Cell Bio - Cell Cycle

Front 1

What happens in S phase?

Back 1

S is for synthesis
- chromosome duplication

Front 2

What happens in M phase?

Back 2

Mitosis and Cytokinesis

Front 3

What are mytosis and cytokinesis?

Back 3

Mytosis - duplicated chromosomes are segregated into a pair of daughter nuclei

Cytokinesis - cell divides into two

Front 4

What are the four phases of the cell cycle?

Back 4

1. G1 - gap phase between M phase and S phase
2. S phase
3. G2 - gap phase between S phase and M phase
4. M phase

Front 5

What are G-phases?

Back 5

- time for the cell to grow and replicate organelles and macromolecules, etc
- also provide time to check that everything is going well and environmental conditions are favourable

Front 6

What is a commitment point? Where is it located?

Back 6

- once you pass that point you are committed to the cell cycle and DNA replication
- commitment point is at the end of G1

Front 7

Why is yeast a good model system for the cell cycle?

Back 7

- it reproduces rapidly due to small genome
- easy to manipulate genetically
- proliferate in haploid state so you can just look at one of the chromosomes

Front 8

What are CdC genes?

Back 8

- Cell-division Cycle genes
- affect the cell cycle so as soon as you mutate them it will cause the cell cycle to arrest

Front 9

Why must Cdc mutant genes be conditional?

Back 9

- so that they cause a failure in function only at certain conditions
- usually temperature sensitive in which it fails to function only at high temperatures
- then can control at what point they stop, and at that point they grow abnormally large

Front 10

Why are Xenopus (frog) embryos a good model system for animals?

Back 10

- egg of frog is over 1 mm in diameter and contains 100,00 times more cytoplasm than an average cell in the human body
- fertilization triggers rapid sequence of cell divisions in which cells don't grow but just duplicate again and again which can be observed

Front 11

What is another way to look at frog eggs in the most simple form?

Back 11

- can centrifuge to break open batch of frog eggs and separate the cytoplasm
- can then add sperm nuclei and ATP to this separated cytoplasm, allowing it to go through repeated cycles of DNA replication and mitosis

Front 12

What types of staining and antibodies are used to monitor cell cycle progression?

Back 12

- BrdU (bromo-deoxyuridine) an artificial thymidine shows up newly synthesized DNA
- or use a flow cytometer and fluorescent DNA binding dyes to detect the amount of DNA present

Front 13

What are the three major check points?

Back 13

1. Start checkpoint - before S phase, ensures environment is favourable
2. G2/M checkpoint - before mitosis and after G2
- ensures all DNA is replicated and environment is favourable
3. Metaphase-to-Anaphase Transition - before anaphase
- ensures all chromosomes are attached to the spindle

Front 14

What are Cdks and when are they active?

Back 14

Cyclin-dependent protein kinases
- active only when bound to cyclin

Front 15

What is cyclin? what does it do?

Back 15

- regalates Cdk, activate it and direct it to specific target protein
- concentration changes throughout the cell cycle

Front 16

What is the Statutory Analysis of In personam jurisdiction?

Back 16

Most states have statutes granting their courts personal jurisdiction in 4 situations:
(1) the D is physically present in the state at the time of service of process;
(2) the D is domiciled in the state;
(3) the D consents to jurisdiction in the state;
(4) a long arm statute authorizes personal jurisdiction over the D.

Some long arm statutes simply say they reach the constitutional limit. Such as **CA**

Front 17

What is APC/C and what does it do?

Back 17

- catalyzes ubiquitylation and degradation of 2 main proteins:
1. securin
2. S- and M- cyclins
- as soon as this complex is active all cyclins are degraded and no cyclins remain active
- usually takes place during mitosis once all Cdks have been activated

Front 18

How are Cdks activated?

Back 18

- in the inactive state, without cyclin bound, the active site is blocked by a region of the protein called the T-loop
- cyclin binding causes the T-loop to move out of the active site, resulting in partial activation of the Cdk
- phosphorylation of the Cdk on the T-loop further activates the enzyme by chaning its conformation to enhance substrate binding

Front 19

What are the two ways that Cdk can be inhibited?

Back 19

- Wee1kinase can phosphorylate 2 closely spaced sites above the active site to inhibit activity of the enzyme
- p27 can bind to both cyclin and Cdk distorting the active site and blocking ATP binding

Front 20

What removes Wee1 kinase phosphates from the cyclin-Cdk complex to reactivate it?

Back 20

Cdc25 phosphatase

Front 21

What three things are required for the Cdk kinase activity to turn on?

Back 21

1. Inhibitory phosphate must be removed
2. Activating phosphate must be present
3. Cyclin must be present

Front 22

How does APC/C become active and how does it work?

Back 22

- APC/C is activated in mitosis by Cdc20 activating subunit
- APC/C then transfers multiple ubiquitin molecules onto the target protein to prepare it for degradation

Front 23

What is the SCF complex?

Back 23

Skp/Cullin/F-box Complex

Front 24

What does the SCF complex do?

Back 24

- ubiquitylates certain CKI (Cdk Inhibitor) proteins in late G1, therefore helping to control activation of S-Cdks and DNA replication

Front 25

how does the SCF complex work?

Back 25

- F-Box proteins on the SCF help it to recognize its target proteins (which must be phosphorylated) and then SCF ubiquitylates CKI for degradataion in the proteasome which then allows for activation of CDKs

Front 26

How is DNA replicated during S phase?

Back 26

- In early G1, Cdc6 and Cdt1 associate with the ORC (origin recognition complex) which bring in MCM helicase which starts to unwind DNA. These together from the pre-Recognition Complex.
- S-Cdk triggers S Phase by phosphorylating ORC and Cdc6 leading Cdc6 to be degraded. Cdt1 is then inhibitied by geminin.
- the preinitiation complex binds to ORC until DNA is replicated and then it is released.
- product at the end is 2 sister chromatids.

Front 27

What is cohesin and what does it do?

Back 27

- protein complex with four subunits that form a ring structure and hold sister chromatids together

Front 28

What is DNA catenation?

Back 28

- 2 sister DNA molecules are often intertwined and must be cut by the enzyme topoisomerase II which opens them so they can separate and then reseals them

Front 29

What two things does APC/C trigger the destruction of in mitosis?

Back 29

1. destruction of securin --> liberation of separase --> separae cuts cohesin which allows for chromatid separation
2. destruction of cyclins --> dephosphorylation

Front 30

What is M-Cdk and what does it do?

Back 30

- responsible for all cell rearrangements during early mitosis
- phosphorylates proteins involved in assembly and disassembly of mitotic spindle

- Cdk1 is associated with M-cyclin
- M-Cdk complex is phosphorylated on an activating site by CAK (Cdk-Activating Kinase) and on an inhbitory site by Wee1 Kinase
- the resulting inactive M-Cdk is then activated at the end of G2 by Cdc25 which removes the inhibitory protein
- Cdc25 is further stimulated by active M-Cdk = positive feedback, which is enhanced because M-Cdk also inhibits Wee1

Front 31

What are the three types of microtubules involved in mitotic spindle formation? What do they do?

Back 31

1. Kinetichore microtubules - connect spindle poles with kinetichores of sister chromatids
2. Interpolar microtubules - push centrosomes away from each other
3. Astral microtubules - help position centrosome

Front 32

What are the four motor proteins involved in spindle assembly and what do they do?

Back 32

1. Kinesin-5 - 2 motor domains, move toward plus ends of microtubule sliding the 2 antiparallel microtubules past each other forcing the poles apart

2. Kinesin-14 - single motor, move to minus end, pulls microtubules/poles together

3. Kinesin-4 and Kinesin-10 - attach to chromosomes, waling toward plus end moving them away from the poles

4. Dynein - move to minus end, pulling centrosomes to poles of cell

Front 33

What is bi-orientation?

Back 33

- stable attachment of microtubules pulling equally on each side from the kinetichores of the sister chromatids

Front 34

What happens if incorrect unstable attachment of microtubules to kinetichores?

Back 34

- tension sensing by kinase aurora-B which then phosphorylates microtubule attachment site to decrease affinity for microtubule

Front 35

What is the process for APC/C promoting Anaphase ?

Back 35

- Cdc20 activates APC/C which leads to ubiquitylation and destruction of securin
- allows separase to activate
- separase cleaves Scc1 - a subunit of the cohesin complex holding sister chromatids together allowing them to separate -- anaphase

Front 36

What is the difference between Anaphase A and Anaphase B?

Back 36

Anaphase A - chromosome movement toward the poles depends on depolymerization of kinetichore at the plus and minus end, this sortening of kinetichore microtubules moves daughters to the poles

Anaphase B - two spindle poles move apart with help of motor proteins sliding interpolar microtubules against each other to move centrosomes to opposite poles

Front 37

What is telophase?

Back 37

- disaseembly of mitotic spindle
- reformation of nuclear envelope
- all proteins that were phosphorylated during mitosis are then dephosphorylated to trigger these processes

Front 38

What are the four steps in cytokinesis?

Back 38

1. Initiation - cleavage furrow appears on cell surface
2. Contraction - contractile ring (made of myosin an dactin filaments) contracts
3. Membrane insertion
4. Completion

Front 39

What is cytokinesis triggered by?

Back 39

Dephosphorylation of Cdk

Front 40

What are the steps in contracting the ring?

Back 40

- RhoA is required for activating formins and Rho-activated kinase
- this kinase in turn inhibits myosin phosphatase and promotes myosin II activation
- the kinase and formins are needed for assembly and contraction of actin-myosin ring

Front 41

RhoA is activated by what?

Back 41

RhoGEF

Front 42

How is cytokinesis different in plan cells?

Back 42

- they have a cell wall and therefore cannot make contractile ring
- early cell plate formed by vesicles from Golgi with cell wall material fuse to synthesize a new cell wall

Front 43

What are the three major signaling molecules that determine cell size and number?

Back 43

1. Mitogens - stimulate cell division by activating G1/S-CDK
2. Growth factors - stimulate increase in cell mass by synthesis of proteins/macromolecules without dividing
3. Survival factors - promote cell survival by suppressing apoptosis

Front 44

What are mitogens? What do they do?

Back 44

- more than 50 known mitogens
- can stimulate cell division, cell growth, survival, differentiation, etc
- release brake on CdK activity
- can activate Ras and MAP kinase pahtway, leading to increased Myc
- Myc then increases levels of G1-cyclins by activating transcription for it and therefore increasing levels of G1-CDK

Front 45

What does DNA damage cause?

Back 45

- DNA damage causes protein kinases ATM or ATR and Chk1 or Chk2 to be recruited
- these phosphorylate the gene regulatory protein p53
- Mdm2 usually binds p53 and promotes its ubiquitylation but phosphorylation of p53 blocks this binding so p53 accumulates and stimulates transcription of the gene that encodes p21

Front 46

What happens in unicellular organisms if DNA can't be fixed?

Back 46

- initially arrest cell cycle but eventually resume the cell cycle because life with mutation is preferable to no lie

Front 47

What happens in multicellular organisms if DNA can't be fixed?

Back 47

- weigh the overall health of the entire organism over the individual and the cell will undergo apoptosis

Front 48

What is a way to make the cell arrest or undergo apoptosis?

Back 48

- Abnormally high levels of Myc (stimulated by Ras) cause the activation of Arf which binds and inhibits Mdm2 and therefore increases p53.

Front 49

How is the cell cycle reset in cells with no G1 phase?

Back 49

- in early embryonic cell cycles, Cdc20-APC/C activity rises at the end of metaphase, triggering M-cyclin destruction leading to APC/C inactivation, which allows M-cyclin to accumulate again

Front 50

How is the cell cycle reset in cells with a G1 phase?

Back 50

1. Cdn1-APC/C activity increases in late mitosis, keeping M-cyclin level low in G1
2. Increases Cdk inhibitory proteins (CKIs)
3. Decreased cyclin gene expression
- GI/S-Cdk activity rises in late G1 and releases suppression of Cdk activity - leads to entry into S phase

Front 51

What three factors determine the size of organs and bodies?

Back 51

1. Cell Growth
2. Cell division
3. Cell death