Unit 9

Unit 9: DNA Damage, Repair, and Mutation Worksheet

Define

- Mutations- Physical or chemical abnormality in the structure of DNA which can be good or bad, leads to new alleles which are raw material for recombination

- Recombination- Outcome of cellular processes that cause alleles of different genes to be grouped in new combinations

- Somatic mutations- In single cells during an organisms life and are passed on to daughter cells but NOT offspring.

- Germ-line mutations- Arise in gametes that are inherited in offspring and are present in all of their cells.

- Point mutations- Single-base-pair change in DNA sequence

- Base substitutions- Transition and Transversions; where one base pair is replaced by another

        - Transition- Type of base substitution where a purine is replaced with a purine with         purine or pyrimidine with pyrimidine.

        - Transversion- Second type of base substitution which a pyrimidine with purine or purine         with pyrimidine.

- Base insertion- The addition of one base pair

- Base deletion- The removal of one base pair

- Synonymous mutations- Also called “silent”, changes the codon but not the function (same amino acid)

- Missense mutations- Also called “nonsynonymous” changes the codon and the function (diff amino acid)

        - Conservative mutation- codes for a different amino acid that is similar to the original (may not change the protein)

        - Non-conservative mutation- codes for a different amino acid that is different from the original (most likely changes the proteins)

- Nonsense mutation- changes the codon to a stop codon which prematurely ends translation

- Spontaneous mutations- mutations that occur “naturally” and arise in cells

- Induced mutations- arise through the action of external agents (mutagens)

- Mutagens- Could be radiation, chemicals or infectious agents

- Tautomerization- Spontaneous isomerization of a nitrogenous base from normal to alternative form

- Ionization- process by which an atom in DNA acquires a negative or positive charge

- Transition mutation- Purine substitutes for a purine or a pyrimidine substitutes for a pyrimidine can result from no proofreading

- Fixed permanent mutation- If the MMR system fails it lead to a fixed permanent mutation which means it stays forever

- Depurination- The lose of a purine base (A or G) 

- Apurinic site- Happens when depurination does its a site where it doesn’t specify for a complementary base.

- Deamination- Hydrolytic removal of an amine group, alters three DNA bases that contain an amino group (C, A, and G)

- Oxidative damage- Damage to DNA caused by oxygen species.

- Akylation- Addition of an alkyl group (methyl or ethyl group) to a nucleotide base

- Bulky adducts- Large, covalent chemical modifications to DNA that disrupt its normal structure

- Base analogs- Similar to normal nitrogenous bases

- Base Excision Repair (BER)- Main target is non-bulky damage

- Nucleotide Excision Repair (NER)- Repair bulky adducts and pyrimidine dimer

        - Global genome NER- Removal of bulky adducts and pyrimidine dimers independently of             transcription

        - Transcription-coupled NER- removal of bulky adducts and pyrimidine dimers during                    transcription

- Translesion synthesis- Another line of defense against DNA damage

- Nonhomologous end joining (NHEJ)- a pathway that repairs double-strand breaks in DNA.

- Homologous recombination (HR)- nucleotide sequences are exchanged between two similar     or identical molecules of DNA.

Concepts

- Why do organisms have to tolerate low levels of mutations?- The provide the raw material for evolution

- Why do base substitutions NOT affect the reading frame for protein synthesis?- It’s because they don’t add or delete a base

- Why do base insertions/deletions affect the reading frame of protein synthesis?- It adds or deletes a base which would shift the reading because its like a sentence that adds a word or loses one it doesn’t match up.

- Using the Ras, detail how point mutations can cause major phenotypic consequences- Ras gene could have a missense mutation which changes some chemicals that prevent hydrolysis of GTP which leaves the protein on.

- Detail how a point mutation in a noncoding region can affect the phenotype of cells and organisms- The non-coding are important to DNA replication and transcription so if the point mutations occur then it will still have an effect on the phenotype, also some molecular effects are altered transcription, altered splicing and altered mRNA stability, with that, those can effect the phenotype like a cascade effect.

- What are the 3 main ways that can cause spontaneous mutations- DNA replication mistakes, Cellular environment like water and oxygen species, and transposable elements that can move in the eukaryotic genomes

- How can DNA replication cause spontaneous mutations- when errors occur in the DNA replication process like mismatched base pairs.

- Detail incorporation errors and why they occur- Its mismatching and it occurs because of tautomerization and ionization

- Detail tautomerization and how it can induce a mutation in the genome- Its when spontaneous isomerization (same chemical diff structure) of a nitrogenous base from its normal form to an alternative, Tautomers are the different forms of nitrogenous bases. The normal form is called Keto form and the alternative forms are called Imino/Enol forms which may pair with the wrong base.

- Detail ionization and how it can induce a mutation in the genome- when an atom in DNA acquires a negative positive charge.

- Detail how the failure of MMR can cause mutations in the genome- Brough about by proton exchange between water and hydrogen bonds, nucleotides either gain or lose protons which effects the hydrogen bonding properties which can lead to a mixmatch

- Detail the process of replication slippage and how it can cause mutations in the genome- Slippage occurs during the synthesis of a sequence containing mononucleotide (AAAAAAA) or short tandem repeats (GATGATGATGATGAT), new strand can dissociate from the template strand then re-anneal in a misaligned position.

- How can the cellular environment cause spontaneous mutations- Depurination, Deamination or Oxidative damage which comes from chemical reactions that can happen naturally in the cell.

- Detail how depurination can cause spontaneous mutations- its the loss of purine bas (A or G), brought by hydrolysis of the glycosidic bond between the base and deoxyribose sugar, results in loss of the base of DNA but the phosphodiester backbone remains intact. Happens all the time (2000-10,000 a day in each human cell) If they were to persist it would have damage and it forms an Apurinic Site which dosen’t specify for a complementary base, BUT cells have repair systems that can remove the sites

- Detail how deamination can cause spontaneous mutations- This is the hydrolytic removal of an amine group. Alters the three DNA bases that contain an amino group (C, A, and G)

    Cytosine to uracil

    Adenine to hypoxanthine

    Guanine to xanthine

    5’methylcytosine to thymine

- Detail how oxidative damage can cause spontaneous mutations- Reactive oxygen species (superoxide radicals, hydrogen peroxide, hydroxyl radicals) are normal products of aerobic metabolism of molecular oxygen by mitochondria. 100 types of oxidative DNA modifications have been identified in mammalian genomes

- Detail how akylation can induce mutations- Addition of an alkyl group to nucleotide base which can prevent normal base pairing.

- Detail how bulky adducts can induce mutations- Large, covalent chemical modifications to DNA that disrupt its normal structure, often caused by genotoxic aromatic compounds (smoke and air pollutants) can break bonds.

- Detail how base analogs can induce mutations- Similar to nitrogenous bases which can sometimes replace normal bases, to be mutagenic it has to mispair more often than the normal base pair it replaces, only exist in a single strand.

- Detail how the binding of intercalating agents can induce mutations- Planar molecules that mimic base pairs, can fool the DNA polymerase into inserting extra bases or skipping bases 

- How does ultraviolet light damage bases and induce mutation- Can form various pyrimidine dimers in DNA, side by side pyrimidines can bond together which blocks DNA replication and transcription.

- Explain how ionizing radiation can induce mutations- It generates Reactive oxygen species (ROS) occurs through radiolysis of water which messes up mitochondrial functions, can also directly damage bonds in the DNA or break single or double strands

- What is the key principle that guides the repair systems in a cell?- Nucleotide complementarity 

- What type of DNA damage can be directly repaired? How do cells achieve this?- Only a few can be reversible, it directly reversed the damage base to the normal base, the main two are UV light damage and Base alkylation

- Detail the base excision repair system in cells- Main target of non-bulky base damage (alkylation, oxidation and deamination), damage is detected by a DNA glycosylase, cleabes the glycosidic bond between base and sugar, creates apurinic or apyrimidinic site, AP endonuclease nicks the damaged strand upstream (from the site) DNA polymerase give new nucleotides and DNA ligase seals the new nucleotides.

- Detail nucleotide excision repair system in cells- repairs the bulky, Glpbal genome NER or Transcription-coupled NER, RNA pol II detects mutation, after detection TFIIH is recruited and binds to mutated segment, cleaves, polyemerase, ligase occur.

- Detail how the MMR system repairs mismatched base pairs- recognizes and repairs mismatched bases caused by insertions and deletions of nucleotides MutH cuts the newly synthesized strand containing the incorrect base, DNA polymerase recruited and replaces the excised segment.

- Detail the Translesion synthesis repair system in cells- Last line of defense, initiated by stalled DNA polymerase, once the extension passes the lesion and the TLS polymerase is replaced by the replicative DNA polymerase

- How do cells repair double bond breaks- two mechanisms repair DSBs, two nonhomilogous end joining, homologous recombination

- Detail the nonhomologous end joining mechanism to repair double-strand breaks- initiated when Ku70 and Ku80 detects and binds to each broken end of the DBS, prevents further damage and recruits DNA-PK, that then phosphorylates Artemis, small gaps in DNA are filled in by the DNA polymerase

- Detail the homologous recombination mechanism to repair double-strand breaks- 

requires an undamaged homologous double-stranded DNA template, Both pathways start the same way
○ Broken DNA ends are processed by exonuclease to generate 3’
single-stranded DNA overhangs
○ Overhangs participate in strand exchange with homologous
double-stranded DNA
● The invading 3’ overhang displaces one strand of the
homologous DNA and base pairs to the other (via
recombinase)
○ Creates a structure called a displacement loop (D-loop)
● The invading strand is the extended by DNA synthesis
using the homologous strand as a template
○ After this point, pathways diverge

- Detail the double-stranded break repair system.- 

The other 3’ overhang invades,
creating a four-branched, double
cross-over (Holliday Junction)
● Endonuclease cleave the Holliday
junctions
○ Yields noncrossover or crossover DNA
segments
● Lastly, gaps are filled in by DNA
polymerase
○ DNA ligase seals the remaining nicks

- Detail how the synthesis-dependent strand annealing system repairs double-stranded breaks- 

DNA helicase displace the
extended invading strand
● Original broken chromosome
pieces are annealed together
● DNA polymerase comes in and
synthesizes missing information
○ DNA ligase seals DNA segment