GET ANALLY MILLARED

Question 1

(1) WAYS TO INACTIVATRD CYCLIN/CDKS COMPLEXES

Answer 1

1) Association of CDK inhibitor (e.g. p16INK4) that blocks interaction of CDK4 with Cyclin D
2) Association of CDK inhibitor (e.g. p27) which blocks ATP binding by Cdk2
3) Association of CDK inhibitor (e.g. p21) which blcoks substrate binding by CDK1 and CDK2
4) Tyrosine phos of Tyr15 in catalytic site of CDK1 by Wee1 kinase
5) Proteolytic destruction of cyclin following its Ubi by either the SCF or APC/C E3 Ubi ligases

Question 2

(1) Ways TO TURN ME ON CYCLIN/CDKS COMPLEXES

Answer 2

1) Transcription and translation of cyclins
2) Phosphorylation of T loop of CDK by Cdk-activating kinase (CAK)
3) Proteolytic destruction of CDK inhibitor proteins by SCF ubi ligase
4) Dephos of Tyr15 in catalytic site of CDK1 by CDC25 phosphatase
5) Intracellular re-localisation of active Cyclin/CDK complex

Question 3

(2) Identification of oncogenes process

Answer 3

1) Transfection of mouse fibroblast cells with DNA from human bladder cancer cells
2) Causes rare cells that received that oncogene to divide abnormally
3) Rare cells removed, DNA extracted and analysed
4) Human genes have nearby repetitive seq called Au seq. DNA from 2nd transfected mouse cell cloned into bacterophage
5) Hybridising phase are isolated and human DNA sequenced
6) Confirmation that oncogene has been cloned is by re-intro of clone by itself into mouse cells.

Question 4

(2) Examples of oncogenes that drive G0 cells into G1/S transition

Answer 4

RAS: GTPase in mitogenic signalling = mutation that blocks GTPase
RAF: Protein kinase in mitogenic signalling = mutations that activate kinase

Question 5

(2) SIGNAL TRANSDUCTION PATHWAY IN OCONONEOGESES

Answer 5

1) Activation of mitogen R (e.g. EGF) activates of Ras GTPase
2) Ras activates Raf to MAP kinase
3) MAP kinase enters nucleus and phos + activate a dimeric TF of Fos/Jun
4) Acti Fos/Jun TFs leads to early gene expression
5) Myc TFs triggers delayed response to gene expression. Myc controls Cyclin D, G1 phase cyclin.
6) Mitogen stimulation triggers activation G1 cyclin/CDK complex and entry into cell cycle
7) Overexpression mutation of these proto-oncogenes causes premature entry into cell cycle.

Question 6

(2) Oncogene-induced replication stress causes genome instability

Answer 6

Overexpression of oncogenes forces cells into S phase before ready
-Chromosomal rearrangements

Question 7

(3) Outline base excision repair (BER)

Answer 7

Fixes single bases of DNA

• BER initiated by DNA glycosylases that recognise and remove damaged bases
• Results in single-strand break that can be processes by a short-patches/long-patch

Question 8

(3) Nucleotide excision repair

Answer 8

Removes DNA damage induced by UV-light
-Removal of short ssDNA that contains lesion
-Undamaged ssDNA remains and used as template for DNA polymerase to make short complimentary sequence
-Final ligation to form dsDNA by DNA ligase
TWO TYPES: Global genomic and Transcription coupled.

Question 9

(3) WAT IS Xeroderma pigmentosum

Answer 9

Rare autosomal cancer occurs when both copies of NER genes are inactive. Symptoms include extreme sensitivity to sunlight and predisposition to skin tumours.

Question 10

(3) Mismatched repair (MMR)

Answer 10

Finds mistakes generated during DNA replication which A to C or G to T occurs.

• Endonuclease cuts phosphodiester bonds of DNA sugar phos backbome at point distal to error
• DNA repair pol fills hole with new correct bases
• Ligase reattaches bonds of backbone

Question 11

(3) Double strand break repair by non-homologous end-joining (NHEJ)

Answer 11

Repairs ds breaks in DNA

• Breaks ends are directly ligated without need for homologous template
• Proteins find DNA ends (short streches of ssDNA)
• When overhangs compatible, NHEJ repairs break accurately

Question 12

(3) Double strand break repair by homologous recombination

Answer 12

Genetic recombination which nucleotide seq are exchanged between 2 similar or identical molecules of DNA. Repairs double strand breaks.

1) After ds break occurs, sections of DNA around 5’ ends of break are cut away in process of resection to expose ssDNA
2) Following strand invasion, follows overhanging 3’ end of broken DNA molecule then invades similar or identical DNA molecule that is not broken
3) After strand invasion, Holliday junctions connect 2 DNA molecules
4) Resolution of junctions results in fixing double-strand breaks.

Question 13

(3) Role p53 suppressor protein

Answer 13

A TF that controls expression of genes and can arrest the cell to KILL IT aiding repair of DNA.

• ATM/ATR activated by DNA damage
• These activate Chk1 and chk2
• phos of p53 and binding of p21 gene
• p21 (CDK inhibitor protein) inhibits G1/S-CDK and S-CDK

Question 14

(3) WAT DO Mdm2 ubi ligase

Answer 14

Ensures undamaged cells p53 is ubi and destoryed so there is little p53 protein.
-If DNA is detected, phos p53 occurs binding of Mdm2 to p53 which allows p53 to accumulate.

Question 15

(3) HOW DO ONCOGENE ACTIVATION ACTIVATES P53

Answer 15

1) Hyperactivation of Ras, Myc and E2F function induction of p16INK4 and p19ARF
2) p19ARF causes stabilisation of p53 which can stop cell prolif or apoptosis.

Question 16

(4) Outline dat M phase

Answer 16

1) Nuclear envelope breaks down, MTs emanating from 2 spindle poles are able to search and bind kinetochores
2) Sister kinetochores are bound to MTs from opposite spindle poles, BI-orientation and move to metaphase plate for anaphase A (sis chromatid separation)

Question 17

(4) WAT is Cohesin

Answer 17

After replication, 2 sister chromatids are held together by multi-ring complex called cohesin

• Must be opened to for anaphase to occur.
• HAS Smc1, Smc3, Scc1 (kleisin), Scc3.

Question 18

(4) Movement into anaphase CUT YOURSELF COHESIN

Answer 18

1) Cleavage of Scc1 (kleisin) subunit of cohesin complex is catalysed by separase
2) Separase inhibited by securin during interphase and early mitosis
3) When all sister chromatids are bi-orientated on mitotic spindle, E3 ubiquitin (anaphase promoting complex APC/C) activated by Cdc20 (activator)
4) Releases separase to allow it cleave the kleisin of cohesin so sisiter chromtids align.

Question 19

(4) Mediation of degradation of cyclin B (APC/C)

Answer 19

M to A transition, APC/C polyubi cyclin B which targets for proteasome.
-Cyclin B destruction inactivates CDK1 kinase

Question 20

(4) When MTs are not fully attached to kinetochores

Answer 20

Mad2, BubR1 and Bub3 associate with Cdc20 and prevents activation of APC/C
-Also stops destruction of cyclin B and securin.

Question 21

(1) How does Rb work

Answer 21

1) Cyclin D - CDK4/6 complexes phosphorylate Rb
2) Releases TF, E2F that causes expression of Cyclin E (G1 phase cyclin) and cyclin A (S phase cyclin) forming cyclin E-CDK2 and Cyclin A-CDK2.
3) Cy E/CDK 2 further phos RB and further activate E2F
4) Cy A/CDK2 is inactive due to p27 CDK inhibitor