Lecture 6 - The Cell Cycle: Mitosis and Cytokinesis Flashcards
ILOs
- Describe in some details how the cell cycle is regulated and how MTs and actin orchestrate mitosis and cytokinesis
- Understand how tools in biology have led to the discovery of the cell cycle
- Understand that timing is important for cell function (“when”)
- Bridge the role of sub‐cellular structures to consequences for the cell as a whole, and for health and disease
- Interpret scientific data
The body of an adult human:
- is composed of ~30 trillion cells (3x1013) = 0.5 billion cells per gram!
- 25 million cell divisions every sec of our lives
- every cell contains the same number of chromosomes!
- defects in cell division lead to chromosome abnormalities (development, cancer, etc.)
How do eukaryotes do it?
• common strategy for all eukaryotic cells:
replicate genome DNA synthesis > S phase
segregate genome Mitosis > M phase
- S phase and M phase must be carried out: accurately, only once and in correct order
- S phase and M phase separated by gaps: G1 and G2
G1>S>G2>M
• gaps allow control checkpoints at G1/S and G2/M ensure accuracy
The Cell Cycle
G1‐S‐G2‐M
G2 has twice the amount of DNA and twice the amount of cytoplasm
Mitosis = nuclear divison
Cytokinesis = cell divison
Checkpoints ensure everything is fine
G1, G2 and Mitotic checkpoints
G1 is the environment favourable?
G2 is all DNA replicated? Is DNA damage repaired?
M phase - Are all the chromosomes properly attached to the mitotic spindle?
Data handling practice
A mutant yeast stops proliferating when shifted from 25°C to 36°C. When the cells are analyzed using a machine that sorts cells according to the amount of DNA they contain, these graphs are obtained. Which of the following would not explain the results with the mutant?
A‐ inability to initiate DNA replication
B‐ inability to begin M phase
C‐ inability to activate proteins needed to enter S phase
D‐ inappropriate production of a signal that causes the cells to remain in G1
Answer: B
At 36°C, the cells all have one genome‐worth of DNA, meaning that they have not replicated their DNA and therefore have not entered S phase. Cells that are unable to begin M phase should have two genomes‐worth of DNA, as they would have completed DNA replication and arrested in G2.
The cell cycle is controlled by:
1) cyclically activated cyclin‐dependent kinases (Cdk)
- cyclin dependent kinase only activates when it binds to cyclin
‐ A cyclin binds its Cdk and activates it
‐ The concentration of cyclins oscillates during the cell cycle, not that of Cdk
‐ Activated Cdk make the cell cycle progress
Hartwell and Nurse discovered the cell cycle through the generation of yeast ts mutants by forward genetics
- all cells will pass the point of the cell cycle where the protein is active
- all cells arrest at point of function of mutant protein (tells you where the normal protein was functioning)
Data handling practice
The diagram shows the cell cycle of WT S.pombe cells. P. Nurse isolated a wee1 mutant and a cdc25 mutant. Based on the length of the cells, what do you thing the normal Wee1 and Cdc25 proteins do?
A‐ Both proteins are involved in M phase
B‐ Wee1 is a G1/S protein that blocks entry in G2 C‐ Wee1 blocks entry in M phase
D‐ Cdc25 orchestrates cell growth in M phase
E‐ Cdc25 allows entry in M phase
Answer: C, E
wee1ts can divide, but is small = goes faster through cell cycle = normal Wee1 blocks the G2/M transition. If normal Wee1 blocked entry in G2, then wee1ts cells would be long.
Cells grow in G2, not in M phase so the length of cdc25ts cells is due to a block in G2, not in M = cdc25ts blocks cells in G2, cells do not divide so keep growing = normal Cdc25 induces entry in M.
The cell cycle is controlled by: 2) phosphorylation
Phosphorylation is a molecular switch
- kinase = enzyme that catalyses the addition onto a substrate, of a phosphate group from ATP (phosphorylation). E.g. Cdc2 and Wee1
- phosphatase = removes a phosphate « de‐phosphorylate ». E.g. Cdc25
- some factors can modify the activity of kinases or phosphatases, incl. other kinases or phosphatases. E.g. Cdc25 and Wee1 regulate Cdc2.
Wee1 and Cdc25‐ Yin and Yang
- Wee1 is a negative regulator of cell division
- Cdc25 is a positive regulator of cell division
- Wee1 is a kinase; Cdc25 is a phosphatase
- S. pombe Cdc2 is now known to be a Cdk1 and Cdc13 a cyclin B
Human Wee1 is in an anti cancer trial
The cell cycle is controlled by:
3) destruction (proteolysis) of the cyclins
‐ Ubiquitylation by the Anaphase Promoting Complex (APC)
Why is Cdk1 (Cdc2) so important?
- Cdc2 is itself a kinase. It has at least 200 substrates.
- phosphorylates as cell crosses G2/M boundary:
- Mitotic spindle forms
- Cytoskeleton disassembles
- Golgi/ER fragments
- Nuclear envelope breaks down
- Chromatin condenses
- Nucleolus disperses
Mitosis: IPMAT
The aim of mitosis is to segregate an equal number of chromosomes into each daughter cell
- Interphase - Prophase - Pro-metaphase - Metaphase - Anaphase - Telophase
The critical role of MTs: the mitotic spindle
- Astral microtubules - attach the spindle to the rest of the cell and position the spindle
- Kinetochore microtubules - bind to other kinetochores, measures the distance between two centrosomes
- Interpolar microtubules
The kinetochore
Kinetochore = set of proteins that link the centromere to the kinetochore MTs
- the centromere binds the microtubules
/!\ centromere ≠ centrosome!
What moves chromosomes?
1) Regulation of MT polymerisation/ depolymerisation
Dynamic instability of MTs is used during mitosis
- depolymerisation of MTs
Ndc80 to the rescue
- Ndc80 is a complex of proteins
- Bridges MTs and kinetochore
Regulation of MAPs that regulate MT
E.g. stathmin is inhibited during mitosis
Cdk1 phosphorylates stathmin and inhibits it in mitosis
Stathmin normally binds tubulin and inhibits MT polymerisation.
Its phosphorylation inhibits its interaction with tubulin Phospho‐mutants mimic a phospho. or dephospho.
What moves chromosomes?
2) Pooling and pushing forces‐ MTs motors
Incorrect attachment or segregation of chromosomes can result in aneuploidy
Amphitelic, Monotelic
Failure to segregate chromosomes of parental gametes at meioisis
The Spindle Assembly Checkpoint (SAC)
Cytokinesis
- process that divides the cell in 2 halves
- begins in anaphase, ends in telophase
- in animal cells, cleavage furrow on the cell surface
- uses actin and MTs
MTs control the position and orientation of the division plane
o Some cells divide in their middle, other must divide asymmetrically
o Mitotic spindle is moved and defines site + orientation of cleavage
- 3 proposed models
- symmetrical vs. assymetrical division determines cell fate
Actin and myosin orchestrate the assembly and constriction of the contractile ring
- Contractile ring = actin + myosin
- Work together like in skeletal muscles cell membrane
- Actin‐binding partners though to be deposited by MTs at site of ring formation, to attract and regulate actin
