Introduction / Cell Cycle

Question 1

What are the steps of the signalling cascade once a growth factor binds its receptor?

Answer 1

1. Receptor tyrosine kinase dimerizes then autophosphorylates.
2. Phosphotyrosines activate the MAP-kinase and Ras pathways
3. MAP/Ras activate transcription factors which promote DNA transcription towards the cell cycle

Question 2

What are the major growth factors which utilize tyrosine-kinase receptors?

Answer 2

VEGF - vascular endothelial
EGF (receptor is EGFR) - epidermal
PDGF - platelet-derived
FGF - fibroblast

Question 3

How does RAS work?

Answer 3

It sits attached to the membrane, with a GTP binding domain.

Normally, its inactive form has GDP bound. Once activated by tyrosine kinase phosphorylation with a bridging protein, it has GTP bound.

GTP-bound Ras can activate RAF / MAPK cascades to signal cell proliferation

Question 4

How is RAS inactivated?

Answer 4

GTPase-activating proteins (GAPs) bind to activated Ras proteins (which also have intrinsic GTPase activity of their own) -> enhance GTP hydrolysis and termination of signal

Question 5

Give an example of GAP protein?

Answer 5

Neurofibromin - the GAP protein defective in NF-1

Question 6

What does activation of the MAPK cascade ultimately result in?

Answer 6

Rapid induction of MYC gene for production of MYC protein

Question 7

What are the two functions of MYC?

Answer 7

1. Can form a heterodimer with MAX (MYC-MAX) -> binds DNA and modulates genes promoting cell division -> i.e. cyclin D and CDK4
2. Can accumulate in unfavorable circumstances like nutrient deprivation, growth factor deprivation, or DNA damage and trigger apoptosis

Be aware -> MYC can be made in the absence of growth factors

Question 8

What is the function of CDKs?

Answer 8

Cyclins bind the cyclin dependent kinases (CDKs) at certain times in the cell cycle which phosphorylate key proteins which help the cell move forward

Question 9

What typically holds the cell in G0 (resting) or G1 (presynthetic growth - gap 1) phases?

Answer 9

Unphosphorylated or hypophosphorylated retinoblastoma is bound to the E2F transcription factor complex, inhibiting its function of production of growth factors needed for S phase

Question 10

What occurs generally in the G1 phase?

Answer 10

Cell preps for DNA synthesis -> protein / RNA synthesis with doubling of organelles

Question 11

How does the cell progress from G1 to S phase? When is the restriction point?

Answer 11

1. Growth factors lead to accumulation of cyclin D via MYC.
2. Cyclin D activates CDK which start phosphorylating Rb, allowing E2F to begin producing cyclin E (think E for E2F).
3. G1 restriction point -> point of no return where cyclin E has accumulated enough that the cell cycle will move forward without more growth factor / cyclin D
4. Hyperphosphorylated Rb has now completely lost control of E2F, which stimulates cyclin A from E2F-targeted genes

Question 12

What is the function of cyclin A?

Answer 12

Forms active cyclin A / CDK complexes, which cause binding and activation of DNA polymerase to commence S phase

Question 13

What happens in G2 phase and what cyclin predominates?

Answer 13

Preparation for cell division -> synthesis of microtubules and new membranes

Cyclin B predominates

Question 14

What actually initiates mitosis?

Answer 14

Phosphorylation of the cyclin B / CDK complex and accumulation of these complexes in the nucleus. These processes can be inhibited by G2/M checkpoint mechanisms.

Question 15

What is G1/S/G2 called vs M?

Answer 15

G1/S/G2 = interphase
M = prophase metaphase anaphase telophase

Question 16

When is mitosis said to be over?

Answer 16

When RB protein has phosphates removed -> regenerates the unphosphorylated, inhibitory regulator form

Question 17

How do cell cycle inhibitors work? Give an example of one.

Answer 17

They inactivate the cyclin / CDK complexes or inhibit their formation in the first place (directly / indirectly)

Example: p21

Question 18

What occurs at the G1/S checkpoint and how can it be stopped?

Answer 18

Checks for DNA defects prior to replication

If damage is detected -> increased activation of p53 tumor suppressor protein -> p53 upregulates DNA repair as well as p21 gene, which is a CDK inhibitor

Question 19

What happens if the DNA is successfully repaired at the G1/S checkpoint?

Answer 19

p53 will induce a protein to degrade itself -> essentially committing indirect suicide

Question 20

What happens if the DNA is unsuccessfully repaired at the G1/S checkpoint?

Answer 20

p53 will induce proapoptosis genes (i.e. BAX) and repress pro-proliferation / antiapoptosis genes like cyclins and Bcl2, leading to apoptosis / senescence

Question 21

What happens at the G2/M checkpoint? How can the cell be stalled here?

Answer 21

Cell checks for DNA defects after replication and before separation of the chromatids

If damage is detected, stall can occur via p53-dependent or independent mechanisms

Question 22

What major cyclins are present at each phase of the cell cycle?

Answer 22

G1 - cyclins D -> E
S - cyclin A
G2 - cyclin B
M - cyclin B

DEAB