Lecture 12

Question 1

What is polarity?

Answer 1

This is a difference in the structure, composition or function between the two polls of a cell, such as the apical/basolateral in an epithelial cell or the axon/dendrite in a neuron. In epithelial cells, this also means location of a protein in a specific location

Question 2

The polarity affects what?

Answer 2

where proteins go

Question 3

Epithelial cells develop distinct ______ and ______ domains as an epithelia forms

Answer 3

apical

basolateral

Question 4

Proteins must be sorted and directed to apical and basolateral membrane for the ____ transport function of epithelial cells to function

Answer 4

ion

Question 5

What is the first step to forming an epithelia?

Answer 5

1. Cell basement membrane and cell-cell interactions

Question 6

Describe the first step to forming an epithelia in a lab

Answer 6

You start off with a cell that doesn’t have the typical cell shape. It is not connected to the basal lamina and the Na+/K+ ATPases and ENaC are not in any specific location. These single cells when they want to form an epithelia is they bind to the basement membrane through hemidesmosomes but the ion channels are still in random places. This binding to the BM helps to orient the channels. Once they are bound to the BM, they start to make connections with each other and we get intercellular connections forming. *the ion channels are at their appropriate membrane

Question 7

Describe how an epithelia is formed in the body

Answer 7

In the body epithelia, there are founder cells (a type of stem cell) and if we get (eg.) a scratch, the cells divide and new cells form with the BM through hemidesmosomes. The new cells then form intercellular connections with neighbouring epithelial cells, helping to form an enlarged epithelium

Question 8

Which intercellular connection forms first after the cells bind to the BM?

Answer 8

adherens junctions

Question 9

Describe how adherens junctions are formed

Answer 9

1. nectins form to make initial cell to cell contact
2. when bring the cells in close contact which allows E-cadherin to form a homodimer with E-cadherin on another cell. This requires Ca2+
3. the cytoplasmic tails of the E-cadherin bind catenins which links to the actin cytoskeleton
4. Catenins also link nectin and cadherin complexes to pull all the proteins together to make the adherens junction

Question 10

Which processes are required for establishing epithelial polarity?
A. Neighbouring cells interacting with each other through adherens junctions.
B. Connections between epithelial cells and their basement membrane.
C. Nectin proteins from neighbouring cells connecting with each other.
D. All of the above.

Answer 10

D. All of the above.

Question 11

What is the third step to forming an epithelia, after forming a connection with the basement membrane, and then forming adherens junctions?

Answer 11

step 3: small GTP binding proteins are activated and step 4: activation of polarity complexes

Question 12

Describe step 3: small GTP binding proteins are activated

and set 4: activation of the polarity complexes

Answer 12

Adherens junctions form and the adherens junctions recruit a number of other proteins including small GTP binding proteins (CDC42). CDC42 activates PKC to help form polarity complexes.

Question 13

What are the four polarity complexes that are formed by PKC?

Answer 13

1. PKC
2. PAR
3. CRB
4. SCRIB

Question 14

What do polarity complexes do? What is the advantage of this?

Answer 14

form tight junctions

these are right close to the apical surface so it helps to form apical and basolateral domains

Question 15

What is the importance of the polarity complexes?

Answer 15

they maintain the apical and basolateral polarity

Question 16

Describe step 5: forming tight junctions

Answer 16

these are formed from claudins, occludins and JAMs and they are connected to the cytoskeleton through ZO proteins and it prevents the movement of proteins between apical and basolateral domains

Question 17

What are the three roles of the tight junction?

Describe these

Answer 17

• barrier: limits the passage of ions and molecules between cells in the paracellular pathway
• gate: can allow certain solutes to flow through the paracellular pathway
• fence: prevents the movement of proteins between apical and basolateral domains

Question 18

Describe step 6: positioning of three protein polarity complexes

Answer 18

Atypical PKC is activated by GTP binding proteins such as CDC42. This aPKC is the only polarity protein that has any enzymatic activity (the other ones just serve as scaffolding, holding the cell in place to maintain a polarity). PKC phosphorylates CRBS and SCRB. This is the signal for these to move to their respective parts of the cell (apical and basolateral respectively). When this happens, we get mutual exclusion

Question 19

Which protein polarity complexes sits apically?

Answer 19

PAR and CRB

Question 20

Which protein polarity complexes sit basolaterally?

Answer 20

SCRIB

Question 21

What is mutual exclusion?

Answer 21

When SCRB is in the basolateral bit, and CRB is in the apical bit, it prevents the basolateral proteins moving to the apical membrane and vice versa

Question 22

Summarise the of formation of TJs and epithelial polarity (7 steps)

Answer 22

1. Interactions between neighbouring cells, and between cells and basement membrane.
2. Interaction between cells forms adherens junction.
3. Small GTP proteins (cdc42) activated.
4. Cdc42 activates aPKC - a polarity complex protein.
5. Tight junctions start to form.
6. Positioning of PAR and CRB polarity complexes to apical domain, and SCRIB complex to basolateral domain.
7. Apical-basolateral polarity established.

Question 23

Crumbs defines the basolateral domain and scribble defines the apical domain BECAUSE aPKC is the only polarity complex with enzymatic activity

Answer 23

D – First false, second true

Question 24

When epithelial cells need to divide, we need to pull apart these junctions and polarity complexes and we have to be able to reform them if needed. How does this happen?

Answer 24

through TGF-β

Question 25

What does TGF-β do?

Answer 25

TGF-β binds to its receptor in the membrane which will disrupt these adherens junctions, tight junctions and polarity complexes in one of two ways:

1. it will directly inhibit the PAR complex (which helps to define apical domain)
2. it activates SMADs

Question 26

what are SMADs?

Answer 26

These are transcription factors which, when activated, will translocated into the nucleus, so there will be an increase in the expression of Snail, Zeb and Twist.

Question 27

What do Snail, Zeb and Twist do?

Answer 27

These down regulate a huge number of proteins involved in tight junction and adherens junction formation which causes the down regulation of cadherins, claudins, occludins and ZO proteins which means we can’t have adherens or tight junctions

Question 28

What is the relevance of breaking up adherens and tight junctions?

Answer 28

it is one of the first steps in epithelial mesenchymal transition (EMT)

Question 29

What is epithelial mesenchymal transition (EMT)?

Answer 29

How a cell can go from an epithelial cell to a mesenchymal cell to allow it to divide and then come all the way back to an epithelial cell again

Question 30

How does a cell go from an epithelial cell to a mesenchymal cell to allow it to divide and then come all the way back to an epithelial cell again?

________ inhibits/downregulates _________ complexes so the ______ junctions and _________ junctions are ________ _______. The cells start to lose their _________. You also lose your connection to the ________ _________ as we lose the expression of ___________. We also get rearrangement of the ____________ filaments towards the front end of the cell

Answer 30

TGF-β inhibits/downregulates polarity complexes so the tight junctions and adherens junctions are falling apart. The cells start to lose their connections. You also lose your connection to the basement membrane as we lose the expression of hemidesmosomes. We also get rearrangement of the actin filaments towards the front end of the cell

Question 31

Why is EMT important when it comes to cancer?

Answer 31

If cells go through EMT but don’t go back again, they remain as mesenchymal cells. Their growth rate is really quick and they are able to migrate and invade

Question 32

What four things do mutations in polarity complex proteins cause?

Answer 32

• Decreased formation or lack of tight junctions so barrier, gate and fence functions compromised.
• Changes in cell-cell adhesion and cell movement.
• Changes in location of apical and basolateral proteins.
• Cancer

Question 33

When epithelia are fully polarised with AJs and TJs formed this structure inhibits ________ ________ pathways, and promotes differentiation (epithelia takes on specialised function):

ZO can sense there is a _______ cell density there and so the ZO stop __________ ________ that promote cell division close to the ________ junctions so they can’t _________ into the ___________ and help initiate cell _________. This means the epithelia can carry out normal functions such as _______ and ____________

Answer 33

cell division
ZO can sense there is a high cell density there and so the ZO stop transcription factors that promote cell division close to the tight junctions so they can’t translocate into the nucleus and help initiate cell division. This means the epithelia can carry out normal functions such as absorption and secretion

Question 34

If we have some sort of damage to the epithelia (eg. we cut ourselves) What do the ZO proteins do?

Answer 34

They can sense that we have a low cell density and they release transcription factors that promote cell division which allows them to translocate into the nucleus and increase the expression of genes that will help initiate cell division.

Question 35

What is important about cancer?

Answer 35

Genetic changes in cancer cells of epithelia may promote EMT: cell division increases and loss of epithelial polarity.
If cancer cells are mesenchymal through genetic changes in the epithelial cells, the tumour cells can leave the epithelial tumour, invade into the blood stream, migrate around the body, invade a new site and they can become epithelial again