Lecture 3 - Eukaryotic Cell Division

Question 1

What does Cyclin A-Cdk2 cause?

Answer 1

It triggers replication origin firing to initiate DNA replication during S phase

Question 2

What happens at the G2/M phase checkpoint?

Answer 2

1. Ensure that all of the chromosomes have been accurately replicated without mistakes or damage
2. Cell size and energy/protein reserves are assessed
3. Can sister chromatids separate correctly?
4. Are all chromosomes attached to the mitotic spindle?
   Between Interphase and Prophase

Question 3

What happens at the Metaphase-Anaphase checkpoint? Also known as Spindle Assembly Checkpoint (SAC)

Answer 3

1. Checks that conditions are OK to enter M phase
2. Has DNA replication occurred correctly?
3. Are preparations for M phase complete?
4. Cell size
   If yes, then the cell enters the late stages of M phase
   Between Metaphase and Anaphase

Question 4

How was the dominant M Phase Promoting Factor (MPF) found?

Answer 4

When M phase cells and cells in any other phase were mixed it was observed that the interphase cell would always start to segregate its chromosomes early (even if replication was incomplete). Both nuclei would enter mitosis synchronously

Question 5

How does MPF work?

Answer 5

MPF promotes the entrance into mitosis (M phase) from the G2 phase by phosphorylating multiple proteins needed during mitosis

Question 6

How is MPF controlled?

Answer 6

It is controlled by the periodic accumulation and degradation of Cyclin B during cell cycle progression

Question 7

How does Cyclin B work?

Answer 7

Their levels gradually rise during G2 and then associates with the M phase Cdk, Cdk1

Question 8

What does the mitotic spindle do?

Answer 8

It moves segregated sister chromatids to opposite ends of the dividing cell in Anaphase

Question 9

What is the mitotic spindle composed of?

Answer 9

Microtubules - dynamic polymers of tubulin

Question 10

Where do the microtubules form the mitotic spindle interact with the sister chromatids?

Answer 10

At protein complexes called Kinetochores

Question 11

Where to the microtubules from the mitotic spindle interact with the spindle poles?

Answer 11

At the centrosomes

Question 12

How are microtubules arranged?

Answer 12

They are composed of linear chains of tubulin dimers that are organized into a hollow cylinder

Question 13

How are microtubules regulated?

Answer 13

1. A rapidly growing microtubule has GTP-bound subunits at its plus end forming a ‘GTP-cap’ structure.
2. New subunits can be added to this structure to promote MT growth from the + end
3. If this ‘GTP cap’ is lost (due to GTP hydrolysis to GDP) then the subunits begin to disassemble and the MT shrinks

Question 14

When do kinetochores form?

Answer 14

G2 phase

Question 15

Where do kinetochores form?

Answer 15

At the centromeres of replicated chromosomes

Question 16

Explain + and - ends of microtubules

Answer 16

+ ends project away from the spindle pole

- ends are anchored at the spindle poles by centrosomes

Question 17

What do Interpolar microtubules do?

Answer 17

Interpolar microtubules from the two poles interdigitate (interlock like fingers) at the spindle equator

Question 18

What do astral microtubules do?

Answer 18

They radiate out from the poles into the cytoplasm

Question 19

What ensures correct bio-orientation of sister chromatids?

Answer 19

A trial and error mechanism

Question 20

What does the SAC do until correct bio-orientation is achieved?

Answer 20

The SAC functions to inhibit progression into the later stages of mitosis

Question 21

Why do replicated sister chromatids not separate from each other before Anaphase?

Answer 21

Adhesion of sister chromatids to each other prevents chromosome separation before Anaphase

Question 22

Function of cohesin

Answer 22

Holds sister chromatids together

Question 23

Two distinct mechanisms by which Cohesin is removed at the SAC

Answer 23

1. Cohesin rings at the tips of the chromosomes are easily lost/removed without any enzymatic activity - they are loosely held together
2. Cohesin rings at centromeric regions are very tightly bound and need to be removed by a proteolytic enzyme known as Separase

Question 24

What is the function of Separase?

Answer 24

It removes cohesin rings at centromeric regions

Question 25

How does the APC/C initiate sister crhomatid separation?

Answer 25

1. Active M-phase cyclin B/cdk1 complexes initiate the assembly of the mitotic spindle and these MTs attach to sister chromatids via their kinetochores
2. Active M-phase cyclin B/cdk1 complexes phosphorylate and activate the APC/C complex
3. APC/C binds to Cdc20 protein
4. APC/C induces ubiquitinylation and destruction of a protein known as Securin
5. This allows Separase to cleave the remaining Cohesion complexes that are holding the sister chromatids together

Question 26

What does Securin do?

Answer 26

Normally holds the proteolytic enzyme Separase in an inactive state during metaphase

Question 27

What else does the APC/C result in during Anaphase?

Answer 27

Destruction of remaining cyclins (S phase or M phase) leaving the cell with little or no Cdk activity during Anaphase