DNA Damage and Repair Flashcards
What is the size of a ‘spur’ and how many ionization events?
4 nm in diameter
3 ion pairs
What is the size of a ‘blob’ and how many ionization events?
7 nm
12 ion pairs
What is the approximate width of dsDNA? length of a turn?
2 nm
3.4 nm
What predominates with exposure to low LET? high LET? (blobs vs. spurs)
low LET is sparsely ionizing, spurs predominate
high LET is densely ionizing, blobs predominate
How common is the following damage/cell with 1Gy?
1) ionization events
2) base damage
3) SSBs
4) DSBs
5) cell death
1) 100k
2) >5k
3) 1k
4) 40
5) 0.63 (SF = 0.37)
How does a dicentric chromosome form?
Two separate chromatids have a DSB, the free ends fuse –> p1/centromere/q1p2/centromere/q2
How does a ring form?
The same chromatid has DSB on both ends, the free ends circularize
How does an anaphase bridge form?
Sister chromatids on the same chromosome have DSB, the two sister free ends fuse. These are unable to segregate nicely during anaphase, and random breaks are induced (with loss of DNA in a daughter)
Approximately what total body dose is needed to see chromosome aberrations in lymphocytes?
> 0.2 Gy. Above 4 Gy, lymphocytes undergo apoptosis and chromosome damage is not seen
What is the process of base excision repair?
Involved proteins: APE1, XRCC1, PARP, PolB, Lig3
1) APE1 and PARP interact with the site to remove the base/backbone
2) XRCC1 stabilizes the induced break while PolB repairs it
3) Ligase3 seals the backbone repair
What is the process of nucleotide excision repair?
Involved proteins: XP family, RPA, ERCC1, CSA, polD, Lig1
1) XPA proteins and CSA recognize the lesion
2) XPA and RPA stabilize the ssDNA ‘bubble’
3) ERCC1 and XPG excise the segment
4) Pol-delta synthesizes the repair strand
5) Ligase I seals the backbone
When is NER utilized vs BER?
BER is for small, single base disruptions whereas NER is for larger, bulky lesions (UV, alkyl groups, platinum)
What is the process of MMR?
Involved proteins: MSH, MLH, Exo1, polD, Lig1
1) MSH and MLH recognize/stabilize the mismatch
2) Exo1 removes the lesion
3) RPA stabilizes the ssDNA while pol-delta synthesizes
4) Lig1 seals backbone
Essentially the same ‘repair’ steps as NER
What causes a defect in MMR? What does this cause?
Lynch syndrome
Microsatellite instability
What is the initiation of DSB recognition and repair?
ATM and ATR recognize the DSBs, phosphorylate gH2ax, and recruit the MRN complex (Mre11, Rad50, Nbs1)
Then, there is a competition to undergo HRR or NHEJ, facilitated by 53BP1
p53 and Chk1/Chk2 are activated as well
What is the function of ATM?
Recognition of DSB
Regulation of MRN activity (resection of ends)
What is the function of 53BP1?
53BP1 inhibits the exonuclease activity of the MRN complex, prohibiting HRR
What is the NHEJ pathway?
Key proteins: Ku70, Ku80, Artemis, PKcs, Pol-lambda
1) Ku70/Ku80 dimer binds free ends
2) DNA-PKcs is recruited to juxtapose the free ends
3) Artemis removes overhangs
4) XRCC4/Pol-L/Lig4 then ligate the free ends
When does NHEJ predominate?
during G0/G1, when there is no sister chromatid as a template
What is the HRR pathway?
Key proteins: MRN complex, RPA, Rad51, Rad52, BRCA1/2,
1) ATM recruits the MRN complex (rad50) and BRCA1 to the site of damage
2) Mre11 creates a 3’ ssDNA overhang
3) BRCA2 facilitates the recruitment of RPA to stabilize the ssDNA and rad51 to allow strand invasion of the sister chromatid
4) Rad52 then removes rad51 after invasion to allow DNA duplex formation
5) A helicase helps unwind the template and pol synthesizes through the Holliday junction
What disease is associated with NER deficiency?
Xeroderma Pigmentosum
What disease is associated with CSA/CSB deficiency? What path is this in?
Cockayne Syndrome (NER) Photosensitivity without cancer risk
What is disrupted in Lynch Syndrome?
MSH/MLH
Are BRCA1/2 patients radiosensitive?
Surprisingly no
