Module 5 second half

Question 1

cdc 2

Answer 1

cyclin dependent kinase (CDK)

Cdc 2+ = wild type = wee form

Cdc 2- = recessive = excess of wee1 or deficit of cdc25

Cdc 2D = deficit of wee1 (kinase inhibits mitosis) or excess of cdc25 (phosphatase - drives mitosis)

Question 2

Kinase and phosphatase cascade

Answer 2

controls entry into mitosis

Wee 1 - phosphorylates Y15 of mitosis cdk, inactivating it
CAK - activated the Miro to f kinase by phosphorylation
Cdc25 - removes an inhibitory phosphate from mitotic cdk

Question 3

Mitosis promoting factor (MPF)

Answer 3

mitotic cyclin/CDK

produced in G2 but kept phosphorylated (inhibited) until cell ready for mitosis

Question 4

Checkpoints

Answer 4

• points in cell cycle where cell can stop if something has gone wrong
• driven by proteins that inhibit cell cycle progression

Question 5

Mitotic checkpoint

Answer 5

DNA replication incomplete

ATR1 – CHK1 – (inactivates) – Cdc25

(ATR1 - checkpoint protein that is activated by single stranded DNA)

Question 6

G2/M summary

Answer 6

1. yeast mutants identified crucial players in G2/M transition, including activators and inhibitors of mitotic cyclin/CDK.
2. phosphorylation of different CDK residues can inhibit and activate the mitotic cyclin/CDK.
3. checkpoints ensure that cell has complete one ask before initiating the next. they target the major regulators of the cell cycle

Question 7

Mitotic cyclin/CDKs and nuclear envelope

Answer 7

• nuclear lamina support nuclear envelope
• lamins A,B and C form lamina structure
• mitotic cyclin/CDKs phosphorylate the lamin proteins, causing meshwork to breakdown

Question 8

APC/C ubiquitin-protein ligase

Answer 8

ubiquitinates mitotic proteins at key stages of mitosis
1. induces anaphase - metaphase-anaphase transition
2. induce late steps in mitosis (late anaphase) (along with Cdc14)

Question 9

Phosphatase PP2A

Answer 9

associated with centromeres and prevents cohesins near the centromere from being removed by phosphorylation - therefore they must be removed another way

Question 10

Metaphase/anaphase transition

Answer 10

separation of chromatids

APC/C initiates this by inducing cohesin removal at the centromere.
(ring that surrounds chromosomes)
- cohesively keeps everything together (to remember)

1. cohesins are composed of smc proteins and Scc1
2. separase is a protease that cleaves Scc1
3. securin binds separase, inhibiting it
4. when all kinetochores bind MTs, Cdc20 binfd to APC/C
5. APC/C polyubiquitinates (dooms) securin
6. separase, uninhibited, cleaves Scc1
7. chromaids are free to separate. this define anaphase

Question 11

Degradation of mitotic cyclins

Answer 11

APC/C specificity changes late in anaphase once chromatids have separated

Cdc1 becomes active and leads the APC/C complex to the mitotic cyclin for degradation

Question 12

Ubiquitination in G1-S phase transition

Answer 12

CKI binds to CDK/cyclin - inactive

SCF targets CKI and ubiquitinates it so then the CDK/cyclin is now active
- ready for S phase (?)

Question 13

3 mechanisms of cell specification

Answer 13

cell to cell interactions
- cell beside you determines your fate

asymmetric
- identical inside but ones slightly larger

morphogen gradients
- Bicoid inhibits caudal = which creates tails
- nanos inhibits hunchback

CAM

Question 14

Necrosis

Answer 14

unplanned cell death

• rupture of cell and spilling contents to surrounding tissue
• caused by variety of attacks on cell
• toxins, burns, infection, inflammation

Question 15

Apoptosis

Answer 15

programmed cell death

• cell chopped up and packaged for removal. DNA fractionated, cell membranes pinch off into small structures
• fragments labelled for macrophages so they cleaned up through phagocytosis

1. EGL1 is produced by apoptosis signal
2. EGL1 binds to CED-9, releasing CED-4
3. CED-4, released from mitochondria; forms octamers in cytoplasm
4. CED-4 octamers convert an inactive CED-3 precursor to active CED-3
5. CED-3 is caspase, destroying various proteins

Question 16

CED-3

Answer 16

when active:
- cleaves lamins (nuclear envelope dissolves)
- activates endonucleases (DNA is digested)
- attacks cytoskeletal components (cell structure)
- attacks cell-cell adhesion proteins (anchors for membrane)
- cleaved (activates) itself (runaway chain reaction)

Question 17

CED-3 mutant

Answer 17

no apoptosis

Question 18

CED-4 mutant

Answer 18

no apoptosis

Question 19

CED-9 mutant

Answer 19

all cells die

Question 20

ED1 mutant

Answer 20

no apoptosis

Question 21

Phosphatidylserine (Ps)

Answer 21

marks apoptotic particles for phagocytosis
- PS is moved to outside of membrane of apoptotic cells, marking them for phagocytosis by macrophages

Question 22

cancer cells select for mutations in two types of genes

Answer 22

proto-oncogenes
- genes that normally promote cell growth. when mutated or amplified they become oncogenes

tumour-suppressor genes
- normally inhibit cell cycle progression. when theyre mutated, cells divide out of control

Question 23

Retinoblastoma

Answer 23

• rare childhood cancer
• heritable and sporadic cases
• caused by mutations in Rb gene

= loss of heterozygosity

Question 24

The G1 restriction point is a target of cancer

Answer 24

• Rb rlly important for G1 of cell cycle
• 2 major kinases in G1 that promote entry into S-phase = G1 CDK + G1/S CDK
• these kinases phosphorylate Rb
  (if no mitogens = go into G0)
• restriction point (R or START): cell committed to another round of cell division and is no longer mitogen sensitive
• Rb represses E2F-E2F promotes expression of replication genes
• G1 phosphorylates Rb and Rb falls off E2F

Question 25

Rb

Answer 25

poor substrate = needs to be phosphorylated multiple times a repressor of E2F txn factor

Question 26

Inactivation of APC/Cdh1 (new commitment point?)

Answer 26

APC/Cdh1 needs to be inactivated before cells enter S-phase - G1/S CDK phosphorylated Cdh1 - Emi1 leads another Ub ligase to destroy Cdh1

Question 27

p16

Answer 27

binds CDK, preventing cyclin/CDK joining. Expression of p16 is driven by stresses placed on the cell

Question 28

G1cyclin/CDKs

Answer 28

phosphorylate (inactivat) Rb

Question 29

E2F

Answer 29

A TF that drives expression of S-phase genes

Question 30

p53

Answer 30

gatekeeper of cell - detects conflicts in cell and can activate several pathways to fix them - loss of p53 doesn't make a cancer cell, it predisposes a cell to cancer. occurs through loss of tumour suppressor function - ....

Question 31

transcriptional regulation

Answer 31

- distinct genes can be turned on or off - distinct mRNAs translated or repressed and localised to different regions of the cell

Question 32

epigenetic regulation

Answer 32

marks on DNA that regulate gene expression and are inherited by daughter cells, but independent of DNA sequence permanently bind Rb to E2F = cell cycle arrest

Question 33

Rb pathway

Answer 33

1. cell signals that it wants to go unto S-phase and so P16 which is inhibiting G1/S CDK goes away (depends on stress of cell - once cell is no longer stressed it released and this continues) 2. g1 phase = G1/S cyclin binds to CDK and creates complex 3. G1/S cyclin CDK phosphorylates Rb (which is inhibiting E2F) 4. Rb leaves E2F and it is activates to create TFs for DNA replication and go into S phase

Question 34

Terminal differentiation

Answer 34

permanent cell cycle exit - differentiated cells become refractory to proliferate signals - G1/CDK inhibitors and Rb family members play major roles in cell cycle exit - terminal differentiation is distinct from senescence (td = permanent withdrawal from cell cycle)

Question 35

Cancer mutations

Answer 35

Gain of function = promote proliferation Loss of function = growth suppress, avoid checkpoints and genomically unstable Dominant negative = act like loss, mutant bind to wild type

Question 36

Regulation of gene expression

Answer 36

Epigenetic: histone acetylation (opens chromatin) and methylation (closes chromatin) Transcriptional: proteins that promote or inhibit gene expression