Endogenous DNA DamagePossible Consequences of DNA DamageDNA Repair PathwaysIR => ROS DamageUV DNA DamageAlkylating Agents: Reactive ElectrophilesChemically Modified DNATypes of DNA RepairDirect Repair (DR)Direct Repair by MGMTDirect Repair by AlkB like enzymesBase excision repair (BER)BER: Initiated by a mono-functional glycosylase on alkylated DNA(i.e. the AAG DNA Glycosylase)BER: Initiated by a bi-functional glycosylase on oxidized DNA(i.e. OGG DNA Glycosylases / AP Lyases)Three Distinct Ways to Address Oxidative DamageNucleotide Excision Repair (NER)Mismatch Repair (MMR)Signaling by MMR and a Mutator PhenotypeRecombinational Repair (RR)Non Homologous End Joining (NHEJ)Sensing DNA DamageDNA Damage and DNA RepairThomas [email protected]@email.unc.eduDNADouble HelixPurinesPyrimidinesNitrogen Bases of DNADNAEndogenous DNA DamageEndogenouse Source Number of lesionsHydrolysis Depurination 18,000 Depyrimidaination 600 Cytosine Deamination 100 5-methylcytosine deamination 10Oxidation 8-oxoG ~1,000 - 2,000 Ring saturated pyrimidines ~2,000 Lipid Peroxidation products ~1,000Nonenzymatic methylation by SAM 7-Methylguanine 6,000 3-Methyladenine 1,200 1-methyladenine and 3-methylcytosine NDNonenzymatic methylation by nitrosated polyamines abd peptides O6-methylguanine20-100Environmental Sources of DNA DamageReplication problemsIf DNA repair corruptedPossible Consequences of DNA DamageDNA DamageVia UV, oxidation or methylationdamageMutationCell DeathNothingCauses of Cancer and AgeingDNA Repair PathwaysComplexityNucleotide Excision RepairNucleotide Excision RepairMismatch RepairMismatch RepairBase Excision RepairBase Excision Repair+ Recombination based repairDirect RepairMgmtMgmtS-CH3SHDirect RepairMgmtMgmtS-CH3SHMost Complexhttp://www.rndsystems.com/mini_review_detail_objectname_MR03_DNADamageResponse.aspxDNA Damaging Agents and Specific Types of DNA DamageWhat is Ionizing Radiation?Ionizing radiation carries enough energy to ionize an atom or molecule (that is, to completely remove an electron from its orbit).H O H O ++ e-hv2 2H2O + H++ OHDNA Repair and Mutagenesis By Errol C. Friedberg, Graham C. Walker, Wolfram Siede 1995IR => ROS DamageDNA Repair and Mutagenesis By Errol C. Friedberg, Graham C. Walker, Wolfram Siede 1995UV DNA Damagecis-syn and trans-syn diastereoisomers of cyclobutane dimers and the pyrimidine (6-4) pyrimidone adducts Biol Chem, Vol. 275, Issue 16, 11678-11685, April 21, 2000Formation of the Main UV-induced Thymine Dimeric Lesions within Isolated and Cellular DNA as Measured by High Performance Liquid Chromatography-Tandem Mass Spectrometry* Thierry Douki, Magali Court, Sylvie Sauvaigo, Francette Odin, and Jean CadetChemical CarcinogensDirect Acting CarcinogensAlkylating agents Anticancer drugs (cyclophosphamide, chlorambucil, nitrosureas)Procarcinogens that require metabolic activationPolycyclic and heterocyclic aromatic hydrocarbonsBenz(a)anthraceneBenzo(a)pyreneAromatic amines, amides, azo dyes2-Napthylamine2-AcetylaminofluoreneNatural plant and microbial productsAflatoxin B1GriseofulvinOthersNitrosamineVinyl chlorideHeavy metals (Nickel & chromium)Insecticides, fungicidesPCBsAlkylating Agents: Reactive ElectrophilesSimple alkylating agents found in tobacco smoke are an example: most agents: Sn1: Sn2: methyl radicalsNNONH2NNNNHONH2dRdR10% SS0.1%1.0 %6.0 %70%326378CGNNHOOCH3NNNNNH2dRdR0.4 %2.0 %0.1%20 %SS10%784323TAChemically Modified DNACarcinogenesis. 2004 Jun;25(6):1045-51.Alekseyev YO, Hamm ML, Essigmann JM.Biochemistry 2002, 41, 9535-9544Natalia Tretyakova,*,‡,§ Brock Matter,‡ Roger Jones,| and Anthony ShallopTypes of DNA Repair• Direct Repair (DR)• Base excision repair (BER)• Nucleotide excision repair (NER)• Mismatch Repair (MMR)• Recombinational repair (RR)Direct Repair (DR)AlkB + Fe(II)06-meG DNA Methyltransferase06-meG DNA MethyltransferaseCH3Cys06-meG DNA MethyltransferaseCH3CysNCH3NCH3CH3OO2+ α-ketoglutarateCO2+ succinate+ HCHODirect Repair by MGMTMgmtMgmtS-CH3SHNONNONHOOCH3NH2NONNONHOONH2NONHOOOOCH3CH3NONHOOOOCH3Direct Repair by AlkB like enzymesABH1, ABH4 - ABH8?Base excision repair (BER)“Lots of flavors depending on the damage, initiating enzymes, and downstream processing”AAG SubstratesNNN+NHONH2*CH3NN+NNHONH2*CH3NN+NNNH2*CH3NNNNNH*NNNNHO*NNNHNHONH2O*NNNHNHONH2CH2CH2OH*NNNHNHONH2CH2CH2Cl*NNNNONH*7-MeG 3-MeG 3-MeAεA Hx 8-oxoGεG 7-HeG 7-CEGBER: Initiated by a mono-functional glycosylase on alkylated DNA(i.e. the AAG DNA Glycosylase)Oxidative DNA DamageOGG1 and MUTYHSubstratesRemoves ARemoves 8-oxoGNature 447, 941-950 (21 June 2007) |Sheila S. David1, Valerie L. O'Shea & Sucharita KunduBER: Initiated by a bi-functional glycosylase on oxidized DNA(i.e. OGG DNA Glycosylases / AP Lyases)Curr Biol. 1997 Sep 1;7(9):R576-9.Cunningham RP.Three Distinct Ways to Address Oxidative DamageCurr Biol. 1997 Sep 1;7(9):R576-9.Cunningham RP.BER: Other enzymes and downstream decisionsDNA Repair (Amst). 2007 Mar 3; : 17337257 A unified view of base excision repair: Lesion-dependent protein complexes regulated by post-translational modification. Karen H Almeida , Robert W SobolNucleotide Excision Repair (NER)http://www.youtube.com/watch?v=CASw429qBUULesions Removed by NERUVcyclobutane dimerN2– BPDE-dGNucleotide Excision Repairhttp://www.rndsystems.com/mini_review_detail_objectname_MR03_DNADamageResponse.aspxxeroderma pigmentosum (XP) is caused by a defectin nucleotide excision repair mechanismsXP is categorized in at least eight complementation groups according to the capacity of the body to repair DNA. These groups (i.e., genetic subtypes) are labeled A through G, plus the XP variant:.Wide range of symptomsblistering or freckling on minimum sun exposure premature aging of skin, lips, eyes, mouth and tongue; with significant increased incidence of cancer in these same areas blindness resulting from eye lesions or surgery for skin cancer close to the eyes progressive neurological complications including: developmental disabilities mental retardation The median age of onset of skin cancer is 8 years, nearly 50 years younger than in the general populationxeroderma pigmentosum (XP) is caused by a defectin nucleotide excision repair mechanismshypermutability after UV irradiation in cell culturespropensity to
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