wk11-DNA damage II

Example of alkylated base

-O6-methylguanine -causes mispairing: •makes H-bond with thymine instead of cytosine -MGMT= the repair enzyme for O6-methylguanine •if methyl grps are not removed→DNA replication of the mispair→transition mutations -v. common in cancer •inhibiting/mutating MGMT that repairs O6-methylguanine mutation→cancer

Describe the different locations that TRNEs can occur at in relation to disorders

-SBMA, HD, SCA1: •expansion is within the coding sequence of the gene •expansion=CAG repeat •CAG encodes a glutamine codon→repeats cause encoded proteins to contain long tracts of glutamine(polyglutamine tracts)→proteins aggregate→aggregation of proteins/protein fragments carrying glutamine repeats is correlated with disease progression (e.g. brain malformation in HD) -Fragile X syndromes (FRAXA and FRAXE): •expansions located in noncoding regions of genes •repeat produces CpG islands(methylation hotspots) that become methylated→can silence gene transcription and cause loss of proteins -DM: •hypothesized that expansions cause abnormal changes in RNA structure→disease symptoms

2 reasons why the public concerned about mutagens

-a lot of agents can act as mutagens that permanently alter the structure of DNA 1-mutagenic agents are often involved in the development of human cancers 2-new mutations may be deleterious→people might want to avoid mutagens→prevent gene mutations that may have harmful effects on their future offspring

Trinucleotide Repeat expansion (TNRE)

-causes several human genetic diseases =the phenomenon in which a repeated sequence of 3 nucleotides can increase in # from one gen. to the next >displacement or addition of more than one nucleotide (triplet)

UV-light photodimerism

-causes structural defects→DNA helix is distorted •cause problems for DNA based processes -Transcription may be blocked -cyclobutane pyrimidine dimers produce a king in DNA -cyclobutane dimers in red and normal DNA in green •45C fro normal DNA

Base analogues as chemotherapy for cancer

-compounds like 5BU as therapeutic agents to introduce mutations: 1-compounds are incorporated only into DNA of actively replicating cells(e.g. cancer cells) which are more prone to take it up 2-when incorporated compounds cause many mutations in the cells(increase mutational burden)→apoptosis of cancer cells -side effect: other actively dividing cells(e.g. skin and epithelial cells of digestive tract) also incorporate 5BU→unwanted side effects e.g loss of hair and appetite

2 main types of physical mutagens

-depends on length of wavelength and energy 1-ionizing radiation: short wavelengths and high energy→penetrate deeply 2-non-ionizing radiation: long wavelengths and low energy→don't penetrate deeply

Single base changes

-e.g. deamination, alkylation, base analogues -affect the sequence of DNA but not grossly distort its overall structure →does not affect overall B-form of DNA -does not affect the actual processes of transcription/replication→exert damaging affect on future gens. through consequence of change in DNA sequence -conversion of one base into another that is not properly paired with the partner base→mismatch persists only until nxt replication→locked in and alters the DNA sequence forever

Intercalating agents

-exert their effects by directly interfering with the DNA replication process -contain flat structures that intercalate/insert themselves between adjacent base pairs in the DNA→distorting DNA helical structure -distortion of helical structure/backbone→abnormalities in transcription and DNA replication •if DNA containing these mutagens is replicated→single nucleotide additions/deletions can occur in newly made daughter strands→frameshift mutation Examples: •acridine dyes •proflavin •ethidium bromide •cisplatin=chemotherpay drug

Mechanism of trinucleotide repeat expansion

-formation of a hairpin during DNA replication leads to an increase in the length of a DNA region if it occurs in a newly made daughter strand: 1-DNA polymerase slips off the DNA after repeat sequence is synthesized 2-hairpin quickly forms 3-DNA polymerase hops back onto the DNA and continues with DNA replication 4-DNA pol synthesizes most of hairpin region twice 5-Depending on how DNA is repaired(usually non homologous end joining)→may result in trinucleotide expansion

Alternative hypothesis: Mechanism of trinucleotide repeat deletion

-formation of hairpin b4 DNA replication in the template strand→a deletion in the trinucleotide repeat (also common) 1-DNA replication occurs and DNA polymerase slips over the hairpin→DNA in hairpin is not replicated 2-if trinucleotide repeat sequence is abnormally long→changes frequently occur during gamete formation→further gens. may have trinucleotide repeat sequences that are longer/shorter than their parents 3-certain DNA repairs that involve short regions where DNA is replication→might occur in the somatic cells of individuals who already have abnormally long trinucleotide repeats in a particular gene→cause those repeats to become even longer and more harmful→severity of some TRNE disorders get worse with age

Mutations due to trinucleotide repeats

-humans and other species have certain genes and chromosomal locations that contain regions where trinucleotide sequences are repeated in tandem • in normal ppl: sequences transmitted normally from parent to offspring without mutation •in persons with TNRE disorders-length of trinucleotide repeat ↑ above a certain critical size →disease symptoms →in some diseases becomes prone to expansion →e.g. trinucleotide repeat CAG

Non ionizing radiation

-including UV light -contains less energy and longer wavelength -penetrates only the surface of an organism such as the skin -UV light causes the formation of cross-linked thymine dimers→may cause mutations when DNA strand is replicated→manifests as melanoma -higher incidence of skin cancer among ppl who have been exposed to lrg amounts of sun during their lifetime→ppl apply sunscreen to prevent harmful effects of UV light •sunscreens contain organic combounds e.g. oxybenzone that absorb UV light or reflect it

Photodimerism

-involves dimer formation between adjacent pyrimidine rings on same strand and loss of hydrogen bonding -2 products: 1-cytclobutane pyrimidine dimer (CPD) •75% of UV induced damage •covalently bond with each other instead of complementary H-bond pair with adjacent strand •serious problem to DNA replication and DNA polymerase 2-(6-4) photoproducts -different kind of covalently bonded

Mechanism of how TNRE occurs

-key aspect of TRNE=triplet repeat can form a hairpin(stem-loop) -consistent feature of triplet sequences associated with TRNE=contain at least one C and one G →can form CG base pairs→promotes hairpin formation -formation of hairpin during replication can lead to a lot of problems: •increase or decrease in length of DNA→disease phenotype >polymerase can slip off >hairpin pulls strand back >DNA polymerase will hop back on and begin synthesis from a new location→loss of sequence •if changes occur during gametogenesis→offspring will have v. diff. #s of repeats→can also ↑ repeats in somatic cells→increase severity of disease with increasing age

How does a tautomeric shift cause a mutation?

-must occur immediately b4 DNA replication -in the double stranded B DNA the bp holds the bases in their stable forms→a tautomeric shift can occur when the strands unwind -Example: 1-T base undergoes a tautomeric shift just b4 replication of the complementary strand 2-during replication: the daughter strand incorporates a G opposite the T→base mismatch 3-mismatch could be repaired by the proofreading function of DNA polymerase/ mismatch repair system 4-If the repair systems fail→nxt round of DNA rep→CG bp (not correct TA bp) 5-1/4 daughter cells inherits the CG mutation

Where does alkylation take place?

-on bases and phosphate backbone: •nitrogen and free oxygen atoms external to the base ring systems •nitrogen atoms in the base ring systems except those linked to deoxyribose •non-bridging oxygen atoms in phosphate groups -number is likelihood of position to be alkylated→key regions where alkylation takes place -one of most common alkylation on guanine

Adduct formation

-polycyclic aromatic hydrocarbons: 1-cisplatin • a polycyclic HC •v. common chemotherapeutic agent •introduces adducts in the cell and cause apoptosis in the cell 2-benzo[a]pyrene •carcinogenic constituent of cigarette smoke •adducts introduced to guanine in the cells found in the epithelial lining of the lungs→interferes with high fidelity DNA replication→more mutations→cancer

structural distortions

-provides a physical impediment to the processes of transcription/replication 3 ways: 1-adduct formation(cisplatin) 2-non-ionizing radiation e.g. photodimerism by UV light 3-intercalation

ionizing radiation

-short wavelength and high energy -can alter DNA structure -includes X-rays and gamma rays -can penetrate deeply into biological materials→produces chemically reactive molecules(free radicals)→free radicals alter the structure of DNA in many ways: 1-base deletions 2-oxidized bases 3-single nicks in DNA strands 4-cross-linking 5-chromosomal breaks

Tautometric Shifts

-temporary change in base structure(isomerization) -tautomers=bases which can exist in keto and enol or amino and imino forms •forms can interconvert by a chemical rxn→involves the migration of a H atom+switch of a single bond and an adjacent double bond -common stable form of guanine and thymine=keto form •at a low rate: G and T interconvert→enol form -common stable form of adenine and cytosine=amino form •at a low rate: A and C interconvert→imino form -enol and imino forms of these bases are found in small amounts •can still cause mutation→do not conform to the AT/GC rule of base pairing •if one of bases is in the one of the rare forms→H-bonding will promote TG and CA bps

Types of mutagens that cause induced mutations

1-Chemical mutagens 2-physical mutagens

human diseases that involve TRNEs

1-Spinal and bulbar muscular atrophy(SBMA) 2-Huntington's disease(HD) 3-Spinocerebellar ataxia(SCA1) 4-fragile X syndromes (FRAXA and FRAXE) 5-myotonic muscular dystrophy (DM)

3 types of UV

1-UV-A: 320-400nm •majority of UV light reaching earth •little DNA damage 2-UV-B: 295-320nm •~10% of UV light reaching earth •responsible for most DNA damage in skin 3-UV-C:100-295nm •wavelength maximum of DNA-highly damaging •little reaches earth's surface due to ozone layer

2 unusual features of some TRNE disorders

1-anticipation/dynamic mutation : a phenomenon in which the severity of the disease progressively worsens in future gens. 2-Anticipation doesn't occur in all TRNE disorders and usually depends on whether the disease is inherited maternally or paternally: •HD: anticipation more likely to occur if mutant gene is paternally inherited •DM: anticipation more likely to occur if mutant gene is maternally inherited →results suggest that TRNE can happen more frequently during oogenesis/spermatogenesis depending on the particular gene involved =germ cell specific events in TRNE disorders

3 types of chemical mutagens

1-base modifiers 2-intercalating agents 3-base analogues

Base modifiers-Deamination

=base modifiers that act by replacing an amino grp with another chemical grp -Example: nitrous acid(HNO2) replaces amino grps with keto grps(=NH2 → =O) = deamination •can change C→U and A→hypoxanthine •modified bases don't pair with appropriate nucleotides in the daughter strand during DNA replication >U pairs with A and hypoxanthine pairs with C

Base modifiers-Alkylation

=base modifiers that disrupt the appropriate pairing between nucleotides by alkylating bases within the DNA -methyl,ethyl grps, or larger alkyl grps (red) are transferred and covalently attached to the bases on DNA -in some cases it distorts the double helix -Examples of alkylating agents: •nitrogen mustard(type of mustard gas)>used as a chemical weapon in WWI→damages skin,eyes, mucous membranes, lungs and blood-forming organs •ethyl methanesulfonate(EMS)

Base analogues

=chemical bases not found in nature(man-made) that can become incorporated into daughter strands during DNA replication -Example: •5-bromouracil= a thymine analogue→can be incorporated into DNA instead of thymine •5BU bps with adenine •but 5BU bps has a high rate of tautomeric shift and bps with guanine→if this occurs during DNA replication →mutation in which a T-A bp→5BU-G bp(transition) •nxt round of DNA replication: template strand containing guanine produces a GC bp→5BU promotes change of AT bp to GC bp→introduces mutations to the DNA

base modifiers

mutagens that act by covalently modifying the structure of bases→base changes after replication