external factors controlling cell division

Question 1

what is cell behaviour

Answer 1

the term used to describe the way cells interact with their external environment and their reactions to this, particularly proliferative and motile responses of cells.

Question 2

what external influences are detected by the cell *

Answer 2

chemical - hormones, GF, ion concentrations, ECM, molecules on other cells, nutrients, dissolved gas (O2/CO2) concentrations

physical - mechanical stress (movement), temperature (for us regulated narrowly, so doesnt have a massive effect), the topography/layout of the ECM and other cells

Question 3

what external factors can influence cell division *

Answer 3

all of the factors that are detected - eg if dont have enough nutrients, proliferation can slow down

however with respect to cancer - GF, cell-cell adhesion, cell-ECM adhesion

Question 4

describe cell spreading *

Answer 4

not a passive gravity, driven process - cells will spread even when upside down - it is controlled by the cytoskeleton

energy is required to modulate cell adhesion and teh cytoskeleton during spreading

it varies betweebn cell type

as cells spread, they lose their blebs

Question 5

how do we know that adhesion influences cell proliferation *

Answer 5

microengineer surfaces with patches that were adhesive of various sizes, on a non-adhesive background - therefore cells could only spread on the patch

compared to cells in agar (cells dont proliferate in agar - they cant sense it, so feel like they are in nothing)

GF were present

cells in agar - very small probability that they would enetr S phase and proliferate

on adhesive patch the probability of proliferating increased with the size of the patch

therefore, we know cells require to be binded to the ECM to be competent to respond to GF

Question 6

how do we know the importance of cell spreading *

Answer 6

fibronectin (a molecule where cells would adhere) was put in a single dot and compared to the same amount of fibronectin in different spots

in single patch - cell dies by apoptosis

when fibronectin is spread out - cell can bridge over the non-adhesive surface = proliferation and survival

therefore cell survival depends on the arrangement of the contact

Question 7

what is important about the cell-ECM adhesion that allows proliferation and survival *

Answer 7

cells need to be attached to ECM and allowed to spread to begin protein synthesis and proliferation

therefore attachment to ECM may be required for cell survival - this is anchorage dependance

Question 8

describe how a cell phenotype can be determined by the composition of the matrix *

Answer 8

in interstitial matrix ie with Type 1 collagen - mammary epi doesnt differentiate into secretory cells - instead lose ball of cells are formed without normal cell-cell junctions

in basal lamina matrix ie type 4 collagen and laminin - mammary cells organise into organoids and are polarised, differentiated and secretory

Question 9

how does a cell recieve information about its surroundings from its adhesion to ECM *

Answer 9

cells have receptors (membrane proteins) on their surface which bind specifically to ECM molecules

the cytoplasmic domain of these molecules are linked to the cytoskeleton

therefore there is mechanical continuity between the ECM and interior of cell

Question 10

describe integrins and their role in sensing external env *

Answer 10

they are heterodimer complexes of a and b subunits that associate extracellularly by their head regions - ligands bind here

each of the leg regions spans the membrane - short cytoplasmic tail

there are>20 a/B combinationss that bind to specific short peptide sequence of ECM proteins

eg a5B1 binds to arg-gly-asp this is tehe RGD sequence

peptide sequences (eg RGD) are found in >1 ECM eg RGD is in fibronectin, vitronectin, fibrinogen and others

most integrins link to cytoskelton via actin binding proteins ie link proteins (ecept a6B4 integrin complex in helidesmosomes linked to cytokeratin [intermediate filament] network)

the integrin complexes cluster to form local adhesions (or hemidesmosomes - a6B4) - the clusters are involved in signal transduction

some integrins bind to specific adhesion molecules on other cells eg in immune system and clotting

Question 11

explain signalling to and from the ECM receptors *

Answer 11

integrins can transduce signals - ECM binding to an integrin complex can stimulate the complex to produce a signal inside the cell - this is ‘outside in’ integrin signalling

Question 12

describe the conformational changes to integrin during signal transduction *

Answer 12

they can be flexed or extended - switching between forms alter ability to bind to ligands - therefore cell-ECM adhesion and hence the signalling can be switched off

when flexed - legs are bent

in extended conformation, the legs are open

Question 13

describe ‘outside in’ signalling *

Answer 13

a cell can recieve information about its surroundings from adhesion to ECM

integrins recruit proteins that help with signalling and recruiting cytoskeleton - these proteins cluster - they recruit src (important in signalling) and Fak (phosphorylates src and influences its proliferative capacity)

the composition of the ECM will determine which integrin binds and therefore what signals the cell recieves - altering the phenotype of the cell

focal adhesions sense the mechanical properties of their surroundings - when the proteins experience force they open up, changing their properties ie they can recurit differnet proteins

the amount of force that is generated at an adhesion depends on the force generated by the cytoskeleton and the stiffness of the ECM

Question 14

describe inside-out signalling *

Answer 14

a signal generated inside the cell (eg because of hormone binding) can act on integrin complex to alter the integrin’s affinity for ECM - ie activates the receptor

this occurs in inflammation/blood clotying - switches on adhesion of circulating leukocytes, and switches on the integrins on platelets otherwise they would be adhesive all the time

Question 15

describe the conformational changes to an integrin complex during activation and signalling *

Answer 15

low affinity - bent conformation = weak or no binding to ligand

high affinity - extended conformation = strong binding to ligand

in some cells might always be in high affinty/always a signal, in others low is prefered until signal

signal from inside the cell acts on the low affinity integrin complex to promote the switch to the extended high affinity conformation - this is inside out activation - switches adhesion on

ECM ligand binds causes the further opening of the leds - exposes the binding sites for the recruitment of cytoplasmic signalling molecules - this is outside in activation

Question 16

describe the density dependance of cell division *

Answer 16

at high cell density the cells compete for growth factors

when cells in culture form a confluent monolayer - they stop proliferating and slow down other metabolic activities

this used to be known as contact-inhibition

however - if you add more GF to a certain area proliferation is stimulated again and cells grow on top of each other

this indicates it is competition for GF not cell-cell contact that inhibits division

Question 17

what mechanism to GF use to alter gene expression and proliferation of cells *

Answer 17

ERK pathway - MAPK

Question 18

what are the signals that control proliferation of tissue cells *

Answer 18

GF - depends on density

ECM - depends on anchorage

they activate identical signalling pathways - MAPK

however, individually they are weak/transient

together activation is strong

therefore the separate pathways act synergistically

Question 19

what are the different types if contact interaction between cells *

Answer 19

short term - transient interactions between cell which dont form stable cell-cell junctions

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

Question 20

describe how cell to cell contact between non-epi cells differs from epithelial cells *

Answer 20

epi cells - their job is to form junctions and layers

non-epi - when collide they dont form cell-cell contacts - tehy repel each other by paralysing mptility at the contact site - this promotes formation of a motile front at another site (ie form lamellipod) and the cell moves in the opposite direction

this is contact inhibition of locomotion and is responsible for preventing multilayering of cells in culture/in vivo

Question 21

describe long-term cell-cell contacts *

Answer 21

some cells stringly adhere and form specific cell-cell junctions (adherens junctions, desmosomes, tight junctions, gap junctions)

this happens in epithelial cells and endothelial cells - they form layers, and neurons forming synapses

junctions in epi: continuous belts (zonula) or as discrete spots (macula) also desmosomes and gap junctions

Question 22

descripe contact induced epithelial spreading *

Answer 22

contact between epi cells leads o the mutual induction of spreading and actin polymerisation - so the total spread area of contacted cells is more than the sum of the 2 separated cells - this could cause a stable monomer

Question 23

how does cell-cell adhesion affect proliferation *

Answer 23

the molecules involved in cell to cell junctions are Ca dependant

if you remove the ca there are no junctions - this activates MAPK and decreases p17^KIP1 = high proliferation

if ca is added again - cell to cell junctions form = inactivated MAPK = increased p17^KIP1 = low proliferation because p27 is an inhibitor of proliferation

get the same effect if add adhesion blocking Ab = high proliferation, remove Ab = low prolif

Question 24

describe the molecular organisation of teh adherens molecules (cell-cell junctions) *

Answer 24

have cadherins that have linking molecules that link them to the cytoskeleton

cadherins are Ca dependant, homophilic cell adhesion molecules - they bind to identical molecules on adjacent cells

inside the cell molecules inc B-catenin and a-catenin link cadherin to the actin filament

Question 25

describe adenomatous polyposis coli (APC) (

Answer 25

inherited colon cancer - there are many inherited family forms

the APC gene-product is a protein involved in the degredation of B-catenin which links teh cadherin junctions to the cytoskeleton

when B-catenin is bound - the plasma membrane is involved in forming cell junctions so is sequested

when cytoplasmic B-catenin is increased - if APC complex is active - b-catenin is rapidly degraded; if APC inactive (in cancer) - B-catenin associates with a molecule called LEF-1 forming a TF taht influences gene trasncription = proliferation

Question 26

other than B-capetin - what adhesion associated pathways influence contact induce inhibition of proliferation *

Answer 26

clustering of cadherins after cell-cell contact is known to alter the activation of small GTPases eg Rac is activated and Rho is inhibited - this can iduce proliferation

some GF receptors are associated with cell-cell junctions - this reduces their capacity to promote proliferation

Question 27

what happens if a cell loses its behavioural constraints *

Answer 27

they proliferate uncontrollablu - lose density dependance of proliferation

are less adherent and will multilayer - lose contact inhibition of locomotion and anchorage deopendance

epithelia break down cell-cell contacts

not Hayflick limited - express telomerase

ie they become cancer

Question 28

describe the loss of contact inhibition in cancer cells *

Answer 28

they pile up so form a solid tumour and invasion (ploughing through otehr tissues)

Question 29

what are proto-oncogenes *

Answer 29

receoptors, signalling intermediates and signalling targets

if the genes coding for a component of a signalling pathway is mutated so that the protein is constituitively actibe - the pathway will be permenantly on

this is the mechanism of short-circuitig leading to uncontrolled proliferation asa result of loss of GF dependance

Question 30

definition of an oncogene *

Answer 30

mutant gene which promotes uncontrolled cell proliferation

Question 31

definition of a proto-oncogene *

Answer 31

normal cellular gene corresponding to the oncogene

Question 32

examples of proto-oncogenes and their oncogene *

Answer 32

EGFR

Ras (signalling intermediate) - V12Ras - mutation - in 30% of all cencers - in 90% of pancreatic cancers

c-Raf (signalling intermediate) - v-Raf - deletion of regulatory domain

c-Jun (TF) - v-Jun - deletion of a regulatory domain

Question 33

mechanism for uncontrolled proliferation of tissue cells *

Answer 33

if both GF pathway and ECM pathway are switched on all time - no longer need the external signals = proliferation w/o the normal constraints

if only have 1 mutation - wont be fully uncontrolled - need both

Question 34

describe benign tumours *

Answer 34

encapsulated and dont invade

Question 35

describe malignant tumours *

Answer 35

invade and metastasise - cause the most problems

Question 36

describe local invasion and met *

Answer 36

major feature of cancer is its ability to spread

most human cancers are carcinomas (epithelial)

in order to metastasise - cells must break away form tumour, travel in bv or lymph, lodge at a distant site, leave the vessel, ultimately establish a secondary tumour

cell-cell adhesion must be broken down

the cells must be motile

degradation of ECM must occur - matrix metaloproteinase (MMP) levels increased in order to break thorugh basal lamina and ECM

lose contact inhibition of locomotion to travel

the degree of cell-cell adhesion is an indicator of how differentiated the primary tumour is, and indicates its invasiveness and the prognosis

Question 37

examples of GF *

Answer 37

EGF - epidermal GF

PDGF - platelet derived GF