Lecture 1&2 - The cell cycle and cell cycle control Flashcards
What are the four phases of the cell cycle and what happens in each?
G1: Cell grows and carries out normal metabolism, cell receives external signals to enter the cell cycle and the organelles duplicate
S: Dna replication and synthesis
G2: Cell grows and prepares for mitosis, resolves problems
M: Mitosis and Cytokinesis
What is Quiescence?
When a cell is in an inactive state and not actively dividing, a cycling of G1 (G0)
- can last for years
- must receive signals to exit G0
How did Walther Flemming illustrate the major stages of the cell cycle?
Stained cells genetic material
-Observed that in each cell cycle the interphase chromatin is condensed into transmissible chromosomes then decondensed
What are the major componants of DNA condensation?
- Nucleosomes (DNA wrapped around histone protein cores)
- Solenoid (Winding of the nucleosomes on top of each other)
- Mitotic chromosome scaffold
What occurs in the first stage of DNA condensation? (Nucleosome)
- 146bp length of DNA is wrapped around an octamer of histone subunits, made up of pairs of: H2A, H2B, H3, H4, forming a Nucleosome
- H1 tightens the connections between DNA wrapped around the histone octamer
- Creates a ‘beads on a string’ structure
- results in a 6-fold reduction in length
What is the function of the H1 molecule in the nucleosome?
Tightens the connections between DNA wrapped around the octamer by binding to linker DNA 20-80bp long
-stabilises chromatin structure
What occurs in the second stage of DNA condensation? (Solenoid)
Nucleosomes wind up on top of each others, forming chromatin fibres 30nm thick = Solenoid
-results in a 40 fold reduction in length
What occurs in the third stage of DNA condensation?
After coiling and supercoiling, chromosome fibres attach to the mitotic chromosome scaffold
- results into 46 chromosomes with total length ~200µm
- 10,000 fold reduction in length
For what percentage of time is a cell in M phase?
5%
What is the structure of microtubules in Interphase and the processes in the 5 stages of Mitosis?
Interphase: Microtubules are long and diffuse, not attached to centromere
Prophase: Chromosomes condense
Prometaphase: Nuclear envelope breaks down
Metaphase: Chromosomes align at the spindle equator
Anaphase: Sister chromatids separate along the spindle as the astral microtubules grow
Telophase: Cleavage furrow constricts around the central microtubules and central spindle as nuclei reassemble
What is the cleavage furrow?
An indentation of a cells surface which begins the progression of cleavage leading to cytokinesis
What are the three types of astrotubules formed when the spindle apparatus forms, and what are they?
- Kinetochore microtubules: attaches to kinetochores of condensed chromosomes and moves towards centrosomes
- Polar microtubules: emanates from 2 centrosomes and overlap with each other. Push against each other to force centrosomes towards cell plates
- Astral microtubules: extend from centrosome to cell membrane and anchor to the periphery of the cell. Pull centrosome towards one of the poles
What is the kinetochore?
Sequences of DNA in the centromere where microtubules bind
What experimental benefits can be gained from the natural syncrony in fly embryos?
can study many cells at the same stage at the same time
Give two examples of chromosome segregation gone wrong
- Anaphase bridges: when telomeres of sister chromatids fuse together and fail to completely segregate into their respective daughter cells
- Aneuploidy: inaccurate segregation of chromosomes
What is the chromosome complement at each of the four stages of the cell cycle?
G1: 2N
S: 2N/4N
G2: 4N
gametes: N
How can multiple cell cycle synchrony be achieved?
Chemical inhibition
-blocks cell cycle before mitosis or before S phase leading to aggregation of cells at a particular stage in the cell cycle
-check by flow cytometry - measures the DNA content of a cell (SS, labelling, fluorescent peak) Normally smallest amount of light scattered in G1, smear for S, largest in G2
Natural synchrony - use process of quiescence to synchronise
What are the chemicals that can be used to attain multiple cell cycle synchrony?
Nacodazole (G2) Cytochalasin B Thymidine (G1) Aphidicolin Flow cytometry analysis - Nacodazole shows biggest fluorescent peak for larger cells (G2); Thymidine shows biggest fluorescent peak for smaller cells (G1)
What are the benefits of Natural synchrony over chemical inhibition when achieving experimental cell cycle synchrony?
- adding chemical may prevent validity
- more natural and can stimulate cells with a chemical signal to enter cell cycle at the same time
What are five classic experiments that identified the cell cycle?
Cell fusion experiments - Roa and Johnson Discovery of cyclins - Hunt Discovery of MPF - Lokha and Masui Discovery of cell cycle genes - Hartwell Discovery of Cdc2 - Nurse
Describe the Cell Fusion Experiments done by Roa and Johnson
Aim - (1)discover what happens when take a cell in one stage of cell cycle and fuse with one in another; (2)what happens when cells in cell cycle phases are exposed to different diffusible factors
(1) combined:
Cells in S + G1 => all cells were in S phase. Hypothesised that it was by an S phase specific factor (SPF)
Cells in S + G2 => No change. Therefore, SPF factor not recognised by G2 nucleus
found there was a difference in G1 cells being able to respond and G2 not.
(2)MPF (mitosis promoting factor)
Combined G1 and G2 cells => neither produce any dominant chemical signal
M phase cells fusing with any other cell in a different phase the M phase is dominant, acts on all cells in any phase
Consequently M phase cells produce MPF
What is MPF?
Mitosis promoting factor or Maturation promoting factor: diffusible factor that triggers mitosis
Why use Xenopus eggs as an experimental organism?
- rapid cleavage divisions
- lay hundreds of eggs at the same time
- fertilisation and maturation is external and visable
- only get one gamete, all the nutrients go to one cell resulting in a robust cell with high volume of enzymes and nutrients
What is the process of Oocyte maturation in frogs that Masui observed, and what did he conclude?
Oocyte is arrested at G2 phase, where an influx of progesterone stimulate progression into Meiosis I followed by meiotic interphase. The egg is released and the cell cycle is arrested again at metaphase meiosis II. Fertilisation then occurs and the cell cycle continues and undergoes the first cleavage. (post fertilisation colours segregate to show first division)
Concluded - there are 2 key points of regulation in meiotic cell cycle