OZZIE SKINNER MT(AMT), CLT, MAOM JAMES A. HALEY VA HOSPITAL MOLECULAR DIAGNOSTICS 03/23/2013

1. DNA Mutations p OZZIE SKINNER MT(AMT), CLT, MAOM JAMES A. HALEY VA HOSPITAL MOLECULAR DIAGNOSTICS 03/23/2013 Pocket Hercules

2. MUTATIONS What and how do mutations occur? A mutation is a permanent change in the DNA sequence of a gene • Can cause problems with growth, development, and function of the body’s systems • Mutations in a gene's DNA sequence can alter the amino acid sequence of the protein encoded by the gene • These changes can affect many genes along the chromosome and disrupt the proteins made from those genes • The effects of structural changes depend on their size and location, and whether any genetic material is gained or lost • Can affect the DNA sequence of a cell's genome or the DNA or RNA sequence of a virus

3. MUTATIONSWhat and how do mutations occur? • Mutations range in size from a single DNA building block (DNA base) to a large segment of a chromosome • Gene mutations occur in two ways: they can be inherited from a parent or acquired during a person’s lifetime • Hereditary (or germline) mutations: are passed from parent to child • Acquired (or somatic) mutations: occur in the DNA of individual cells at some time during a person’s life • These changes can be caused by environmental factors or if a mistake is made as DNA copies itself during cell division • Acquired mutations in somatic cells (cells other than sperm and egg cells) cannot be passed on to the next generation • Some genetic changes are very rare; others are common in the population • Genetic changes that occur in more than 1 percent of the population are called polymorphisms

4. MUTATIONSAffects on health and development • A gene is a recipe for making a protein • Each cell depends on thousands of proteins to do their jobs in the right places at the right times • By changing a gene’s instructions for making a protein, a mutation can cause the protein to malfunction or to be missing entirely • When a mutation alters a protein that plays a critical role in the body, it can disrupt normal development or cause a medical condition • A condition caused by mutations in one or more genes is called a genetic disorder • Incompatible with life: changes occur in genes that are essential for development, and often disrupt the development of an embryo in its earliest stages

5. MUTATIONSAffects on health and development • Only a small percentage of mutations cause genetic disorders • Some mutations alter a gene’s DNA base sequence but do not change the function of the protein made by the gene • Often, gene mutations that could cause a genetic disorder are repaired by certain enzymes before the gene is expressed (makes a protein) • Although most mutations that change protein sequences are harmful or neutral, some mutations have a positive effect on an organism (e.g. 32 base pair deletion in human CCR5/Delta 32 mutation/HIV resistance) “Mutations are often Bad, but,……………………..…..………… some can be Good”

6. MUTATIONS First, Smallpox, then The Black Death of 1348 • Came out of the East, and reached the shores of Italy in the spring of 1348. • Unleashed a rampage of death across Europe. • Within three years, anywhere between 25% and 50% of Europe's population had fallen victim to the pestilence. The plague presented itself in three interrelated forms: • The bubonic variant (the most common) + swellings or buboes that appeared on a victim's neck, armpits or groin. + Victims had a life expectancy of up to a week + Infected fleas that attached themselves to rats and then to humans spread this bubonic type of the plague. • Pneumonic plague +attacked the respiratory system and was spread by merely breathing the exhaled air of a victim. + life expectancy was measured in one or two days. Oriental rat flea “Regurgitates”

7. MUTATIONS The Black Death, 1348 • Septicemic version of the disease attacked the blood system • Having no defense and no understanding of the cause of the pestilence, the men, women and children caught in its onslaught were bewildered, panicked, and finally devastated • The plague disease, generally thought to be caused by Yersinia pestis. (Gram negative, bipolar staining coccobacilli) • The "black death" again devastated Europe during the 14th century over a 5 year period causing 25 million deaths • Who was to blame?

8. MUTATIONS The Black Death, 1348 - DNA evidence Survivors • However, some were able to survive the devastating Black Plague, such as the residents of Eyam, England • The village had quarantined itself in order to stop the plague from spreading beyond the village Investigation • In October 2010 a multinational medical group investigated the role of Yersinia pestis in the Black Death • They tested DNA and protein signatures specific for Y. pestis in human skeletons from widely distributed mass graves all over Europe that were associated archaeologically with the Black Death. • The new research ends the debate about the etiology of the Black Death, and decidedly demonstrates that Y. pestis was the causative agent of the epidemic plague that devastated Europe during the Middle Ages The obvious question, How did the residents of Eyam survive? .

9. MUTATIONS They developed the Delta 32, CCR5 Mutation in the gene for the human chemokine receptor 5 (CCR5) How HIV Enters the White Cells – The Mechanism Some rare strains of HIV infect via the CXCR4 receptor so by no means does a positive result for CCR5 delta 32 allow someone to engage in risky behavior Chromosome 3

10. MUTATIONS Mutations are caused by: • Radiation: • Alpha particles, Beta particles, Gamma rays, Cosmic rays, and X-rays, can wreak havoc on the molecules of your DNA • Cytosine and Thymine– are most vulnerable to radiation that can change their properties • The effect of the radiation may not be to kill the cell, but to alter its DNA code in a way that leaves the cell alive but with an error in the DNA blueprint • Viruses • Viruses that use RNA as their genetic material have rapid mutation rates • An advantage since these viruses will evolve constantly and rapidly, and thus evade the defensive responses of e.g. the human immune system.

11. MUTATIONS Mutations are caused by: • Transposons: (jumping genes) • Sequences of DNA that can move or transpose themselves to new positions within the genome of a single cell • The mechanism of transposition can be either "copy and paste" or "cut and paste“ • Oncogenes (genes that cause tumors) may be activated by the random reshuffling of transposons to a position adjacent to the oncogene • Barbara McClintockfound that they were responsible for a variety of types of gene mutations: insertions, deletions and translocations • In developing somatic tissues like corn kernels, a mutation that alters color will be passed on to all the descendant cells • This produces the variegated pattern which is so prized in Indian corn

12. MUTATIONS Mutations are caused by: During DNA replication: (SNP/Point mutation) • The replacement of a single base nucleotide with another nucleotide of the genetic material, DNA or RNA • Single-nucleotide polymorphisms may fall within coding sequences of genes, non-coding regions of genes, or in the intergenic regions (regions between genes) • Often the term point mutation also includes insertions or deletions of a single base pair

13. MUTATIONS Mutations are caused: • During DNA replication: (SNP/Point mutation) • One can categorize point mutations as follows: • Transitions: replacement of a purine base with another purine or replacement of a pyrimidine with another pyrimidine Purines are Pyrimidines are Adenine and GuanineCytosine , and Thymine • Purines are the double ringed nitrogenous bases • Pyrimidines are the single ringed nitrogenous bases • Transversions: replacement of a purine with a pyrimidine or vice versa. • Approximately two out of three single nucleotide polymorphisms (SNPs) are transitions. PUGA2

14. MUTATIONS Mutations are caused: • During DNA replication: (SNP/Point mutation) • Point mutations can also be categorized functionally: • Nonsense mutations: Premature stop codon in a DNA sequence or in the transcribed mRNA • Nonsense mutations are known to cause Cystic fibrosis, Beta thalassaemia , and Hurler syndrome • Start codon (ATG) Methionine • DNA stop codons are TAG, TAA, and TGA • RNA stop codons are UAG, UAA, and UGA Lack the enzyme to break down long chains of sugar molecules

15. MUTATIONS Mutations are caused by: • During DNA replication: (SNP/Point mutation) • Point mutations can also be categorized functionally: • Missense mutations: point mutation in which a single nucleotide is changed, resulting in a codon that codes for a different amino acid • Missense mutations are known to cause sickle-cell disease sickle-cell disease Glutamine Aspartic acid

16. MUTATIONS Mutations are caused by: • During DNA replication: (SNP/Point mutation) • Point mutations can also be categorized functionally: • Silent mutations: DNA mutations that do not result in a change to the amino acid sequence of a protein • A point mutation that changes a codon to a synonym (one that codes for the same amino acid) is also known as a synonymous mutation • About 50 genetic diseases have been linked to synonymous mutations • This includes silent mutations in CFTR, the gene that is linked with Cystic fibrosis.

17. MUTATIONS Mutations are caused by: • Deletions: • A mutation in which a part of a chromosome or a sequence of DNA is missing • Deletion is the loss of genetic material • Deletions can be caused by errors in chromosomal crossover during meiosis (process of cell division in sexually reproducing organisms) • Small deletions are less likely to be fatal; large deletions are usually fatal • Deletions genetic disorders can range from: Male infertility to Duchenne muscular dystrophy to “Cri du chat” Example Original The fat cat ate the wee rat. Deletion The fat ate the wee rat.

18. Notice anything unusual? Cri du chat syndrome / 5p- syndrome

19. MUTATIONS Mutations are caused by: • Duplications • Duplications occur when part of a chromosome is copied (duplicated) too many times • This type of chromosomal change results in extra copies of genetic material from the duplicated segment • Duplications arise from an event termed unequal crossing-over that occurs during meiosis between misaligned homologous chromosomes (corresponding chromosomes from each parent) • Duplications of oncogenes are a common cause of many types of cancer Example Original The fat cat ate the wee rat. Duplication The fat cat cat ate the wee rat.

20. MUTATIONS Mutations are caused by: • Inversions • An inversion involves the breakage of a chromosome in two places; the resulting piece of DNA is reversed and re-inserted into the chromosome • Genetic material may or may not be lost as a result of the chromosome breaks • An inversion that involves the chromosome’s constriction point (centromere) is called a pericentric inversion • An inversion that occurs in the long (q) arm or short (p) arm and does not involve the centromere is called a paracentric inversion Example • Original: The fat cat ate the wee rat. • Insertion: The fat tar eew eht eta tac.

21. MUTATIONS Mutations are caused by: • Inversions • On the left of the diagram is human chromosome number 5 (of 23) • On the right of the diagram is the matching chromosome from a chimpanzee • The region from p14.I to q14.I, the order has been flipped • Evolutionist calls this a inversion mutation between Man and Chimp • Overall, there are nine inversion mutations between man and chimpanzee Chromosome 5

22. MUTATIONS Mutations are caused by: • Isochromosomes • An isochromosome is a chromosome with two identical arms • Instead of one long (q) arm and one short (p) arm, an isochromosome has two long arms or two short arms • As a result, these abnormal chromosomes have an extra copy of some genes and are missing copies of other genes Associated diseases • Girls with Turner syndrome • Pallister-Killian syndrome • Some tumors

23. MUTATIONS Mutations are caused by: • Dicentric chromosomes: human chromosome 2 • Unlike normal chromosomes, which have a single constriction point (centromere), a dicentric chromosome contains two centromeres • Dicentric chromosomes result from the abnormal fusion of two chromosome pieces, each of which includes a centromere.  These structures are unstable and often involve a loss of some genetic material Associated diseases Myelodysplasia (MDS): A collection of conditions that result when bone marrow cells produce damaged cells that don't mature properly Acute Myeloid Leukemia (AML)

24. MUTATIONS Mutations are caused by: Ring chromosomes • Ring chromosomes usually occur when a chromosome breaks in two places and the ends of the chromosome arms fuse together to form a circular structure • The ring may or may not include the chromosome’s constriction point • In many cases, genetic material near the ends of the chromosome is lost • Ring chromosomes may form in cells following genetic damage by mutagens like radiation, they may also arise spontaneously during development Associated diseases • Ring chromosome 20 syndrome - epilepsy • Ring chromosome 14/13-mentalretardation • Ring chromosome 15 - dwarfism

25. MUTATIONS Mutations are caused by: • Translocation:juxtapositioning • When chromosomes break and the fragments rejoin to other chromosomes • A gene fusion may be created when the translocation joins two otherwise separated genes, the occurrence of which is common in cancer • It is detected on cytogeneticsor a karyotype of affected cells • Translocations can be balanced or unbalanced

26. MUTATIONS Mutations are caused by: • Translocation:balanced translocation • In a balancedtranslocation, pieces of chromosomes are rearranged but no genetic material is gained or lost in the cell • When a person's cells divide to create egg or sperm cells for reproduction, the egg or sperm cells can end up with extra genetic material or missing genetic material • This could lead to miscarriage depending on which chromosome and genes are affected • In about 4.5% of all couples with recurrent miscarriages, one or both parents has a balanced translocation

27. MUTATIONS Mutations are caused by: • Translocation:unbalanced translocation • An unbalancedtranslocationoccurs when a child inherits a chromosome with extra or missing genetic material from a parent with a balanced translocation • This is usually in the form of a loss of genetic material, called Monosomy and a gain to genetic material, which is Trisomy Monosomy/ Trisomy Both types of are “aneuploidy” : (having an abnormal number of chromosomes)

28. MUTATIONS Mutations are caused by: Trisomy 21/DownsS.

29. MUTATIONS Mutations are caused by: Monosomy X - genotype 45, X,

30. MUTATIONS Different Types of Mutations

31. MUTATIONS Myostatin-is a gene that limits muscle growth • The protein is a member of a superfamily of molecules called transforming growth factors beta (TGF-b) or growth differentiation factor 8 • Discovered that a mutation in the myostatin gene is responsible for increased muscle mass, also known as 'double muscling‘ • Scientists hope that myostatin inhibitors may be able to treat diseases such as muscular dystrophy • There is some concern about the possibility of abuse of myostatin inhibitors by athletes • In humans, myostatin is encoded by the MSTNgene • Animals lacking myostatin, have significantly larger muscles Transgenic Rainbow Trout

32. MUTATIONSMyostatin-related muscle hypertrophy Who wants some “rump roast?” “It taste God awful; You said it tasted like chicken”. You lied mama, you lied. If you see me coming at you, don’t run. My muscles are so tight, I couldn’t catch a cold. myostatin modulator Siblings

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