External factors controlling division and behaviour of normal and cancerous cells Flashcards
What is cell behaviour?
The ways in which cells interact with their external environment and their reaction to this, particularly proliferative and motile responses of cells
What chemical external influences are detected by cells?
Hormones Growth factors Ion concentrations ECM Molecules on other cells Nutrients Dissolved gas concentrations
What physical external influences are detected by cells?
Mechanical stresses
Temperature
Topography
Layout of the ECM and other cells
What external factors can influence cell division?
All external factors can potentially influence cell division but ones that are best understood are:
Growth factor
Cell-cell adhesion
Cell-ECM adhesion
What will happen if you place an isolated cell on a culture medium?
It will settle down on the surface due to gravity and then spread across the culture medium and usually obtain some polarity- front and back then it will become motile (front is usually the motile part)
Is the cell spreading a passive process?
No as energy is required to modulate cell adhesion and the cytoskeleton during spreading
When cells were suspended in agar, what percentage entered the S phase?
8%
When cells were put on a small adhesive patch that didn’t allow them to spread out fully, what percentage of cells entered the S phase?
30%
When cells were allowed to stick to a large adhesive patch which allowed them to spread out fully, what percentage of cells entered the S phase?
90%
What do cells require to be able to respond to growth factors?
To be adhered and spread- natural cell-ECM adhesions are required for proliferation
What interaction do most cells have with fibronectin?
They will stick to it
If you have a defined small patch of fibronectin and put a cell on it, what will happen?
The cell will stick but it can’t spread so will die of apoptosis
If you take the same amount of fibronectin and distribute it over a number of small spots and put a cell on it, what will happen?
The cell will be able to spread, survive and grow
Why do cells require to be attached to ECM (and a degree of spreading)?
To begin protein synthesis and proliferation- it can be required for survival (epithelia and endothelia)
What is anchorage dependance?
Requirement of attachment to ECM for survival
How is their mechanical continuity between the ECM and the cell interior?
Cells have receptors on their surface which bind specifically to ECM molecules and these are often linked at their cytoplasmic domains to the cytoskeleton
Which is the most important of matrix receptors?
Integrins
What are integrins?
Heterodimer complexes consisting of alpha and beta subunits
How do integrins bind to the ECM?
Via their heads
What do each of the tail regions of integrins do?
They cross the plasma membrane and project into the cell
How many alpha and beta subunits are there in integrins?
10 alpha
8 beta
More than 20 known combinations
What does each integrin subunit combination bind to?
Short specific peptide sequence
How are integrins linked to the actin cytoskeleton intracellularly?
Via actin-binding proteins
What is an exception to integrins linking to actin cytoskeleton via actin-binding proteins?
Alpha6 beta4 intern complex found in epithelial hemidesmosomes- linked to cytokeratin (intermediate filament) cytoskeleton
What do integrin complexes cluster to form?
Local adhesions (most) or hemidesmosomes (alpha-6beta-4)
What are the integrin clusters involved in?
Signal transduction
What does the duel function of integrins allow?
Cells to interpret the matrix composition of the environment
Many integrins are also able to bind to specific adhesion molecules on other cells, what is this particularly important in?
Immune system and blood clotting
How promiscuous are integrins?
Some bind to more than once ECM molecule (sloots)
Some only bind to a single ECM molecule
What are the two ways that ECM receptors can transmit signals?
From outside-in
From inside-out
What is inside-out signalling?
A signal generated inside the cell (e.g. due to hormone binding to receptor) can act on an integrin complex to alter the affinity of an integrin (e.g. for ECM)
Give a common example of inside-out signalling?
Platelets have integrins on their surface which are inactive as you don’t want them to stick all the time but when needed to activate, inside out signals will activate the integrins to become high affinity and start to stick
What is outside-in signalling?
Cells can receive information about its surroundings from its adhesion to ECM - can alter phenotype of cells
How are integrins switched from low affinity state to high affinity?
Inside-out signal- due to an external factor such as growth factor or hormone
What does ligand binding cause an integrin to do?
Ligand binding also causes a change in conformation, the legs separate and cytoplasmic signalling molecules can then bind and that binding will then allow signalling to take place- this is outside-in
When mammillary epithelium was cultured in a medium containing type 1 collagen, what happened?
The cells formed balls of cells that were clumpy and loosely associated with each other
When mammillary epithelium was cultured in a medium containing basement membrane proteins (laminin etc), what happened?
Cells organise themselves into organoids that had a polarised epithelium, there was an empty space in the middle and the cells had polarity- the cells were well differentiated and even produced milk proteins
What does the difference between mamillary epithelium growth in type I collagen and basement membrane proteins show?
ECM has a profound effect on the phenotype of the cells
As cells become more densely packed, what happens to the rate of proliferation?
Slows down
What was thought to be the cause of the decreasing cell proliferation in response to increasing cell density?
Cells running out of space
What is the cause of the decreasing cell proliferation in response to increasing cell density?
The availability of growth factors- density is too high so wasn’t enough growth factor available for cell division- density-dependence of cell division
What do growth factors trigger?
ERK cascade and cause cell division
What are the two signals that interact to produce proliferation?
Growth factor (density dependence)
ECM (anchorage dependence)
Individually- activation is weak and transient
Together- activation is strong and sustained
What are short-term contact interactions between cells?
Transient interactions between cells that don’t form stable cell-cell contacts
What are long-term contact interactions between cells?
Stable interactions resulting in formation of stable cell-cell junctions
What happens when most non-epithelial cells collide?
They don’t form stable cell-cell contacts as they don’t like touching each other and they actually repel one another
How do non-epithelial cells repel each other when they collide?
Contact inhibition of locomotion- Paralysing motility at the contact site, this promotes the formation of a motile front at another site of the cell so the cell moves away in the opposite direction
What is contact inhibition of locomotion responsible for?
Preventing multi-layering of cells in culture and in vivo
Name some types of long term cell-cell contacts?
Adherens junctions, desmosomes, tight junctions and gap junctions
What sort of cells always strongly adhere together and form specific cell-cell junctions?
Epithelial and endothelial cells- form continuous layers
Neurones forming synapses and myocardial tissue as well
What are the two main types of cell-cell junctions found in epithelia?
Zonula (belts)
Macula (discrete spots)
What happens when epithelial cells make contact?
Contact induced spreading of epithelial cells- mutual induction of spreading which would form a stable junction between cells
As an investigation into the effect of cell-cell contact in epithelia on cell proliferation, what was the difference between a cell culture with normal levels of calcium and low calcium?
Normal calcium:
MAPK inactivated
Increased level of p27KIP1
Low proliferation
Low calcium- no junctions form as junctions are dependent on calcium:
MAPK activated
Decreased level of p27KIP1
High proliferation
What happened when this experiment was repeated with adhesion blocking antibodies instead of calcium?
Same effect
Why are adherens junctions like the master junctions of cells?
They control the formation of other types of junctions
What does an adherens junction consist of?
Cadherin which binds to similar molecules on adjacent cell
What is cadherin?
Ca2+ dependent, homophilic cell adhesion molecule
What does cadherin bind to intracellularly?
Beta-catenin which is associated with alpha-catenin which in turn links to the actin cytoskeleton
What is adenomatous polyposis coli (APC)?
Inherited form of colon cancer- number of familial forms
What is the APC gene product involved in?
It is a protein involved in degradation of beta-catenin in the cytoplasm
Normally any free beta catenin in the cytoplasm is rapidly degraded but what happens if it accumulates in the cytoplasm?
It can associate with LEF-1 to form a complex that acts as a transcription factor
What does the beta-catenin/LEF-1 complex do after being formed?
It goes into the nucleus and influences gene expression and proliferation
What happens in adenomatous polyposis coli to lead to increased proliferation?
APC mutation reduces efficiency of degradation of beta-catenin as APC isn’t functioning properly- this leads to accumulation of beta-catenin in cytoplasm which associates with LEF-1 and increases proliferation
What happens when cadherins are clustered together?
You get changes in the activation of some of the small GTPases including Rac and Rho- this can influence proliferation
You also get association of many growth factor receptors into cell-cell contacts and this can prevent their activation
Under certain conditions, cells can lose their social skills and behavioural restraints, what happens as a result?
Proliferate uncontrollably (lose density dependence)
Less adherent and will multilayer (lose contact inhibition of locomotion and anchorage dependence)
Epithelia break down cell-cell contacts
Not hayflick limited- express telomerase and become immortal
What is a consequence of the loss of contact inhibition of locomotion for the progression of cancer?
It allows invasion of surrounding tissue
What is a proto-oncogene?
If a gene coding for a component of a signalling pathway is mutated so protein is constitutively active- pathways will be permanently turned on (Ras)
e.g. Receptors, signalling intermediates and signalling targets
How would you describe the way in which protooncogenes lead to cancer?
A mechanism of short-circuiting leading to uncontrolled proliferation as a result of loss of growth factor dependence
What is the definition of an oncogene?
Mutant gene which promotes uncontrolled cell proliferation
What is the definition of a photo-oncogene?
Normal cellular gene corresponding to oncogene
Why is Ras a particularly important oncogene?
It is mutated in 30% of all cancers
How can a cancer cell proliferate if it has lost anchorage dependence and density dependence but a normal cell can’?
One of the signals further downstream has been permanently turned on
