Lectures 3-4

Question 1

Types of inherited single gene defects

Answer 1

-Retinoblastoma, a childhood eye cancer (Rb)
-Li-Fraumeni syndrome (p53)
-BRCA1
-2 predisposition to breast and ovarian cancer (APC)

Question 2

Two forms of Retinoblastoma

Answer 2

-Familial (Hereditary): Multiple tumors, affecting both eyes, early onset, 6x increased cancer risk.
-Sporadic (Non-hereditary): Single tumor, affecting one eye, later onset.

Question 3

Phases of cell cycle:

Answer 3

-G1
-S
-G2
-M
-G0

Question 4

G1:

Answer 4

Cell make decisions about growth versus quiescence

Question 5

S phase:

Answer 5

DNA is synthesized

Question 6

G2:

Answer 6

Making sure all the DNA is replicated with fidelity

Question 7

M Phase:

Answer 7

-2 new cells are made through the division of the genetic material
-mitosis (prophase, prometaphase, metaphase, anaphase, telophase) and cytokinesis

Question 8

G0:

Answer 8

Resting, nonproliferative state of cells that have withdrawn from the active cell cycle

Question 9

What features do checkpoints have:

Answer 9

-Allow internal feedback control
-Allow control from external stimuli

Question 10

Types of cell cycle checkpoints:

Answer 10

-G1/S (determines if DNA synthesis can begin)
-Intra-S and G2/M (Prevents the replication of damaged DNA until repaired)
-Spindle Checkpoint (ensures chromosomes are aligned correctly to spindle. If not mitotic arrest doesn’t proceed to Anaphase)

Question 11

Progression to S phase depends on?

Answer 11

The presence of growth factors in the G1 phase up until the restriction point.

Question 12

What is the restriction point.

Answer 12

A checkpoint where a cell must make the decision of whether to proceed to S or to enter G0. At this point growth factors no longer influence progression.

Question 13

CDK Inhibitors:

Answer 13

-INK4/CDKN2 (p16, p15, p18, p19).
-CIP/KIP/CDKN1 (p21cip1, p27kip1)

Question 14

How does INK4 inhibit?

Answer 14

They bind to CDK4/6 which prevents it from binding to cyclin D

Question 15

How does the CIP/KIP/CDKN1 inhibit?

Answer 15

They can inhibit all of the other cyclin-CDK complexes by blocking their ATP binding site.

Question 16

What is CAK

Answer 16

-Stand for CDK activating kinases.
-Work by phosphorylating the activation loops.

Question 17

What is CD25

Answer 17

It is a class of phosphates that remove inhibitory phosphorylation’s that affect CDK2

Question 18

How are cyclin Ds activated?

Answer 18

By growth factors causing a signaling cascade resulting in transcription factor AP-1 which regulates key cyclins.

Question 19

How is cyclin E/CDK2 activated?

Answer 19

CDK4/6 sequestering p21 and p27

Question 20

Function of active cyclin E-CDK2:

Answer 20

It phosphorylates p27 which marks it for degradation by the proteasome, allowing for more active cyclin E-CDK2 complexes to be generated and leading to hyperphosphorylation of RB.

Question 21

What happens when RB is hyperphosphorylated

Answer 21

Hyperphosphorylation makes RB inactive which allows for the release of E2F and HDAC which are needed for S phase gene expression.

Question 22

Nuclear RB protein is a member of?

Answer 22

The “pocket proteins”

Question 23

What is HDAC

Answer 23

It is an enzyme (histone deacetylase) that when bound to RB it blocks S phase gene expression.

Question 24

What is E2F

Answer 24

-E2F is a key transcriptional regulator of S-phase genes
-Positive regulator of cell cycle progression
-Active E2Fs1,2,3 induce expression of late G1 genes, required for S phase.

Question 25

How is E2F1 degraded

Answer 25

In the G1/S transition cyclin A and CDK2 phosphorylates both E2F and DP subunits; leading to their dissociation, and loss of transcriptional activities.

Question 26

What are E2Fs1,2 and 3a

Answer 26

They are Hsp (heat shock protein)-chaperones also know as ‘activating E2Fs’

Question 27

S-Phase gene expression leads to:

Answer 27

-DNA polymerase, topoisomerase, ligase etc -Initiation Factors -Structural proteins (e.g. Histones) -Proteins involved in synthesizing DNA precursor nucleotides -Spindle assembly checkpoint protein e.g MAD2

Question 28

What cyclin is involved in G2/M Phase Progression

Answer 28

Cyclin B partnered with CDK1

Question 29

What keeps CDK1 inactive

Answer 29

The kinases WEE1 and MYT1

Question 30

What activates CDK1

Answer 30

Two independent events: 1. CAK-mediated phosphorylation 2. Cdc25-mediated dephosphorylation

Question 31

Targets of Active CDK1/Cyclin B:

Answer 31

– phosphorylates serine residues on lamin (breakdown of nuclear membrane) – Phosphorylates histones – Phosphorylates microtubule associated proteins (altered behavior during mitosis)

Question 32

What are CDC25 Phosphatases

Answer 32

-They are a family of phosphatases (A, B, C) that dephosphorylate specific tyrosine and threonine residues on CDKs (which inhibit CDK function) -Also dephosphorylates inhibitory phosphorylation residues on CDK2 in G1

Question 33

Which cell cycle stages are CDC25s involved in

Answer 33

G1-S and G2-M transitions.

Question 34

Why are CDC25 phosphatases targets of the checkpoint machinery during DNA damage

Answer 34

Because they allow for progression through the cell cycle, so in order to prevent this they are inactivated or degraded.

Question 35

Overexpression of CDC25A and B is seen in?

Answer 35

High grade tumors and poor prognosis cancers.

Question 36

What are checkpoint kinases (CHK)

Answer 36

They are kinases activated in response to DNA damage and target CDC25.

Question 37

What is Spindle Assembly Checkpoint

Answer 37

It ensures proper segregation of sister chromatids by inhibiting metaphase to anaphase transition until all chromosomes are attached correctly to the mitotic spindle.

Question 38

What are Cohesins?

Answer 38

Protein complexes that hold together sister chromatids.

Question 39

What is separase?

Answer 39

It is a protease which cleaves cohesins

Question 40

What happens when chromosomes are unaligned?

Answer 40

Separase is kept inactive by securin and Cdk1-cyclinB

Question 41

What happens when chromosomes become aligned?

Answer 41

CyclinB and securin are ubiquitinylated by the Anaphase Promoting Complex (APC)-CDC20 complex and degraded. This leads to the activation of separase, which releases sister chromatids, allowing for metaphase to anaphase transition.

Question 42

MYC represses expression of what?

Answer 42

p15 and p21

Question 43

TGFb reduces what expression?

Answer 43

1. c-MYC expression-Smad3 and E2F4/5 2. p107 shut down c-MYC expression

Question 44

TGFb-mediated repression of MYC is lost in what cancer?

Answer 44

Breast cancer

Question 45

How does TGFB induce expression of p21 and p15?

Answer 45

TGFb signalling: Smad2/3 associate with Smad4 heterotrimeric complex moves to nucleus and collaborates with the transcription factor Miz-1 to induce expression

Question 46

P21 and P15 are repressed in what cancers?

Answer 46

Any cancers where MYC is overexpressed

Question 47

How does TGFb block cell cycle progression.

Answer 47

By preventing phosphorylation of RB, which prevents it from releasing E2F.

Question 48

What is Akt

Answer 48

It is a serine/threonine kinase which is activated downstream of growth factor receptors. Its function is to promote cell cycle progression.

Question 49

3 functions of Akt:

Answer 49

-Inhibition of CDK Inhibitors (p21 and p27). -Promotes cell cycle progression by blocking the actions of GSK3b. -Blocks the actions of a pro-apoptotic protein Bad.

Question 50

How does Akt inhibit CDKIs p21 and p27?

Answer 50

Akt phosphorylates them which leads to their translocation from the nucleus to the cytoplasm where they are degraded by the proteosome

Question 51

Targets of GSK3b

Answer 51

It has inhibitory actions on MYC and CyclinD1.

Question 52

What does Autonomous cell division mean?

Answer 52

The cell no longer depends on positive and negative growth factors (i.e cells are now cancerous).

Question 53

What pathways play a central role in transmitting oncogenic signals.

Answer 53

-The PI3K-AKT pathway -The RAS-RAF-MEK-ERK pathways

Question 54

Increased Akt activity is frequent in what cancers?

Answer 54

Breast cancer.

Question 55

Tumor grade can be linked with?

Answer 55

Higher pAKT expression and reduced nuclear P27.