The cell cycle and its regulation Flashcards
Describe the reasons why different cells divide at different rates
- Embryonic vs adult cells (early frog embryo cells - 30 min)- embryonic cells divide much faster
- Complexity of system (yeast cells - 1.5 - 3 h)
- Necessity for renewal
(intestinal epithelial cells - ~20 h
hepatocytes - ~1 year)- however hepatocytes can proliferate much faster upon injury (liver lobules can re-grow)- shows how tightly regulated the system is - State of differentiation (some cells never divide -
i. e. neurons and cardiac myocytes) - Tumour cells?- lose ability to control proliferation and differentiation- not regulated
Describe the antagonistic relationship between differentiation and division
When a cell is differentiating- it is not dividing
When a cell is dividing- it is not differentiating
This is why once a cell has differentiated- they never divide
Outline the relevance of appropiate regulation of cell division
Premature, aberrant mitosis results in cell death
In addition to mutations in oncogenes and tumour suppressor genes, most solid tumours are aneuploid (abnormal chromosome number and content).- due to dysregulaiton of cell cycle and mitosis
Various cancer cell lines show chromosome instability (loose and gain whole chromosomes during cell division)
Perturbation of protein levels of cell cycle regulators is found in different tumours - abnormal mitosis
Contact inhibition of growth
Attacking the machinery that regulates chromosome segregation is one of the most successful anti-cancer strategies in clinical use
What is meant by contact inhibition of growth
Cells grow normally by detecting neighbouring cells and know when to stop growing (spatial awareness) when there are enough cells to form a functional organ/tissue.
What is meant by the cell cycle
Orderly sequence of events in which a cell duplicates its contents
and divides in two.
Outline the 3 key events in the cell cycle
Duplication
Division
Co-ordination
Co-ordination is extremely important as if you divide before duplicaitng, the daughter cells will not have enough organelles or DNA to be viable
What is meant by the M-phase of the cell cycle
M-phase: Mitosis (Division)
Nuclear division
Cell division (cytokinesis)
need to divide both the nuclear and cytpolasmic contents
What is meant by interphase
Interphase (Duplication)
DNA
organelles
protein synthesis- to make new organelles
Why is the M-phase the most vulnerable part of the cell cycle
Mitosis - most vulnerable period of cell cycle:
Cells are more easily killed (irradiation, heat shock, chemicals)
DNA damage can not be repaired- so if a chromosome splits unevenly here- it will perpetuate into the daughter cells
Gene transcription silenced
Metabolism reduced
Everything is focussed into dividing the cell
This is why the M-phase is the shortest part of the cell cycle (1 hour)
What happens during the 3 stages of interphase
G1 – the cell makes sure that it has everything that is necessary for duplication
S – DNA replication, protein synthesis and replication of organelles
G2 – the cell checks that everything is ready to enter mitosis and that the DNA has been replicated without any errors (any errors are repaired here)
What is Go
G0 - cell cycle machinery
dismantled
Cell has left the cell cycle
Most cells are here
This is where they perform their function (regeneration, secretion, motility etc)
What happens during the S phase of the cell cycle
DNA replication
Protein synthesis: initiation of translation and elongation increased; capacity is also increased (increased ribosomes)
Replication of organelles (centrosomes, mitochondria, Golgi, etc)
in case of mitochondria, needs to coordinate with replication of mitochondrial DNA- need to replicate mtDNA too
Describe the structure of a centrosome
Consists of two centrioles ( mother and daugher centrioles)
Each centriole has barrels of nine triplet microtubules
The mother and daughter centrioles are held perpendicuarly to each other by inter-connecting fibres and a cloud of electro-dense material .
What is the function of a centrosome
Centrosomes are organelles near the nucleus of a cell, which contain centrioles (mother and daughter), and from which spindle fibres develop in cell division (to form the highways for chromosomal segregation)
Describe the life cycle of centrosomes during mitosis
The mother and daughter centrioles separate in G1
Then the mother produces another daughter and the daughter produces another mother, resulting in the formation of 2 centrosomes (the duplication takes place during S phase)
One centrosome will go to each daughter cell
Describe the role of the centrosomes during the M-pahse
Nucleation sites appear (gamma-tubulin ring complexes)
NOTE: nucleation is the assembly and polymerisation of microtubules which are needed for chromosomal segregation
List the different stages of mitosis
Prophase à prometaphase à metaphase à anaphase à telophase à cytokinesis.
Describe the condensation of chromatin during prophase and its importance
Short region of DNA double helix- 2nm
The double helices wrap around histones to form beads-on-a-string (11nm)
The beads then fold in on themselves to compact furher to form the 30nm fibre of packed nucleosome (each bead is a nucleosome)
It is then compacted to form a chromosome scaffold (300nm) and then further wrapped (700nm) until you get a chromosome (1400 nm)
Important as condensed chromatin is less likely to break off, easier to segregate and transport around the cell.
Describe the condensed chromosomes in prophase
Condensed chromosomes - each consists of 2 sister chromatids, each with a kinetochore attached to the centromere
What is a kinetochore
Protein complexes that are attached to each sister chromatid – they are important in detecting the attachment of microtubules in chromosomal segregation
What can be seen at the start of prophase
Heterogenous nucleus (due to condensation of chromosomes) Centrosome and formation of microtubles outside the nucleus
What can be seen by the end of late prophase
Replicated chromosomes condense
Duplicated centrosomes migrate to opposite sides of the nucleus and organize the assembly of spindle microtubules
Mitotic spindle forms outside nucleus between the 2 centrosomes
Describe spindle formation
Radial microtubule arrays (ASTERS) form around each centrosome (microtubule organizing centers - MTOC)
Radial arrays meet (from each centrosome)
Polar microtubules form (the ASTERS change properties)
N.B. Microtubules are in a DYNAMIC state- where the polar microtubules are constantly polymerising and depolymerising- so that there length and shape will continually be changed
What is the key feature of metaphase
Chromosomes aligned at equator of the spindle.
What are the two phases or prometaphase
Prometaphase: early prometaphase
late prometaphase
What happens during early prometaphase
Breakdown of nuclear membrane (important as we need to seperate the nuclear material in mitosis)
Spindle formation largely complete
Attachment of chromosomes to
spindle via kinetochores (centromere region of chromosome)- chromosomes captured by these microtubules (via kinetochores) once the nuclear membrane is broken down.
What happens during late prometaphase
Microtubule from opposite pole is captured by sister kinetochore
Chromosomes attached to each pole congress to the middle
Chromosome slides rapidly towards center along microtubules
What is CENP-E
CENP-E = centromere protein E (kinetochore tension sensing
Senses the stretch on the chromosome from the microtubule
Summarise anaphase
Paired chromatids separate to form two daughter chromosomes
Cohesin (multi-protein complex) holds sister chromatids together
Anaphase A and B
What are the 3 main types of half-spindle
Kinetochore microtubule – attached to kinetochores
Polar microtubule – attached to a microtubule array from the other centrosome
Astral microtubule – microtubule originating from a centrosome that does not connect to a kinetochore
What happens during anaphase A
Cohesin is broken down and the microtubules get shorter so the chromatids start moving towards their respective poles
What happens in anaphase B
1-Daughter chromosomes migrate towards poles
2-Spindle poles (centrosomes) migrate apart
These two forces provide considerable segregation of the DNA in the cell- such that are large area is cleared for easy segreagation of the contents
Are all the microtubules moves to each pole in anaphase
No, some remain in the centre- to maintain the highways for chromosomal segregation.
What happens in telophase
Daughter chromatids arrive at the pole and the nuclear envelope reassembles
Assembly of a contractile ring of actin and myosin filaments where the cells are going to split
The contractile ring squeezes the cell so it divides into two daughter cells
NOTE: cleavage furrow = where the cell is going to be cleaved
What is important to remember about telophase
Last phase of mitosis
Describe what happens in cytokinesis
Cytoplasm cleaved in two (by contractile ring)
Chromatin recondenses
Nuclear substructures reform
Interphase microtubule array reassembles.
What is the mid-body
The point where the actin-myosin contractile ring is formed- seen at the end of cytokinesis still- as a tail between the two daughter cells.
Essentially, what is the spindle assembly checkpoint
Senses completion of chromosome alignment and spindle assembly
(monitors kinetochore activity)
Ensures that all the chromosomes are arranged in the middle of the cell and they each chromatid is attached to a mictorubule.
How does the spindle assembly checkpoint monitor chromosomal attachment
The cell makes use of negative signals:unattached chromosomes send a stop signal to the cell-cycle control system.
The kinetochore has proteins that emit a signal when the kinetochore is NOT attached to microtubules
When a microtubule attaches to the kinetochore, it stops emitting the signal
At the end of metaphase, you want all the kinetochores to stop sending signals before you can proceed to anaphase
Name two proteins that allow the kinetochores to detect spindle attachment
attachment.
CENP-E (monitors tension on chromosomes from microtubules).
BUB-protein kinase (this dissociates from the kinetochores when the chromatids are properly attached to the spindle – then they go on to signal progression to anaphase)
Describe the importance of the spindle assembly checkpoint
Active during prometaphase and metaphase
If a dividing cell were to begin to segregate its chromosomes before all the chromosomes were properly attached to the spindle, one daughter cell would recieve an incomplete set of chromosomes, while the other would recieve a surplus. Both situations can be lethal, thus the cell ensures that every last chromosome is attached properly to the spindle before it completes mitosis.
What can cause aneuploidy
Mitotic checkpoint defect
Mis-attachment of the spindles (so chromatids end up at different poles to the ones that they should be at)
Aberrant mitosis (production of an abnormal number of centrosomes leads to abnormal division of the chromatids, and abnormal cytokinesis)
What are the four different types of spindle attachment
Amphitelic – normal spindle attachment
Syntelic – both kinetochores of the sister chromatids are attached to spindles from one centrosome
Merotelic – one kinetochore of one of the sister chromatids is attached to spindles from both centrosomes
Monotelic – one kinetochore of one of the sister chromatids is attached to a spindle, the other is unattached
What is important to remember about syntellic attachment
The kinetochores may or may not be producing checkpoint signals.
What are the consequences of syntellic attachment
Both sister chromatids at the same pole (duplicated chromosome)
What is the consequence of merotellic attachment
Mulitple competition for the same chromatid
Chromatid stays in centre of the cell
Chromosome lost at cytokinesis.
Describe how aberrant centrosome/DNA dupliation can lead to aneuploidy
Aberrant centrosomes:
o DNA and centrosome duplication leading to too many centrosomes (often 4) and a multipolar spindle aberrant cytokinesis.
Broadly speaking, how can anti-cancer therapies work
Anti-cancer therapy by inducing gross chromosome mis-segregations
Describe the use of checkpoint-kinase inhibitors as anti-cancer therapies
Checkpoint kinase (CHKE1 and CHKE2) – Serine threonine kinase
activation holds cells in G2 phase until all is ready
inhibition leads to untimely cell transition to mitosis
These inhibit the the attachment-error mechanism- leading to abnormal segregation of chromosomes in mitosis and thus early apoptotic cell death
Describe the use of taxanes and vinca alkaloids as anti-cancer therapies
Taxanes and vinca alkaloids (breast and ovarian cancers)
Alters microtubule dynamics
Produces unattached kinetochores
Causes long-term mitotic arrest. (as cell cannot progress to anaphase without loss of these signals- don’t want cells to spend ages in mitosis as the cell is vulnerable during this time- so the cell is just killed off.
Why don’t chemotherapies cause as much damage to normal cells
As cancer cells divide faster- so these will be targeted more.
What can go wrong during the cell cycle
e.g. Cell is not big enough or DNA damage
What happens when something goes wrong during the cell cycle
. Cell cycle arrest
at check points (G1 and spindle check point)
can be temporary (i.e. following DNA repair)
- Programmed cell death (apoptosis)
DNA damage too great and cannot be repaired
Chromosomal abnormalities
Toxic agents
Cell cycle progression aborted and cell destroyed
What are the main checkpoints of the cell cycle
During G1 (triggered by growth factors- ensure the cell has everything it needs to be able to duplicate and divide).
Just before mitosis to check for DNA damage
Metaphase-anaphase checkpoint (spindle assembly checkpoint)
Describe the potential impact of tumours on the cell cycle checkpoints.
G1- Increased production of grwoth factors or growth factor receptors- to increase the frequency and speed f the cell cycle
G2- blocks checkpoint so all cells enter mitosis- even if they have unrepaired DNA damage
Spindle assembly- mitosis continues irrespective of whteher the chromosomes are properly alligned or not- this can lead to aneuploidy
These mechanisms are ultimately why tumour cells are so dysregulated.
Describe the de-regulation of the cell cycle during tumorigenesis.
Tumours can prevent the cell from entering Go
and thus prevent the dismantling of the cell cycle apparatus- so the cell continually divides.
What triggers the cell to enter Go
In the absence of stimulus, cells go into Go (quiescent phase)
Most cells in the body which are differentiated to perform specific functions
Cells are not dormant, but are non-dividing
How can cells exit Go
Exit from G0 highly regulated - requires growth factors and intracellular signalling cascades
Summarise signalling cascades
Response to extracellular factors
Signal amplification
Signal integration
Modulation by other pathways (during relay and amplification steps)
Regulation of divergent responses (for example metabolic, gene expression or cytoskeletal changes).
Describe peptide growth factors
Epidermal growth factor (EGF); Platelet-derived growth factor (PDGF)
Respective receptors found as monomeric, inactive state
Receptor Protein Tyrosine Kinase (RPTK)- E.C binding domain and I.C tyrosine kinase domain.
Describe signalling by peptide growth factors
Receptors are monomeric, ligands are dimeric
In the presence of a ligand:
Receptors form dimers
are activated by phosphorylation
Which 3 amino acids can be phosphoylated
Tyrosine
Serine
Threnonine
Describe what happens during protein phosphorylation
transfer of phosphate from ATP to a hydroxyl groups
What are the consequences of protein phosphorylation
The added phosphate group (negatively charged) can alter protein function by:
causing a change in shape (conformation) leading to change in activity (+ve or –ve)
creating a docking site for another protein
What does receptor activation trigger
Different phosphorylation events
kinase cascades ( to different sites other than the tyrosine kinase domain).
binding of adapter proteins (which allows control of divergent pathways)
Summarise the protein kinase cascade
The proteins regulated by the kinases are often other kinases and so activation of one kinase activates another to activate another.
Each kinase also has a corresponding phosphatase to turn the kinase off- allows for regulation.
This leads to:
o Signal amplification.
o Signal diversification.
o Opportunity for regulation.
What are the 3 actions of prophase
o Chromosomes condense.
o Duplicated centrosomes migrate to opposite sides of cell and become MTOC.
o Mitotic spindle forms between 2 centrosomes.
What can a permanent activation of a cyclin lead to
Driving of a cell through a checkpoint.
