RANZCR-Like RCB Flashcards
The primary cellular target of ionising radiation is DNA.
i. List the four (4) main types of DNA lesions caused by therapeutic ionising radiation.
(2018, Q6, a i)
1 marks
1) ds-DNA break
2) ss - DNA break
3) Base modifications
4) Interstrand crosslinks
Define the terms:
ii. i Sublethal damage (SLD)
ii. ii Potentially lethal damage (PLD)
(2018, Q6, a ii)
1 marks
SLD = IR induced cellular injury that can be repaired.
PLD = IR induced cellular injury that can be potentially be repaired if conditions are appropriate - e.g cell in s-phase when damage occurs
Describe the mechanism of action of the following protein complex units at sites of double-strand DNA breaks:
- ATM-MRN
- DNA-PKcs-KU
(2018, Q6, b)
4 marks
MRN senses DSB. Activates/forms complex with ATM
ATM transduces signal:
- Phosphorylates gammaH2AX
- Phosphorylates CHK2
gammaH2AX makes DNA accessible and recruits proteins for HR repair.
Activated MRN-ATM also activates CHK2 which activates p53 and cell cycle arrest/promotes apoptosis
Ku 70 and 80 detect DSB, binding to site
DNA-PKcs-KU complex forms (transducer):
- Recruits and forms complex with Artemis protein
Artemis endonuclease activity begins NHEJ repair
Describe the processes of non-homologous end joining (NHEJ) and homologous recombination (HR).
(2018, Q6, ci)
7 marks
NHEJ
1) Detection: DSBs detected by Ku70 and Ku80 proteins which bind to site (also protect ends)
2) Recruit: DNA-PKcs recruited - autophosphorylates and phosphorylates other proteins.
3) Artemis (protein complex) recruited to DNA break, forms complex with DNA-PKcs and is activated by phosphorylation.
4) Processing: Artemis endonuclease activity processes the DNA ends ready for ligation.
If non-blunt end of DNA i.e. a 3’ or 5’ overhang, then the absent DNA can be generated accurately by polymerases.
5) DNA ends ligated
Homologous Recombination uses sister DNA with the same sequence as a template for repair. After activation as above:
1) Single stranded filaments made around DSB
2) Filaments coated with RPA
3) RAD51 displaces RPA and SEARCHES and INVADES sister chromatid
4) HELICASES UNWIND DNA and hold open SEPARATION FORK and polymerases SYNTHESISE 5’-3’ along separation fork.
5) Finally crossover points are CUT by RESOLVASES, and ligated
Detection→Recruitment/activation→Processing→Ligation
Compare the characteristics of these two DNA repair mechanisms (HR and NHEJ).
(2018, Q6, c ii)
2 marks
HR = Slow, accurate, cell phase dependent (s, early G2) NHEJ = Fast, can be inaccurate if blunt ends joined, any cell cycle phase.
Using a diagram, illustrate the phases of the cell cycle.
2019, Q12 a
2.5 Marks
G1 (G0) - S - G2 -M (M+Cytokenesis)
Briefly describe the cell activity that occurs in each cell cycle phase.
(2019, Q12 b)
2.5 Marks
G1: Growth of proteins required to be functional cell/perform normal metabolic roles. confirms conditions appropriate for replication.
S: Synthesis - DNA unwound, replication produces 46 chromosome pairs. DNA error checking and repair. prior to G2
G2: Growth - production of proteins necessary for mitosis
Mitosis: spatial separation of daughter DNA (along with intracellular organelles proteins) to allow cytokinesis.
Identify three (3) key checkpoints in the cell cycle and name the associated protein complexes that regulate cell cycle progression at these checkpoints
(2019, Q12 c)
3 Marks
1) G1/S Checkpoint. CDK 4 and 6 form complexes with CyclinD, CDK2 complex with cyclin E, (phosphorylation Rb and release E2F) , p53, p21, MRN-ATM, CHK1
2) S-interphase checkpoint. CDK 2 and cyclin A. ATR-ATRIP, CHK2, CDC25A/C
3) G2/M (early and late) Checkpoint. CDK 1 complex with Cyclin B, MRN-ATM CHK2, ATR-ATRIP-CHK2, CDC25A/C
Several key proteins such as Rb protein, p53 and p21 prevent progression through the cell cycle at a checkpoint. Discuss in detail the mechanism by which these proteins act to regulate the cell cycle at this checkpoint.
(2019, Q12 d)
5 Marks
Rb: G1 protein bound to E2F. Phosphorylated by CyclinD-CDK,4,6 activating E2F increasing cyclin E-CDK2 and promoting progression to S-phase
p53: Arrest cell cycle, promote apoptosis (G1/S, G2/M), DNA damage detected (dsDNA break activates MRM-ATM- CHK2, ssDNA break ATR-ATRIP-CHK1) CHK1/2 activates p53. p53 UPREGULATES p21 which inhibits cyclinD-CDK4 (reducing E2F)
Describe the relative radiosensitivity of cells in the different phases of the cell cycle and on the same set of labelled axes, draw cell survival curves for each of the phases of the cell cycle to illustrate this.
(2019, Q12 e)
5 Marks
In order most to least.
M>G2late>G2early>G1>S
M - Damage cannot be retired: Condensesd chromosomes make damage difficult to access, limited time for detection and repair, HR repair does not occur.
G2 late - Increasingly condensed chromosomes difficult to access/detect damage
G2 Early - Less condensed, more accessible sister DNA for HR repair
G1 - Multiple checkpoints for arrest and repair
S - DNA most accessible for HR repair and checkpoint response.
Graph: y-axis = SF, x-axis dose (Gy) 1 Gy step
Draw M and G2 close and early responding, S pretty curvy
Define the FIVE ‘Rs’ of radiobiology and using examples, comment on how each ‘R’ can influence cell survival or treatment delivery during a course of radiation treatment.
(2018, Q3, a)
5 Marks
1) Repair: Repair of IR damage to normal tissues and tumour. E.g given enough time between fractions potentially lethal damage may be repaired.
2) Repopulation: Process by which clones or normal tissue replace cells killed by radiation. E.g Hyper fractionation may allow depopulation with radio resistant clonogens.
3) Redistribution: cells in more radio resistant phases of the cell cycle during 1 fraction, progress (redistribute) to more sensitive phases by time of next fraction. E.g fractionation can exploit this to decrease cell survival
4) Re-oxygenation: Hypoxic cells are less sensitive to low LET radiation, and have improved cell survival. E.g Fractionation increases the chances of treating transiently hypoxic cells in an more normoxic phase over the course of XRT. As normoxic cells die in earlier stages of treatment,
5) Radio-resistance. Some tumours/normal tissue is more E.g allow a higher dose to be given
Define the α/β value and outline ONE limitation of the linear quadratic model.
(2018, Q3, b)
1 Marks
ffs
