Checkpoint controls I

Question 1

What is growth and division regulated by?

Answer 1

Progression through the cell cycle

Question 2

What does the genetic material of 2 daughter cells depend upon?

What do defects in these cause?

Answer 2

The PRECISE execution of 2 processes:
1) Faithful REPLICATION of a cell’s genome in S phase

2) The proper ALLOCATION of the resulting duplicated DNAs to a daughter cell during M phase

Defects = cancer

Question 3

What is the function of a checkpoint in the cell cycle?

Answer 3

Surveillance/monitoring mechanisms which monitor each step of cell cycle progression

Question 4

When can/can’t cells progress through the cell cycle?

Answer 4

Can progress:
- ONLY if the pre-requisite step has been completed successfully

If not - cells cannot progress until the problems have been addressed

Question 5

What happens if the problems which are causing the cell cycle to halt cannot be repaired?

Answer 5

The cell undergoes APOPTOSIS

Question 6

Describe the G1/S phase checkpoint

Answer 6

Cell will not be permitted to enter S phase if the genome is in need of repair

Question 7

Describe the intra-S phase checkpoint

Answer 7

DNA replication paused in response to DNA damage, allowing DNA to be repaired

Question 8

Why can S phase increase in time?

Answer 8

Time taken for the DNA to undergo DNA repair (S phase pauses in response to DNA damage

Question 9

Describe the G2/M checkpoint?

Answer 9

A cell will NOT proceed through this checkpoint until DNA REPLICATION has been completed

OR

There is DNA damage

Question 10

Describe the spindle assembly checkpoint?

Answer 10

The metaphase/anaphase checkpoint:

• Cell is not permitted to enter anaphase until all of the chromosomes are properly assembled on the mitotic spindle DURING METAPHASE

Question 11

What happens if the chromosomes are incorrectly assembled on the mitotic spindle?

Answer 11

Daughter cells could have 2 different genomes

Leads to the ACCUMULATION of mutations that could lead to the formation of cancer

Question 12

How do cancer cells behave differently to normal cells at checkpoints?

Answer 12

In normal cells, if there is DNA damage - the cells fail to progress through the cell cycle and don’t proliferate

In cancer cells - LOTS of DNA damage but STILL continue to PROLIFERATE

Question 13

What mutations must occur in cancer cells so that they can go through the cell cycle?

Answer 13

INACTIVATION of one or more checkpoint controls

In addition to acquiring oncogenes and inactivated TSGs

Question 14

What is the R point essential for?

Answer 14

At the end of G1 - in order for the cells to make a decision if to proliferate or become quiescent

Question 15

What is pRb?

Answer 15

A nuclear PHOSPHOprotein that is ABSENT or in a DEFECTIVE form in many tumours

Question 16

What is the molecular governor of the R point?

How?

Answer 16

Rb (Retinoblastoma protein)

• pRb undergoes PHOSPHORYLATION at the SAME time as the advance of cells through the cell cycle
• Hyperphosphorylation of pRb inactivates the protein and allows the advance through the cell cycle

Question 17

What happens when pRb is active?

Answer 17

PREVENTS the cells from going through the R point and the rest of the cell cycle

Question 18

When is pRb activate?

Answer 18

When pRb is unphosphorylated or hypophosphorylated

Question 19

When is pRb inactive?

Answer 19

When INACTIVATED

Question 20

What is pRb phosphorylation governed by?

Answer 20

The components of the cell cycle

Question 21

When is pRb NOT phosphorylated?

Answer 21

In G0

Question 22

What happens to pRb in early/mid G1?

Answer 22

D-type cyclin and CDK4/6 initiate pRb phosphorylation

Causes HYPOPHOSPHOYLATION

Question 23

What is hypophosphoylation of pRb?

Answer 23

Only a SMALL number of residues are phosphorylated

Question 24

What is hyperphosphoylation of pRb?

Answer 24

Phosphorylation of a LARGER number of residues

Question 25

What mediates the hyperphosphorylation of pRb?

Answer 25

Increase of cyclin E increase a R-tpoint Cyclin E/CDK2 mediates the hyperphosphorylation Advancement through late G1

Question 26

What happens to pRb after early/mid G1?

Answer 26

pRb remains HYPERPHOSPHORYLATED for the remainder of the cell cycle

Question 27

When is pRb dephosphoylated? What does this allow?

Answer 27

When the cells exit mitosis Cell cycle can start again

Question 28

What does the E2F transcription factors do? How?

Answer 28

Enables pRb to implement growth vs quiescent decisions: - UNphosphoylated or HYPOphosphorylated pRb bind E2F - HYPERphosphoylated pRb dissociates from E2F

Question 29

What happens with pRb and E2F in G1?

Answer 29

1) pRb is HYPOphosphorylated | 2) pRb binds to E2F - preventing the transcription of E2F dependant genes

Question 30

What happens with pRb and E2F at the R point?

Answer 30

1) pRb is HYPERphosphorylated | 2) E2F is RELEASED - transcription of the genes mediating G1/S transition

Question 31

What happens to pRb and E2F as the cells undergo G1/S transition? Why?

Answer 31

Transcriptional activity of E2F is inhibited by Cyclin A/CDK2 which is formed at the G1/S transition E2F is DEGRADED by UBIQUITINATION

Question 32

What is the length of action of the E2F transcription factor?

Answer 32

Short lived

Question 33

What are the positive feedback loops in pRb/E2F signalling?

Answer 33

1) pRb is HYPERphosphorylated by cyclin E/CDK2 --> ACTIVATION of E2F (released) - One of the target genes of E2F is cyclin E - Further INHIBITION of pRb - Further ACTIVATION of E2F 2) Cyclin E-CDK2 induces the PHOSPHORYLATION of p27^Kip1 (CKI), which is ubiquitinated and degraded - More E-CDK2 released from inhibition

Question 34

What ensures the cell cycle is irreversible?

Answer 34

Positive feedback loops

Question 35

Describe the separation of the sister chromatids during anaphase?

Answer 35

IRREVERSIBLE step: - Cycle will not proceed until ALL the chromosomes are attached to at least 2 spindle fibres from the OPPOSITE poles - If chromosome miss placed - cell will pause mitosis, allowing time for the spindle to capture the stray chromosome

Question 36

What is the progression into anaphase controlled by? How?

Answer 36

The anaphase promoting complex/cyclosome (APC/C): 1) The APC is activated when all the chromosomes are aligned 2) APC/C ubiquitinates SECURIN and causes it to undergo degradation 3) Causes the activation of separase

Question 37

What keeps the 2 sister chromatids together?

Answer 37

Cohesion

Question 38

When is separase inactivated?

Answer 38

Inactivated when in a complex with SECURIN

Question 39

When is separase activated? What happens when separase is activated?

Answer 39

When securin is INACTIVATED When separase is activated: 1) Breaks the link between the 2 chromatids by DEGRADING cohesion 2) Allowing anaphase to initiate and the sister chromatids to be pulled towards the opposite sides of the cell

Question 40

What 2 things are checked at the G2/M checkpoint?

Answer 40

1) DNA integrity (absence of damage) | 2) DNA replication (DNA completely copied during S phase)

Question 41

What is the passage through the G2 checkpoint triggered by?

Answer 41

MPF (mitosis promoting factor)

Question 42

What is a MPF?

Answer 42

A cyclin B/CDC2 complex

Question 43

What happens is the checkpoint mechanism detects a problem with DNA (damage or incomplete replication)?

Answer 43

- Cell cycle HALTED | - Cell attempts to either COMPLETE DNA replication or REPAIR damaged DNA

Question 44

What happens if the cell CAN repair the problem with DNA?

Answer 44

Cell can progress through mitosis

Question 45

What happens if the cell CAN'T repair the problem with DNA?

Answer 45

Cell may undergo APOPTOSIS

Question 46

What is DNA detected by?

Answer 46

The action of 2 kinases: 1) ATM 2) ATR

Question 47

What does ATM detect?

Answer 47

ds breaks in DNA

Question 48

What does ATR detect?

Answer 48

ss breaks in DNA

Question 49

What doe ATM and ATR do when they identify DNA breaks?

Answer 49

1) Phosphorylate downstream signalling 2) Activation of many downstream pathways 3) Leads to cell cycle arrest, activation of DNA repair and apoptosis (if repair not possible)

Question 50

Where do the signalling pathways of ATM and ATR converge?

Answer 50

On P53

Question 51

What is P53?

Answer 51

- A TUMOUR SUPRESSOR that is lost or mutated in ~50% of ALL human tumours - TRANSCRIPTION FACTOR - promotes the expression of genes mediating growth-suppression, apoptosis and DNA repair

Question 52

What is the target gene of P53? What does this do?

Answer 52

p21^Cip1 (a CKI) Blocks the progression of cell cycle