repeats through a chromatid bending back upon itself • Subsequent chromatid breakage at the mispaired repeats and rejoining inversion

  20. Examples of chromosomal inversion • Hemophilia A: Inversion of exons 23-26 in factor VIII gene where the first 22 exons are separated from exons 23-26 and hence leading to inactivation of factor VIII protein.

  21. Mechanisms of Loss of heterozygosity • Loss of one allele, either by a deletion or recombination • Knudson two hypothesis of tumorigenesis Somatic mutation involving a tumor suppressor gene in addition to a germline mutation • Unipaternal disomy two copies of a chromosome or part of a chromosome are passed on from one parent no copies from the other parent

  22. Examples of genetic diseases due to Loss of heterozygosity • Retinblastoma • Breast cancer (BRCA) • Prader-Willi syndrome and Angelman syndrome can be caused by UPD or other errors in imprinting involving genes on 15q. • Beckwith-Wiedmann syndrome abnormalities of imprinted genes on 11p • Paternal UPD on chromosome 14 skeletal abnormalities, mental retardation and joint contractures.

  23. References • Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P. Molecular Biology of the Cell. Ed. 2002 • http://en.wikipedia.org/wiki/Mutation • Strachan and Read . 2004 Human Molecular Genetics 3 • http://www.cmgs.org/ • Hu and Worton. Partial gene duplication as cause of human disease 1992, Human mutation

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