7. External factors controlling division and behaviour of normal and cancerous cells Flashcards
1
Q
What is cell behaviour?
A
The term used to describe the ways in which cells interact with their external environment and their reaction to this, particularly proliferative and motile responses of cells.
2
Q
What external influences are detected by cells?
A
- Chemical - hormones, growth factors, ion concentrations, ECM, molecules on other cells, nutrients and dissolved gas concentrations
- Physical - mechanical stresses, temperature, the topography or ‘layout’ of the ECM and other cells.
3
Q
What external factors can influence cell division?
A
All external factors can potentially influence cell division but the ones ones that are best understood are:
- Growth Factor
- Cell-Cell adhesion
- Cell-ECM adhesion
4
Q
Describe the basic behaviour of cells in culture.
A
- Initially, you get an isolated cell on the culture medium, which will then settle down on the culture surface due to gravity
- It will then spread across the culture medium and it will usually obtain some polarity - it will have a front and a back
- The front is usually the motile part
- This is not a passive process
- Energy is required to modulate cell adhesion and the cytoskeletion during spreading
5
Q
Why is cell-ECM Adhesion important?
A
- An experiment was conducted and the degree to which cells proliferated on adhesive surfaces of different sizes was observed
- If the cells were suspended in agar (non-adhesive) few cells entered the S phase
- If they were able to stick to a small adhesive patch that did NOT allow them to spread fully, a small proportion of cells proliferated
- If the cells were allowed to stick to a larger adhesive patch, which allowed them to spread out fully - almost all the cells started proliferating
- Throughout this experiment, growth factors were available to the cells so this variable did not affect the proliferation of the cells
- The ability of the cells to respond to the growth factors required the cells to be adhered and spread
- So natural cell-ECM adhesions are required for proliferation
6
Q
Explain the importance of cell spreading.
A
- Most cell stick to fibronectin (matrix molecule that is also found in the blood)
- An experiment was conducted and spreading and survival of cells was observed.
- A cell was placed on a defined small patch of fibronectin, the cell stuck but could not spread and died from apoptosis
- The same amount of fibronectin was distributed over a number of small spots, the cells were able to spread and survive
- So it’s not just adhesion that is required for the cells to proliferate, the cells also need to be able to spread to enable the cells to respond to growth factors and proliferate
7
Q
Define Anchorage Dependence.
A
The requirement of attachment to ECM for survival.
8
Q
There is mechanical continuity between the ECM and the cell interior. What does this mean?
A
- Cells have receptors on their cell surface, which binds specifically to ECM molecules
- These molecules are often linked, at their cytoplasmic domains, to the cytoskeleton
9
Q
Explain the importance of integrins in the ECM.
A
- Integrins are the most important of the matrix receptors
- Integrins are heterodimer complexes consisting of alpha and beta subunits
- They bind to the ECM via their heads
- Each of their tail regions cross the plasma membrane and project into the cell
- There are about 10 alpha and 8 beta subunits that form more than 20 known combinations
- Each combination specifically binds a short, specific peptide sequence(e.g. alpha-5 beta-1 fibronectin receptor binds arg-gly-asp (RGD)
- Intracellularly, the integrins are linked, via actin-binding proteins, to the actin cytoskeleton (most integrins do this)
- An exception to this is the alpha6 beta4 integrin complex found in epithelial hemidesmosomes- these are linked the cytokeratin (intermediate filament) cytoskeleton
- Integrin complexes cluster to form local adhesions (most) or hemidesmosomes (alpha-6 beta-4)
- These clusters are often involved in SIGNAL TRANSDUCTION- the integrins are not just an adhesive patch for adhesion of cells, it is also a platform for signalling
- This dual function of integrins allows the cells to interpret the matrix composition of the environment
- Many integrins are also designed to bind to specific adhesion molecules on other cells
- This is particularly important in the immune system and blood clotting
10
Q
A
- ECM receptors (e.g. integrins) can act to transduce signals in both directions
- E.g. ECM binding to an integrin complex can stimulate the complex to produce a signal inside the cell
- The signals can be from ‘outside-in’
- The signals can be from the ‘inside-out’
- A signal generated inside the cell (e.g. as a result of the hormone binding to the receptor) can act on an integrin complex to alter the affinity of an integrin (i.e. alter its affinity for ECM binding) - inside-out integrin signalling
- Integrin complexes could be folded over, in which case they have a low affinity for matrix molecules so they don’t stick particularly well
- There are signals generated within the cell that makes them unfold and go into a high affinity conformation and become sticky
- This is important in the immune system and blood clotting
- Platelets have integrins on their surface but they are inactive - this is good because you don’t want them to stick to everything
- When we do need to activate the platelets, inside-out signals will activate the integrins so they become high affinity and start to stick
- Outside-in signalling:
- A cell can receive information about its surroundings from its adhesion to ECM
- E.g. the composition of ECM will determine which integrin complexes bind and which signals it receives
- This can alter the phenotype of the cells
11
Q
Describe the conformational changes to the integrins.
A
- The integrin on the left is in the low affinity state
- They can be switched on, into the high affinity conformation, by an inside-out signal (this can be due to an external factor such as a growth factor or hormone)
- Once they bind to the matrix, you get other changes taking place
- The 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 the integrins cluster, the molecules act on one another and you get signalling
12
Q
What happens to cells in high density cultures?
A
- For most normal cell types, when you grow them in culture they require a surface to stick to
- As the cells become densely packed, the rate of proliferation starts to slow down
- When they fill up the space provided, they tend to stop division or it becomes very minimal
- It used to be thought that this cessation of proliferation was due to the cells running out of space
- Then an experiment was conducted where a fresh medium containing all the necessary factors for growth was spread across part of the culture surface and the cells that were exposed to these extra growth factors continued to proliferate
- This showed that it wasn’t the contact with neighbouring cells that was preventing cell division, it was actually the availability of growth factors
13
Q
Define density-dependence of cell division.
A
This means that if cell density is too high, there will not be enough growth factor available for cell division.
14
Q
Explain how signals control proliferation of tissues.
A
- Growth factors trigger the ERK cascade and cause cell division
- There is cross-talk between ECM and growth factor signalling
- Signalling from growth factors and signalling from ECM come together to produce proliferation
- So there are 2 signals that interact to produce proliferation:
- Growth factor (density dependence)
- ECM (anchorage dependence)
- So growth factor receptors and integrin signalling complexes can each activate identical signalling pathways (e.g. MAPK)
- Individually, this activation is weak and/or transient
- Together, this activation is strong and sustained
- These separate pathways act synergistically (together)
15
Q
Describe the junctions between cells.
A
- Short-term: transient interactions between cells that do NOT form stable cell-cell junctions
- Long-term: stable interactions resulting in the formation of stable cell-cell junctions
18
Q
Describe cell-cell junctions in epithelia.
A
20
Q
How does cell adhesion affect proliferation?
A
- An experient conducted used cells in culture at normal levels of calcium (physiological levels) and in low calcium (enough to survive but low)
- The reason for removing the calcium is that many of the cell-cell junctions are calcium-dependent
- When the calcium was removed, the cells were still at high densities (and they were force to touch one another) - but were not forming junctions –> the junctions broke down
- When the junctions were absent:
- MAPK much more activated
- Decreased level of p27KIP1 (inhibitor of proliferation in the cell cycle)
- HIGH proliferation
- When calcium was reintroduced and the junctions reformed:
- MAPK inactivated
- Increased level of p27KIP1
- Low proliferation
21
Q
Describe the molecular organisation of adherens junctions.
A
- These are sort of like the master junctions of cells - they control the formation of other types of junctions
- It consists of a Cadherin (calcium dependent, homophilic cell adhesion molecule) which binds to similar molecules on the adjacent cell
- Intracellularly it binds to beta-catenin, which is associated with alpha-catenin, which, in turn, links to the actin cytoskeleton
- beta-catenin was found to be much more important than just being a molecule linking adherens junctions to the actin cytoskeleton
22
Q
Define Adenomatous Polyposis Coli (APC).
A
- APC is an inherited form of colon cancer - there are a number of familial forms
- The APC gene-product is a protein that is involved in the degradation of beta-catenin
26
Q
Explain the consequence of loss of contact inhibition in cells.
A
- Normal cells usually form a nice monolayer
- Cancerous cell lose contact inhibition of locomotion and they will pile up on each other
31
Q
Why is Ras an important proto.oncogene? Name other proto-oncogenes and their corresponding oncogene.
A
Ras is a particularly important oncogene - it is mutated in 30% of all cancers
32
Q
Why is anchorage dependence not important in cancer?
A
- In cancer you have usually lost anchorage dependence (ECM binding) and density dependence (growth factor)
- A normal cell would not be able to proliferate if it loses both these signals
- However, a cancer cell doesn’t require either of these signals because one of the signals that is further downstream, has been permanently turned on
- The pathways has been ‘short-circuited’
33
Q
How does a primary carcinoma cell metastasise?
A
