DNA repair disorders

Question 1

Xeroderma pigmentosum

Answer 1

Rare autosomal recessive disorder
Clinically heterogenous
Dry scaly skin (xeroderma)
Abnormal pigmentation (pigmentosum)
Predisposition to skin cancer
30% show neurological abnormalities

Question 2

NER pathway

Answer 2

DNA damage recognition (XPE/C)
DNA unwinding. Open pre-incision complex (XPB/D)
Double DNA incision
Oligonucleotide excision and repair synthesis
DNA ligation

Question 3

XP cellular features

Answer 3

Reduced DNA repair synthesis (unscheduled DNA synthesis)
Increased sensitivity to damaging agents
Increased mutability

Question 4

Complementation groups

Answer 4

8 different groups (one for each defective protein)
Groups XP-A,B,C,D,E,F,G are NER deficient
XP-V (variant) are NER proficient (another defect gives same phenotype)
ABDFG have severe sunburn from early age. Early diagnosis. Neurological abnormalities
CE have normal sunburn. No neurological abnormalities

Question 5

XPC

Answer 5

XPC copurifies with hHR23B protein.
Binds diverse, helix distorting, damage containing substrates
Binds 6-4 photoproducts (helix distorting) more strongly than cyclobutane pyrimidine dimers (mild destabilisation)
Detects and binds free bases opposite damage

Question 6

XPE

Answer 6

XP-E cells mildly UV sensitive, near normal NER
XPE (DDB2) complexed with DDB1 in UV-DDB (DNA damage binding protein)
DDB binds to damaged DNA and flips out damaged bases
Creates ssDNA to facilitate XPC binding
Expression induced by p53 in response to UV

Question 7

DNA unwinding

Answer 7

XPB (3’ to 5’) and XPD (5’ to 3’) are ATP dependent DNA helicases
Subunits of TFIIH (recruited to damage via XPC)
RPA binds undamaged strand
XP-B patients rare
XP-D patients more common (XPD is dispensable)

Question 8

Damage verification

Answer 8

XP-A cells are the most UV sensitive group (no NER capacity)
XPA verifies damage and positions repair machinery correctly
XPA interacts with RPA, TFIIH and XPF-ERCC1
Stabilises open pre-incision complex

Question 9

XPF-ERCC1 and XPG

Answer 9

XPF-ERCC1 introduces nick at ss/dsDNA junction, 5’ of ssDNA
XPF interacts with TFIIH and ERCC1 interacts with XPA

XPG nicks at ss/dsDNA, 3’ of ssDNA
XPG interacts with TFIIH and XPA

Question 10

XP and skin cancer

Answer 10

Damage remains in DNA and blocks normal replicative polymerases
DNA polymerases stall/switch
Transiently replaced by translesion synthesis polymerases (TLS)
TLS have relaxed fidelity (allow DNA damage to occupy active site) and no proof reading activity