!!!13.2 DNA Replication and Repair

Question 1

Localization of DNA Replication

Answer 1

• Replication sites distributed throughout nucleus in distinct sites
• Replication forks localized within 50 to 250 sites, called replication foci.
• The clustering of replication forks may provide a mechanism for coordinating the replication of adjacent replicons on individual chromosomes.

Question 2

Distribution of Histone Core Complexes. How histone nucleosomes associate on newly associated DNA.

Answer 2

• beads on string 1st level of orginizatoin. So what happens after DNA is replicated? need to get nucleosome on the new strand.
• (H3H4)2 tetramers remain intact
• H2A/H2B dimers separate from each other
• mix of new and old to form beads on a string.

Question 3

Epigenetic State of Replicated Chromatin

Answer 3

• DNA that’s being replicating has a certain epigenetic pattern. The pattern is maintained in the newly synthesizes DNA’
• Old histones guide the modification of the new histones.
• If you have old and new histone in close proximity. If it already has chromodomain it’ll recite more to the other one.
• • feedback loop/ old inficending the new histones. The modification is coppied.
• Not entire sure how it happens.

Question 4

The 4 Types of DNA Repair Pathways

Answer 4

1. Nucleotide excision repair
   a) Transcription coupled pathway
   b) Global pathway
2. Base-excision repair
3. Mismatch repair
4. Double strand break repair

Question 5

Pyrimidine (Thymine) Dimer

Answer 5

Thymine dimer mutation. This type of mutation is induced by UV radiatation. Normally H-binding in opposite. If thymines are right next to each other, the UV light created covalent linkages between the thymines. Thimine dimers. So breaks up the H-bond. Term is bulky adduct. They are no longer hydrogen binding. This is a mutation that causes problems because DNA pol has to try to move through bulky adduct.

Question 6

Trasciption coupled path

Answer 6

Related to progression of RNA pol through template and encountering Thymine Dimer. chugging along transcribing and when it encounter this will pause and recruit CSB to the site of the bulky path.

Question 7

Global Path

Answer 7

deals with all DNA. (nbot just DNA that’s being trascibed) lesion signaled by stalled RNA polymerase and CSB protein. TFIIH (XPB and XPD components) unwind the local area around the thymine dimer,. XPC is for global. Then recruits XPC, then start to cleve the DNA out from each side.

Question 8

Nucleotide Excision Repair -5

Answer 8

1) damage recognition mediated by either CSB or XPC
2) Unwinding of DNA by XPB and XPD proteins and helicase subunits of TFIIH
3) Nucleases cut on 5’ (XPF-ERCC1) and 3’ (XPG) sides
4) Filling of gap by DNA pol δ/ε
5) DNA repair synthesis
6) ligation

Question 9

Xeroderma Pigmentosum

Answer 9

• Deficient for repairing damage from UV light
• Nucleotide excision repair deficiency
• Related to Cockayne syndrome
• Deficient for repair of transcriptionally active DNA

Question 10

Base Excision Repair

Answer 10

1) Uracil-DNA glycosylase recognizes uracil
2) Glycosylase cleaves base (cleaves glycosidic bond)
3) AP endonuclease cleaves DNA backbone
3) DNA backbone cleaved

4) Phosphodiesterase activity of polymerase β removes sugar phosphate from excised base
5) Polymerase β fills gap
6) strand is sealed by DNA ligase III

Question 11

Detecting Damaged DNA in Base-Excision Repair

Answer 11

1) DNA glycosylase (hOGG1) inspects base paired to cytosine (all GC pairs)
2) Base flipped out of DNA duplex (oxidized guanine)
3) Base cleaved from sugar
4) Guanine not recognized by active site (returned to DNA)

Question 12

Mismatch Repair

Answer 12

• Recognizes distortion in geometry of base-paired nucleotides
• Distinguishes between newly synthesized and parental strands
• Methylation pattern in prokaryotes
• Method not clear in eukaryotes
• polymerase recognizes the geometry and can recognize correct and incorrect geometry.

Question 13

Double-Strand Breakage Repair

Answer 13

1) detected by heterodimeric ring shaped protein Ku that binds to broken end of DNA
2) Ku recruits protein DNA-PKcs (the catalytic subunit of DNA dependant protein kinase
3) bring ends of the DNA together and phosphorylate the ends
4) joined by DNA ligase IV

non-homologous end joining, no template, just joins the end back together

DNA ligase IV is a specific repair ligase recruited to the hosphoralated ends

Question 14

Homologous recombination

Answer 14

\ breaks can be repaired by homo recombination. \ break in one chromo, after the break, the DNA is widened by exonuclease chewed back, get s.s ends. Those 3’ ends undergo strand. Will use the homo chromo as template to repair the DNA break. Does this with help from Rad 51.

Question 15

Translesion Repair

Answer 15

sometimes DNA is not repaired before replication. Translesion repair can help with this. If it encounters one of these mutations for example a thymine dimer, it can recruit a specialized enzyme called Eta. Eta doesn’t object to thymine dimers it recognizes them and sticks to adenine opposite the thymine dimer. Comes in and takes over for DNA III. Inserts two A nucleotides opposite thymine dimer

Question 16

Cancer, DNA Repair and DNA Replication

Answer 16

•Microsalatiles tend to be mutation hot spots. Errors really common in repeated sequences. TGF-B is a growth factor receptor.
• Colon cancer
15% of cases linked to mutated genes for mismatch repair

Question 17

Instability in TGF-β1 Receptor Gene

Answer 17

• Vascular disease is believed to be the result of excessive wound healing response to chronic injuries
• Hypertension, smoking
• TGF-β1 gene product controls regulatory systems that respond to cellular injury
• Antiproliferative effect
• Damage to receptor is associated with decreased antiproliferative effect

Question 18

Microsatalites

Answer 18

Only thing to take away, microstatlites, especially single are prone to replication errors. Repeating DNA are prone to replication errors. Could ask why. This is because replication slippage.