9-15 Cell Cycle II

Question 1

What are the 6 main stages of M-Phase (Mitosis)

Answer 1

1st: Prophase
2nd: ProMetaphase
3rd: Metaphase
4th: Anaphase
5th: Telophase
6th: Cytokinesis

Question 2

List Main events of Prophase [2]

Answer 2

• Replicated sister chromatids condense

* Mitotic spindle assembles outside the nucleus inbetween 2 centrosomes (which had replicated and moved apart)

Question 3

List Main events of ProMetaphase [2]

Answer 3

• ABrupt start w/Nuclear envelope breakdown

* Chromosomes attach to spindle microtubules via their kinetochores and actively move

Question 4

Main events of Metaphase [2]

Answer 4

• Chromosomes are aligned at equator of spindle, midway between spindle poles
• Kinetochore microtubules attach each sister chromatid of a pair to OPPOSITE poles of the spindle complex

Question 5

Main events of Anaphase [2]

Answer 5

*Sister chromatids synchronously separate slowly apart into 2 daughter chromatids (on their own)

*Kinetochore microtubules get shorter and spindle poles also move apart PULLING sisters apart
[[both processes contribute to chromosomes separation! ]]

Question 6

Main events of Telophase [4]

Answer 6

*2 sets of daughter chromosomes arrive at poles of the spindle and DECONDENSE NOW!

• New Nuclear envelope reassembles around each set!
• Formation of 2 nuclei and End of Mitosis is marked!
• Assembly of contractile ring begins cytoplasm division

Question 7

Main events of Cytokinesis [3]

2)How are myosin II and actin distributed during this event?

Answer 7

• Contractile ring of actin and myosin filaments divide the cytoplasm
• Pinching of cell into 2 creates 2 daughter cells(each with one nucleus)
• Reformation of interphase array of microtubules nucleated by the centrosome

2)MOST myosin II rushes to contractile ring to help “pinch” off cytoplasm but only some actin goes to ring. The rest of actin remains in cortex of nascent daughter cells

Question 8

1) What is the nuclear lamin?
2) Structure of Nuclear Lamin?

3)What is Nuclear Lamins relation to Mitosis?

Answer 8

1) basal membrane connecting envelope to inner cell
2) Regular lattice of specialized intermediate filaments

3) In ProMetaphase Lamin is phosphorylated by M-Cdk and because of this [P] Lamin falls apart! = Nuclear Envelope Breakdown

Question 9

A: When is Nuclear Lamin DePhosphorylated and what effect does this make?

B: What enzyme does this DePhosphorylation?

Answer 9

A: Nuclear Lamin is Dephosphorylated during Early Telophase–>Nuclear Envelope REFORMS!

B: Done by phosphatases only when M-Cdk isn’t active

Question 10

When does the Fusion of Nuclear Envelope Fragments and Fusion of Enveloped Chromosomes occur?

Answer 10

Telophase!

Question 11

Where do the Nuclear Membranous fragments used to build ___ ____ actually come from?

Answer 11

Nuclear Membranous fragments for the Nuclear Envelope come from the ER

Question 12

Each Centrosome is made up of a ____ pair and ____ _____.

2) What happens in G1 to these centrosomes?
3) S phase?
4) G2 phase?
5) M phase?

Answer 12

Each Centrosome is made up of a centriole pair/duo and pericentriolar matrix juice

2) G1=2 centrioles of the pair separate
3) S = daughter centriole in each separated centriole starts to grow at right angle near base of their parent centrioles=elongation

4)G2 = Elongation of daughter centriole is complete by G2 and 2 centriole pairs remain close in single complex until beginning of M phase

5;) M=complex splits in two and the 2 halves begin to separate from each other–>centrosome that nucleates its own radial array of microtubules=ASTER

Question 13

Explain how the formation of the 2 poles of the mitotic spindles actually come about when entering M phase?

Answer 13

2 halves of the replicated centrosome (each w/its own centriole pair) surrounded in matrix, will split and migrate apart to innate the formation of 2 poles of the mitotic spindle
happens when entering M Phase

Question 14

What are some key differences between the Mother Centriole and daughter centriole in a centrosome centriole pair? [3]

B: When does the daughter centriole mature into this figure?

Answer 14

MOTHER CENTRIOLE= *Larger / *More Complex
*is the only centriole associated with the matrix that nucleates microtubules

B: daughter centriole matures to Mother during next cell cycle

Question 15

Centrosome is the major _____ of animal cells are usually located in the _______. It consist of ______.
B: Where is the [-] end of each centrosome microtubles?

C: Where is the [+] end of centrosome microtubules

Answer 15

Centrosomes are the major MTOC of animal cells and are located IN CYTOPLASM NEXT TO NUCLEUS. Consist of amorphous matrix w/proteins containing gamma-tubulin rings(which microtubules grow from)

B: [-] of centrosome microtubules are buried inside centrosome

C: [+] end of microtubules are FREE :-) in cytoplasm

Question 16

Microtubule is a stiff hollow tube made of ___ protofilaments aligned in ____. Subunits of these protofilaments are ____ ____ formed from ______.

Answer 16

Microtubule is a stiff hollow tube made of 13 PROTOFILAMENTS aligned in PARALLEL. Subunits of Protofilaments are [alpha/beta TUBULIN HETERODIMERS] formed by tight links between alpha and beta tubulin monomers with all SAME ORIENTATION

Question 17

Which tubulin monomer is the GTP TIGHTLY bound and considered an integral part of the protein?

Answer 17

ALPHA-tubulin monomer

Question 18

GTP and Beta-tubulin?

Answer 18

GTP is less tightly bound to betA-tubulin monomer and is important for filament dynAmics = polymerization/depolymerization

Question 19

How many Kinetochores total have to be captured by microtubule structures during Mitosis?

Answer 19

92!

Question 20

1) Explain MICROTUBULE catastrophe :-/
2) What is the GTP cap?

3)Explain MICROTUBULE Rescue :-)

Answer 20

1) happens when GTP nucleotide hydrolysis occurs FASTER than subunit addition–>cap ends up being lost—>MCT shrinkage and Catastrophe
2) GTP cap is formed by a growing microtubule chain that has multiple GTP-containing subunits at its end)
3) Rescue: GTP-containing subunits still add to shrinking end even despite loss of cap and does it enough to form a NEW cap–>MCT picks growth back up–>Rescue

Question 21

How does the loss of the GTP cap cause depolymerization in Microtubule filaments?

Answer 21

Forced linear conformation of [GDP-containing subunits] by lateral bonds within MCT wall are held solely by the GTP cap –>made of [GTP-containing subunits.] Loss in this cap allows GDP-containing protofilaments to relax into their CURVED form—->disintegration/disruption at shrinking end

Question 22

MAP

2) example?
3) How is this related to the Cdk?

Answer 22

proteins that stabilizes/preferentially binds to Microtubule [+] ends and promote polymerization with less dynamic MCT!
2) ex. XMAP215

3)Cdk phosphorylates MAP–>INactivation

Question 23

Catastrophin [5]
2) example?

3)How is this related to the Cdk?

Answer 23

1) proteins that oppose MAP, INC MCT dynamics, DeSTabilize microtubule polymers on [+] end–>shrinkage
- —>CATASTROPHE!

2) Kinesin-13 from the Kinesin Motor Superfamily
3) Cdk phosphorylate/ACTIVATES Catastrophins in order to make MCT more dynamic

Question 24

List and describe the 3 classes of dynamic Microtubules in mitotic spindles of metaphase

Answer 24

1. Astral MCT=interacts w/cell cortex to orient centrosomes AND eventually pull spindle pole boxes away from each other toward cell surface
2. Kinetochore MCT= attach each chromosome to the spindle pole box
3. Interpolar MCT = connects the two halves of the spindle boxes together while chromosomes are attched

Question 25

Which class of Microtubules undergo dynamic instability, _____ and ______ at their [+] ends

Answer 25

Astral MCT undergo dynamic instability both growing and shrinking in the cell cortex at its [+] end

Question 26

Kinetochore MCT and interpolar MCT both undergo ______ _____ _____ during mitosis. What does this even mean??

Answer 26

Kinetochore MCT and interpolar MCT both undergo CONTINUOUS DEPOLYMERIZATION FLUX during mitosis

• –>pulls both MCT toward opposite spindle pole boxes
• ———–>pulls apart sister chromatids in Anaphase

Question 27

What 3 things does Condensin do?

Answer 27

head domains of its major subunits [Smc2&Smc4] held by 3 subunits catalyze:

• the deep tight condensation of chromosomes
• formation of ring structure that encircles loops of DNA
• hydrolyzes ATP &coil DNA molecules in test tubes

Question 28

A: Describe Cohesin structure

B: How does Cohesion work?

Answer 28

A: 4 subunit complex with 2 main subunits [Smc1&Smc3] that act as coiled-coil proteins with ATPase domain at one end.

B: Scc1 and Scc3 connect the 2 ATPase head domains
–>forms ring structure that encircles sister chromatids

Question 29

1) What is the Kinetochore?
2) How many of these should you see during Metaphase?
3) How many Kinetochore MCT are involved in mammalian cells?

Answer 29

1) Connecting point plaque between [+] end Kinetochore MCT and the surface of the centromere during Metaphase
2) There is 1 Kinetochore PER sister chromatid = 2 total

3) ( 15-20 ) Kinetochore MCT attach to EACH kinetochore for sister chromatids = 40 total for 1 homolog chromosome

Question 30

Centromere

Answer 30

Series of A-T rich alpha satellite DNA sequences repeated thousand of times and then surrounded by flanking pericentric heterochromatin = 1 centromere

Question 31

[T or F] A cell can contain both an Active Centromere and an inactive centromere

Answer 31

TRUE

Question 32

What is the 5 component order (outside–>inner) of the centromere compnenets

Answer 32

5: Kinetochore MCT

4: inner/outer kinetochore plates
3: centromere-specific histone H3 variant chromatin
2: NORMAL histone H3 & dimethylated lysine 4 chromatin

1(most inner): cohesin linking sister chromatids together

Question 33

Explain the 2 forces that are at play in Mammalian Anaphase A

Answer 33

1. Motor proteins operating at kinetochores

2. depolymerization of kinetochore MCT pulling daughter chromosomes toward nearest opposite pole

Question 34

Explain the 2 forces at play in Mammalian Anaphase B

Answer 34

1. Elongation and Sliding Force generated between MCT from opposite poles pushing each other apart
2. Astral MCT as an outward force pulling poles away from each other toward cell surface

Question 35

What are common substrates of M-Cdk kinase and how does M-Cdk affect them? [6]

Answer 35

a: Condensin=chromosome condensation initiation
b: lamina (nuclear)=causes nuclear envelope breakdown
c: Motor proteins=move mitotic spindle
d: MAP=INactivates MAP–>MCT INstability and good dynamics

e: Catastrophin=Activates–>more MCT Instability and good dynamics [shrinking->growing->shrinking]
f: APC-CDC20=activates APC and BEGINS ANAPHASE

Question 36

1) Broken DNA Damage tends to activate ______ which phosphorylates ____ (thus __[activating/deactiving] it!) What does this ultimately do?
2) What’s a 2nd alternative way of blocking M-CdK using the first [missing factor] above? [2]

Answer 36

1)DNA damage activates ATM/ATR Kinases
—>phosphorylate p53–>ACTIVATES p53!
p53[P}=makes [p21/waf1/CKI] which STOPS M-Cdk activity.
——————————————————————————-
2)*ATM/ATR–(activtes)–> [Chk1/2 Kinase] –(P)–>
M-Cdc25 and INACTIVATES M-Cdc25.

**M-Cdc25 normally removes inhibitory phosphate from M-Cdk so if INactive= No Active M-Cdk = No M-phase

Question 37

Extensive DNA Damage is likely to lead to……

Answer 37

Extensive DNA Damage is likely to lead to CELL DEATH BY MITOTIC CATASTROPHE

Question 38

1) APC/C has the ability stop __-phase and “help” unglue ____ ____ during anaphase.
2) HOW DOES IT DO THIS? [3 points]

Answer 38

1) APC/C ubiquitinase ligase can stop {M-phase*anaphase} and “help” unglue sister chromatids during anaphase

2) **Cdc20 activates APC/C–>leads to degradation of
M-cyclin AND ubiquitylation/destruction of binding father securin—>releases separase protease as a result.

**the Now ACtive Separase cleaves Scc1(subunit of cohesion holding sisters together)=release sisters later pulled apart by mitotic spindle boxes.

Question 39

What happens to separase protease in animal cells if Cdks phosphorylate them?

2) what would be result of anaphase Cdk inactivation due to ____ destruction

Answer 39

In animal cells…When Cdks phosphorylate separase protease they inhibit it! –>prevent it from carrying out Anaphase

2) if cyclins were destroyed during anaphase, Cdk would be inactivated as result—->separase protease would now be DePhosphorylated and Free to carry out Anaphase freely

Question 40

1) APC-CDC20 is responsible for…….

2) How is APC-CDC20 activated?

Answer 40

1) APC-CDC20 binds to APC/C and activates its ubiquitinase activity.
2) APC-CDC20 is activated when all kinetochores {92} are attached to microtubules–(leads to)–>Activation of APC/C

Question 41

1) Explain the Mitotic Catastrophe Checkpoint involving Centromere/Kinetichore passenger proteins
2) How does Cancer cells distort this feature for their benefit?
3) What happens in “normal cells” when a cancer cell abnormality like this happens?

Answer 41

1-Kinetochore proteins (5) = Aurora Kinases, survivin,
APC-CDC20, BubR2 kinase and CENPs all dissociate when MCT bind –> promotes “true go” thru anaphase checkpoint

2-Cancer cells OVERExpress Aurora Kinases, Survivin & CENPs while undderexpressing BubR2 Kinase—>Tricks Cell to think Kinetochore proteins have fallen off and Promotes “false/premature go thru anaphase checkpoint”
—->polyploidy/genetic instability.

3)normal cells have a “wait period” the cell will give to wait on sister chromatid separation by APC but IF wait period runs out=MITOTIC CATASTROPHE APOPTOSIS by Taxol!

Question 42

CENPs

Answer 42

CENtromere Proteins apart of [Kinetchore protein group] involved in Anaphase wait signal.

When HIGH in cytoplasm—>ACTIVATES M-Cdk–> [P]binding father Mad2—>releases cdc20 to activate APC/C

Question 43

What 3 things does Jnk do?

Answer 43

1. phosphorylates Jun –> [+] tx!
2. phosphorylates p53————–>APOPTOSIS
3. phosphorylates BcL2—->APOPTOSIS