External factors controlling division and behaviour of normal and cancerous cells

Question 1

What chemical external influences are detected by cells

Answer 1

Hormones
Growth factors
Ion concs
ECM
Molecules on other cells
Nutrients 
Dissolved gas  (O2/CO2) concs.

Question 2

What physical external influences are detected by cells

Answer 2

Mechanical stresses
Temperature
The topography or “layout” of the ECM and other cells

Question 3

Which external factors influence cell division/proliferation

Answer 3

Growth factors
Cell-cell adhesion
Cell-ECM adhesion

Question 4

Describe the adhesion of tissue cells in culture (cell-ECM adhesion)

Answer 4

In suspension, cells do not significantly synthesise protein or DNA
Cell proliferation
Anchorage dependence

Question 5

How does cell-ECM adhesion influence cell proliferation

Answer 5

Cells must be spread out and bound to ECM in order to be competent in responding to soluble growth factors (for protein/DNA synthesis).
This principle is known as Anchorage dependence.

Question 6

Describe what happens to a cell when it is placed on a culture medium

Answer 6

It will begin to settle and spread across the surface
It will gain some sort of polarity
It will become motile

NOTE: this is an active process, it is not because of gravity. Energy is required to modulate cell adhesion and changes in the cytoskeleton during spreading

Question 7

Describe the cell-ECM adhesion molecules

Answer 7

Receptors on cell surfaces which bind specifically to ECM molecules

These are often linked to the cytoskeleton at their cytoplasmic domains

This ensures mechanical continuity between ECM and the cell interior

Question 8

What are integrins

Answer 8

ECM receptors

Heterodimer complexes of alpha and beta subunits

Question 9

Describe the head and leg regions of integrins

Answer 9

integrins associate extracellularly by their “head” regions.
Ligand-binding occurs at the junction of the head regions

Each of the “leg” regions spans the plasma membrane

Question 10

What do the extracellular parts of integrins bind to and give an example

Answer 10

Short, specific peptide sequences (e.g. arg-gly-asp binding to RGD sequence)

Question 11

What do most integrins bind to intracellularly and how

Answer 11

Actin cytoskeleton via actin-binding proteins at focal adhesions

Question 12

What is an exception to this generalisation of integrin binding?

Answer 12

The alpha6beta4 integrin is found in hemidesmosomes in epithelia and it binds to cytokeratin instead

Question 13

What do integrins form when they cluster?

Answer 13

Most integrins – local adhesions

alpha6beta4 integrin - hemidesmosomes

Question 14

Give examples of which specific adhesion molecules on other cells that integrins can bind to

Answer 14

αvβ3 binds to PECAM-1(CD31) on endothelial cells in inflammation
WBC integrin binds to ICAM on endothelial cells – important in inflammation

Question 15

What is the other important purpose of integrins other than cell adhesion?

Answer 15

It is a platform for signal transduction

Question 16

What are the two signalling actions of ECM receptors

Answer 16

Transduce signals (from outside > inside cell) 
Generate signals (from inside > outside cell)

Question 17

Describe outside-in signalling of integrins

Answer 17

Binding of integrin complexes to ECM can stimulate a signal inside the cell
The ligand binds and opens the legs of the complex, allowing cytoplasmic signalling molecules to bind

Question 18

Describe inside-out signalling of integrins and give an example of when it occurs

Answer 18

Growth factors can generate signals inside the cell, which can act on the integrin complex and alter its affinity
Occurs often in inflammation or blood-clotting - “switching on” of circulating leukocytes

Question 19

What does Inside-out signaling activate

Answer 19

Folded integrin complex within the plasma membrane, acting to extend the complex.

This then allows cytoplasmic signaling molecules to bind, promoting signaling and actin assembly

Question 20

What is proliferation dependent on (for integrin complexes and GF)

Answer 20

Density and Anchorage

GF - density
Integrin/ECM - anchorage

Question 21

Describe how density influences proliferation

Answer 21

Proliferation seems to cease when the density of
growth factors (which are signaling for proliferation) is too great
At high density, cells compete for growth factors = Density-dependence of cell division.

Question 22

What is anchorage dependence

Answer 22

Cells must be spread out and bound to extracellular matrix to respond to soluble growth factors and proliferate

Question 23

When cells are dividing on a culture medium, they will stop dividing once they reach the edges of the medium. What was originally thought to be the reason behind this? and what is the actual reason for this

Answer 23

Contact inhibition of cell division

Competition for external growth factors (density-dependence)

Question 24

Describe the ERK MAP kinase cascade

Answer 24

Chain of proteins in the cell that communicates a signal from a receptor on the surface of the cell to the DNA in the nucleus of the cell
The pathway includes many proteins, including MAPK (originally called ERK), which communicate by adding phosphate groups to a neighboring protein, which acts as an “on” or “off” switch.

Question 25

Describe the stimulation of proliferation by the ERK MAP kinase cascade

Answer 25

Growth factor receptors and integrin complexes can both activate the same signalling pathways (like MAPK).
Individually GFs or integrin signaling complexes will only result in weak/transient activation of the pathway
Together, their activation of the cascade is strong and sustained, leading to more profound proliferation

Question 26

What are the two types of contact interactions between cells

Answer 26

Short-term - transient interactions between non-epithelial cells which do not form cell-cell junctions

Long term - stable interactions resulting in formation of cell-cell junctions

Question 27

What happens to non-epithelial cells when they come into contact with one another?

Answer 27

repel one another by paralysing motility at the contact site
Promoting the formation of a motile front at another site on the cell
Causes it to move off in the
opposite direction
This is contact inhibition of locomotion and is partly responsible for preventing the multilayering of cells

Question 28

What are some long-term cell-cell contacts and what are the junctions in the epithelia arranged as

Answer 28

Adherens junctions
Desmosomes
Tight junctions and gap junctions

Continuous belts (zonula) or discrete spots
(macula)

Question 29

What happens when epithelial cells become in contact with one-another

Answer 29

mutual spreading, extending the area of contact and helping to lead to a more stable layer

Question 30

What effect does high calcium levels have on an epithelium?

Answer 30

Formation of cell-cell junctions
MAP Kinase pathway (inactive) - increase p27kIP1
Results in low proliferation

Question 31

What effects do antibodies blocking adherens junctions have on an epithelium?

Answer 31

Formation of cell-cell junctions will not occur in high levels of ABA (high proliferation)

Question 32

What are adherens junctions mediated by and what does it associate with intracellularly

Answer 32

Cadherins
transmembrane (Ca2+ dependent) cell adhesion molecule
Associates with β-catenin intracellularly.

Question 33

Describe adherens junctions structure

Answer 33

β-catenin associates with an α-catenin molecule, which associates with an actin filament.

Question 34

Give an example of a pathology that involves cadherins

Answer 34

Adenomatous polyposis coli (APC), a form of hereditary colon cancer
The affected protein is involved in degrading β-catenin (cell-cell junctions)
Results in an overgrowth of the colon epithelium and thousands of polyps (proliferation)

Question 35

Other than beta-catenins, what are some other mechanisms of cell-cell contact inhbition

Answer 35

Clustering of cadherins after cell-cell contact is known to alter the activation of small GTPases
e.g. Rac is activated, Rho is inhibited: this can influence proliferation

Question 36

What are the consequences of cells losing their behavioural constraints

Answer 36

Proliferate uncontrollably (lose density dependence of proliferation)
Be less adherent and will multilayer (lose contact inhibition of locomotion and anchorage dependence)
Cause epithelial breakdown cell-cell contacts
Be not Hayflick limited, express telomerase

Question 37

What are the consequences of loss of contact inhibition of locomotion for the progression of cancer

Answer 37

Promoting formation of solid tumours

Invasion and metastasis