Lecture 5: Cell Cycle - cyclins, CDKs, CDKIs

Question 1

What is a CDK

Answer 1

Cyclin dependent kinase

Question 2

What is a CDKI

Answer 2

Cyclin dependent kinase inhibitor

Question 3

What are the key control proteins

Answer 3

CDK’s and cyclins.

Cyclins bind to CDKs and affect the latter’s ability to phosphorylate serine and threonine residues of their substrates

CDKs - 40% homology with each other
Cyclins - 100 AA residue long domain

Question 4

How do CDK/cyclins interact with checkpoints

Answer 4

activity levels of particular CDK/cyclins allow the cell to pass through the different checkpoints (G1 to S, intra-S, G2 to M) of the cell cycle

ALSO A SPINDLE CHECKPOINT

Question 5

What is a restriction point?

Answer 5

The G1 to S check point - cyclin D is responsive to growth factors (E.g., Ras pathway) which pushes the cell from G1 to S

(Before its too late. Once past it either happens or the cell self destructs)

Question 6

How are CDKs regulated

Answer 6

Via protein - protein interactions and phosphorylation’s (can be +/-)

Results in conformational change (opening a substrate cleft)

Without binding to a cyclin (or other) its binding to substrates is poor

Question 7

What are cyclns

Answer 7

‘Cyclin box’ - 100 aa to interact with kinase partner (CDK)

Transcriptional and post-transcriptional controls on cyclin levels:
1. transcription: TFs, activity of cyclin/CDK themselves (one cyclin/CDK will lead to increased transcription of the next)
2. Protein abundance: rapid degradation
3. Protein localisation

Question 8

What are CDK concentrations like

Answer 8

VERY CONSTANT

Question 9

What are cyclin concentrations like

Answer 9

VARIES, the concentrations correlate with check points

Question 10

What are D type cyclins

Answer 10

3 types (D1, D2, D3)

Levels can be induced by growth factos (mitogens) = integration of multiple signlalling pathways through control of CDK4 & 6 activation

Control G0 progression to G1

ONCOGENE ACTIVATION DRIVES CYCLIN D EXPRESSION

Different cyclins bind to different CDKs - the right cyclin binding also impacts
its level of activity

Question 11

Can a Cyclin-CDK complex be further regulated?

Answer 11

YES.

They are subject to further positive and negative regulation via phosphorylation

E.g., phosphorylation a C terminus threonine residue by CAK is required for activity

Phosphorylation of two residues near N terminus threonine tyrosine is inhibitory

CDC25 phosphatases remove inhibitory phosphorylation

Question 12

What are the CDKs / cyclins for G1 check point

Answer 12

D1, 2, 3 cyclins
CDK4, 6

Cyclin E1, E2 & CDK2 - critical initiators of DNA replication, chromatin unwinding, DNA polymerases.

Cyclin E transcribed by E2F

Question 13

What are the CDKs / cyclins for S/G2 check point

Answer 13

Cyclin A1, A2 & CDK2
- phosphorylates DP1 (part of E2F complex) inactivating it

Question 14

What are the CDKs / cyclins for M check point

Answer 14

Cyclin B1, B2 & CDK1 (CDC2)
1. directs nearly all processes during mitosis including chromosome condensation and segregation
2. promotoes mitotic spindle formation
3. degradation of cyclin B essentail for exit of M phase

Question 15

Why is CDK1 essential?

Answer 15

Key for mitotic progression

Only possible CDK capable of initiating M phase

CHK & WEE1 mediate DNA replication and M phase via CDK1 activitiy

Question 16

How are CDKs activated by non-cyclin proteins

Answer 16

KSV: K cyclin (~30% identiyy to cyclin D2), binds to CDK6 & 4. Inactivates pRB

RINGO/Speedy proteins (Human Spy1) activate CDK1 & 2 (no similarity to cyclins)

Question 17

How do cyclin/CDK alterations lead to cancer?

Answer 17

slow degradation or bad binding affintities can elgonate certain phases

Question 18

How does Cyclin D1 cause cancer

Answer 18

too much Cyclin D1 = Breast carcinoma leukaemia’s

reduced degradation due to depressed GSK3 activity = diverse tumours

Question 19

How does Cyclin E cause cancer

Answer 19

Over expression = breast carcinoma

Defective degradation due to lo,ss of hCDC4 = endometrial, breast, ovarian carcinomas

Question 20

How does CDK4 cause cancer

Answer 20

Structural mutation (INK4 resistance) = melanoma

Amplification = several types

Question 21

What are the two types of CDKIs

Answer 21

INK4 proteins:
Inhibit CDK4/6

Cip/Kip proteins:
inhibit all other cyclin-CDK complexes

TGF-beta = growth inhibitory by inducing p15^INK4B (& weakly induces p15^Cip1/Waf1)

DNA damage induces p21^Cip1/Waf1

Question 22

How are CDKs regulated?

Answer 22

Cyclin (or RINGO/speedy) binding

CDKIs (these are also regulated)

+/- phosphorylations

Question 23

What do mitogens do?

Answer 23

Inhibit CDKIs by activating PI3K->PKB/Akt (pathway)

Question 24

What is the role of the PKN/Akt pathway?

Answer 24

phosphorylates p21^Cip1/Waf1:
Causes export from nucleus to cytoplasm, no longer engage and inhibit cyclin-CDK complexes

Phosphorylates p27^Kip1: cannot locate nucleus

Question 25

Loss of CDKI activity leads to cancer

Answer 25

Loss of p16^INK4a or Rb allows prevents the cell from stopping proliferation

Question 26

How might p21^Cip1/Waf1 have oncogenic potential