4: Genome Maintenance- DNA Damage And Repair

Question 1

Endogenous DNA Damage

Answer 1

DNA damage during normal cellular function

1. Acid and heat (remove purines/pyrimidines)
2. Oxygen radicals

Question 2

Exogenous DNA Damage

Answer 2

1. Sunlight and UV (pyrimidine dimers and 6-4 photo product)
2. Chemical damage (DNA base adducts)
3. X-rays (single and double stranded breaks)

Question 3

DNA Photodamage from UV

Answer 3

Either covalent dimer (cyclobutane)

Or

(6-4) photo product that stops polymerase

Both make polymerase stop

Question 4

Chemical Mutagens and Carcinogens

Answer 4

Polycyclic aromatic hydrocarbons (PAH) from burning of fossil fuel and tobacco- converted to epoxides by P450 pathway.
Form covalent adducts with DNA bases
(Learned from chimney sweeps)

Question 5

Heterocyclic Amines

Answer 5

Overlooking meats

Covalent adducts with DNA bases

Question 6

Aflatoxins

Answer 6

From molds on peanuts and grains

Covalent adducts with bases

Question 7

Single Nucleotide Damage Repair (examples)

Answer 7

Mismatch repair

Photo-reversal of UV lesions

Base Excision Repair

Nucleotide Excision Repair

Interstrand Crosslink Repair

Question 8

Mismatch Repair (basic)

Answer 8

Repairs normal nucleotides that cannot base pair

Question 9

Photo-reversal of UV Lesions (basic)

Answer 9

Repairs photodimers

Question 10

Base Excision Repair (basic)

Answer 10

Resects and replaces small lesions (modified bases)

Question 11

Nucleotide Excision Repair and Interstand Crosslink Repair (basic)

Answer 11

Repairs big helix distorting lesions

Question 12

Double Strand Break Repair and Interstrand Crosslink (examples)

Answer 12

Homologous recombination
(Uses homologous sequence as template)

Non-homologous end joining
(Resects ends to uncover 2-3 nucleotide micro-homology)

Interstrand Crosslink Repair

Question 13

Trans-Lesion Synthesis (TLS)

Answer 13

Occurs when polymerase stalls at thymidine dimer or adduct.

Switch to different polymerase to guess what base to insert

Then polymerase resumes synthesis (delta)

Cancer cells love this to escape treatment

Question 14

Direct Repair of UV Products by Photolyase Enzyme

Answer 14

Enzyme activated by UV, no more T-T dimer

Question 15

Base Excision Repair (detail)

-from metabolic damage

Answer 15

3 steps:

1. Base removed by glycosylase
2. Phosphodiester backbone broken and deoxy sugar and phosphate removed
3. DNA polymerase and DNA ligase repair gap
   Find lesion, make cut, chew up, come back and insert right ones (small lesions)

Question 16

Nucleotide Excision Repair

Answer 16

Big bulky lesions from all the slides (adjuncts)

Enzyme complex chews large segment and polymerases add good stuff.

Question 17

BER vs. NER

Answer 17

BER: base damage, non-bulky lesions (deamination, oxidation), damage-specific (small amount of repair)

NER: bulky, helix distorting lesions (dimers and adducts), broad specificity (any helix distortion), coupled to transcription (follows RNA polymerase II), more work needed.

Question 18

Double Stranded DNA Break Repair (two pathways)

Answer 18

Homologous Directed Repair
OR
End Rejoining

*From X-Rays

(Most serious DNA damage lesions)

Question 19

D.S. DNA Break Repair (Homology Directed Repair)

Answer 19

Use homologous chromosome to recombine with broken one.

ATM and BRCA1/2- d.s. DNA repair

Question 20

D.S. DNA Break Repair (End Rejoining)

Answer 20

Enzymes put them back together, losing bases.

DNA-protein Kinase and KU (enzymes)

Direct ligation= made pair deletions)

Question 21

Interstand Crosslink Repair (detail)

Answer 21

Damage makes cross links.

Detected by FA protein complex.
Fanconi Anemia (FA)

Repaired by NER and Homology Directed Repair (HDR)

Question 22

Poly(ADP) ribose Polymerase (PARP)

“How to detect single and double stranded break”

Answer 22

Senses single-strand DNA breaks and recruits BER.

Nuclear protein detects and binds to make PAR from NAD+.

“PAR” gets linked onto histones, signal for DNA repair pathway to come deal with break.

Question 23

Xeroderma Pigmentosum Syndrome (XP)

Answer 23

Defect in NER (repair)
Increased risk of skin cancers (basal and squamous carcinomas)
“Children of the night”
8 genes can be mutated
7 of 8 code for NER complex
1 encodes bypass DNA polymerase that can recognize T-T dimers on template and add A-A.

Question 24

Hereditary Breast/Ovarian Cancer

Answer 24

Mutation in BRCA1 and 2 (homologous recombination repair- HR)

Also may link to prostate cancers

“Look like autosomal dominant but are recessive and need a second hit”

Question 25

Ataxia Telangiectasia

Answer 25

Mutations in ATM gene needed for homologous recombination repair (HR), ATM keeps complex together
Ataxia (infancy, incoordination, nystagmus)
Telangiectasia (childhood, bulbar conjunctivae)
Immunodeficiency (B, T cell formation, frequency infections)
Sensitive to X-rays

Question 26

Werner’s Syndrome

Answer 26

Short stature, premature aging (diabetes, osteoporosis, atherosclerotic diseases), cancer

Mutations in WRN gene
(Gene unwinds DNA for homologous recombination and replication)

Question 27

Bloom’s Syndrome

Answer 27

Short stature, narrow face with prominent nose, skin sensitivity to sun, butterfly-shaped facial rash (like Lupus Erthematosis), immunodeficiency, cancer (leukemia), may MR.

Mutation in BLM gene (codes for DNA helicase like WRN, used in homologous recombination)

Question 28

Fanconi Anemia

Answer 28

Short stature, leukemia, digit abnormalities, cafe au lait spots (brown)

Mutations in FANC genes (regulate Interstand DNA Crosslink repair)

FA complex

Question 29

Hereditary Nonpolyposis Colorectal Cancer (HNPCC)

Answer 29

Multiple colorectal carncinomas

Mutations in MSH2,3,6, MLH1, PMS1,2 (mismatch repair pathway)