the cell cycle and its control Flashcards
what are the 3 things that the cell cycle ensures
-chromosome replication
-chromosome segregation
-cell division
The Cell Cycle as a Clock
-Almost all normal cells do NOT proliferate unless stimulated by extrinsic factors
-Other signalling proteins can overrule these stimulatory factors and force a halt to proliferation
-Extracellular signals can induce a post-mitotic, differentiated state = no proliferation
-A mixture of signals has to be integrated by the cell to make the decision to proliferate, be quiescent of differentiate
-existence of a master governor that makes major decision regarding cell fate = cell cycle clock, which operates in the nucleus
whats the structure of the cell cycle
-Prophase: Movement of the centrioles to polar end of the nucleus; condensation of chromosomes
-Prometaphase: Components of the mitotic spindle elongate away from the spindle poles
-Metaphase: Chromosome alignment is complete – pairs of chromosomes are aligned
-Anaphase: Pairs of sister chromatids are separated
-Telophase: Chromosome start to de-condense, nuclear membrane starts to be re-established
-all Passes except M phase we call interphase
why is interphase the main part of the cel cycle
1) G1 phase: cell increases in size, ribosomes, RNA produced and preparation for DNA synthesis
2)S phase: DNA synthesized (chromosomes duplicated)
3) G2 phase: cell checks fidelity of DNA, and preparation for nuclear division
what is the G1 phase
-where cells grow and perform their physiological function. Transit through G1 is driven by external signals (growth factors etc).
-Cells can exit G1 into G0, where they will not-divide and not grow. They can re-enter the cycle from G0
what are the different checks that occur in the duration of the cell cycle
-check for damaged unduplicated DNA- G2
-check for chromosome attachment - M PHASE
-check of DNA damage- G1
-check for DNA damage or staled replication forks- S
-restriction point: check for favourable environment conditions- G0 + G1
what do the checks in the duration of the cell cycle do
-allow for increase to scheduled length of a phase
-facilitate repair processes
-Are known as checkpoints
G1: growth v/s quiescence decision
-Discrete window to consult the extracellular environment: from the onset of G1 phase to an hour or 2 before the G1-to-S transition.
-G1 decision making machinery apparent in the responses of cultured cells to extracellular signals:
=Serum and growth factors removed before the cells have completed 80-90% of G1 ->fail to proceed further and revert to G0 state
=Serum and growth factors removed in the final hr of G1 -> proceed to S, G2 and M phase
-Deregulation of the R-point decision-making machinery accompanies the formation of most types of cancer cells
what are The challenges associated with finding a useful model system
-Genetic approach: Requires cells that have a mutation in a putative cell cycle transition gene
-Biochemical approach: Requires supply of large numbers of cells undertaking the same transition at the same time
advantages of Yeast as a genetic model for cell cycle
-Rapid division rate <1hr
-Cell cycle control genes are HIGHLY conserved
-Yeast can be grown as haploids or diploids
-Easy to grow
-You can tell which phase of the cell cycle the yeast cell is in by just looking. (the length of the cell)
How can we study genes that are crucial for cell survival?
-Genetic tricks allow identification of potentially lethal mutations:
-Diploids can be used to maintain lethal mutations that are then studied as haploids
-Temperature sensitive mutations allow growth at permissive temperature
whats Cell-free mitosis
-One can deplete the cytoplasm of different proteins using antibodies
-One can remove cytoplasm at different stages to study changes (eg in protein phosphorylation) over time
-they are testing proteins- get proteins, inactivate them, test nuclei to see if it undergoes mitosis
biochemical approach: what do they all early embryonic cells to do
-to purify cell cycle regulators
-Something in the Egg cytoplasm can catalyse the transition from G2 to M-phase : The factor was called Maturation Promoting Factor (MPF)
biochemical approach: How do we isolate the proteins that are responsible for cell cycle transitions?
-Frogs eggs grow arrested in G2 to a relatively large size (good for extracting decent amounts of protein)
-Oocytes arrested in G2, then under hormonal control mature and start meiosis BUT when they are laid they become stuck in meiotic metaphase ie M-phase. The are released by fertilization -> to start dividing (without growing)
-So what controls this transition from G2 to M
whats G2/M transition
-You can start to take the egg cytoplasm, separate out the proteins into different classes eg by size, etc
-Inject back into an oocyte and see if it triggers the G2 to M transition
-then re-purrify (lane to the right)
-Eventually you get some consistent bands that trigger the G2/M transition
-These bands are similar to the yeast genes identified
-This identified Cyclin-dependent kinases (CDKs) and Cyclins
G2/M transition: bioinformatics
-A cell cycle transition is controlled by a protein kinase-based machine (AA sequence of CDK)
-These were similar to proteins identified in yeast which perform similar functions
what are protein kinases
-signalling devices which operate to create molecular switches
how do we visualise and quantify kinase activity
-Selective extraction of kinase
-Incubation with a protein substrate and ATP
-Electrophoresis of substrate and imaging
what do Cell cycle transitions involve
-irreversible destruction of cyclins
what happens in other cell cycle transitions?
-We would expect that DISTINCT proteins would need to be switched on (or off) in each transition
-So are different transitions catalyzed by distinct regulator kinases?
-In yeast, there is only ONE cell cycle regulator kinase gene, called Cdk1
-however there are multiple cyclins
what are there multiple of cyclins in mammals
-Cdks
-cyclins
how do Cyclin levels fluctuate during the cell cycle
-Cyclin E: low levels throughout most of G1, rapid increase after the R point
-Cyclin A: levels increase in concert with the entrance in S phase
-Cyclin B: levels increase in anticipation of mitosis
-Collapse of cyclin levels as the cell progresses through the cell cycle -> degradation (ubiquitination-dependent)
-cell cycle can now only progress in one direction
what are the two major control factors
(1)Cyclins/CDKs– this drives the cycle forward
(2) Mechanisms to stop the cycle (and correct) if there are problems ie Checkpoints
what are D- type cyclins controlled by
-extracellular signals
what are cyclin/ CDKs regulated by
-CDK inhibitors
