Ch 7 Mutation• A heritable change in DNA • Random• Source of genetic variation in a species• may be advantageous, deleterious, neutralMutation (+ sexual reproduction) + natural selection = evolutionAdaptation vs mutation• Adaptation (Lamarkian) – not proven– environment induces adaptive change– E. coli in ampicillin – some become resistant in response• Mutation– mutation is random– E. coli DNA mutates randomly – statistically, one will result in amp resistanceTypes of mutations• germ-line mutation– in sperm or egg (meiosis), will be passed to offspring• somatic mutation– body cells, not passed on• spontaneous mutation– natural aspect of DNA replication• induced– UV light, chemicals, X-ray etc.Point mutation• single base substitution, deletion, or additionPoint mutations1. Base pair substitutions– transition = G to A (purines) or T to C (pyrmidines) – transversion = ?2. Missense mutation– Wrong amino acid encoded– May lead to a neutral or non-conservative amino acid change– Example sickle cell – Codon CTC CAC glu val3. Nonsense mutation– Premature stop codon – Codon AAA  UAAlys stop4. Neutral mutation- point mutation does not amino acid folding- codon AAA  AGA lys arg (both basic aa)5. Silent mutation– No amino acid change, usually 3rdposition transitionscodon GCA  GCGarg arg6. Frameshift mutation– Shifts the reading frame --> scrambled message– Often leads to a premature stop codonThe sun was hot but the old man did not get his hat.Sickle cell disease results from a second position transversion which causes a missense mutationOther mutations• Larger insertions, trinucleotide repeats, deletions, duplications, translocation, inversionspg. 475 Fragile X Syndrome• Xq27.3 FMR-1 gene X-linked dominant• 2ndleading cause of mental retardation(1/4000 males, 1/6000 females)• CGG repeat– Normal = 6-54 repeats– Normal carriers = 55 – 200 repeats (premutation expansion of repeat) – Fragile X syndrome = 200 – 1300 repeats(Reprinted from Medical Genetics, 2nd ed., Jorde LB, et al, ©2000, with permission from Elsevier Science.)triplet repeat amplification• FMR-1  FMRP (mRNA binding protein)• FMRP regulates translation of mRNA• Affects synaptic plasticity – signaling between neurons for learning and memory• >200 repeats and FMR1 gene silenced retardation• Caused by slipping of DNA polymerase during replicationCauses of mutations1. Spontaneous mistakes by DNA polymerase = replication errorsMost repaired by DNA proofreading and repair systems2. Induced mutations• ChemicalsMustard gas, industrial waste, PCBs, DDT, ethidium bromide • Base analogues– Example: Bromouracil substitutes for thymine• Radiation – sun (UV), radon, X-ray– Breaks, dimers…– High doses kill cellsEthidium bromideThe Ames test• Indirect assay for carcinogens• Plate 108auxotrophic bacteria (his-)• Add filter disc with chemical• Look for revertants to his+• = mutations• Add rat liver extract to look at metabolismSpontaneous reversion rate 1000X higher rateto His+ colonies with mutagenRepair of mutations 1. Direct reversalExample: Mismatch repair– 3’  5’ proofreading by DNA polymerase Also, UV repair (bacteria) and alkylation repair (prokaryotes)2. Excision repairEx. base excision repair- single strand incision at both sides of error; - Excise mistake - DNA synthesis to replace the gap- Ligation- Also UV repair prokaryotes, methyl mismatch repair, SOS responseDefective excision repair: Xeroderma pigmentosum (1/250,000) 9q34.1• Autosomal recessive• Extreme sun sensitivity– High risk of skin cancer (1000X) average by age 8– Metastatic melanoma• Corneal damageGenetics and Cancer Ch 20Terms• Tumor (neoplasm) = mass of cells– Benign = cells in a single contained mass– Malignant = invades surrounding tissue• cells may break off and move = metastasis• Transformed cell – has lost normal growth controls– Loses contact inhibition• Oncogenesis = initiation of cancerThe cell cycle• Time from one cell division to the next• G1– Gap 1– Make organelles • S– Synthesize DNA• G2– Gap 2– Prepare to divideCheckpoints • Arrest cell cycle to repair damage, or send cell to apoptosis• G1 – Should cell continue to S?• G2-M– Is the DNA replicated– Is the cell large enough?• M– Are chromosomes attached to the spindle?cell cycle regulation (cyclin-Cdk)Cyclins – concentration cyclesCyclin-dependent kinases (Cdk)cyclin binds a Cdkthe Cdk phosphorylates another protein cell continues in cell cycleGrowth factors and receptorsGrowth factorsstimulates cells to divideGrowth factor receptorsbind growth factor at cell membranesignal transduction   cell division– Ex. EGFRSignal transducers– Ex. H-ras EGR  binds to EGFR  ras binds GTPcell divisionFig 20.3Signal transductionBalance between stimulatory and inhibitory signalsMutation in gene encoding regulatory protein can lead to neoplastic cell – Loss of control of cell cycleiGenetics Regulation of cell division in normal cells• Try iactivity in chapter 10 online – tracking down the cause of cancerCancer is genetic• Hereditary cancers– Predisposition genes– Ex. some forms of colon cancer~ 10 % of cancers• Sporadic cancers~90% of cancers• Descendents of cancerous cells all cancerous• Mutagens increase cancer risk• Certain mutations cause certain cancersGenes and cancer1. Proto-oncogenes (Harold Varmus and Michael Bishop Nobel Prize 1989)– Genes involved in cell cycle• If mutated  oncogene• Tumor cells exhibit oncogenes• Dominant mutation• Cancer is a “multi-hit” disease– Mutations in more than one cell cycle gene requiredProto-oncogenes– GFs– GFRs and other protein kinases– Signal transducers• Generate a signal cascade• Activated by GF binding to GFROncogene OverexpressionExpression in cell that does not normally express GFRExpression of mutant GFR or signal transducer  always “on”How do proto-oncogenes become oncogenes?Mutation in coding or promoter region of gene• Point mutation• Deletion• Gene amplification• Translocation2. Tumor suppressor genes• Suppress uncontrolled cell proliferation• Both copies inactivated for loss of function= recessive• Retinoblastoma– Eye cancer develops in childhood– Hereditary – can be passed on– Sporadic –