Extracellular matrix integrins and cell migration Flashcards
Where is extracellular matrix secreted from?
Cells
What components make up the ECM?
Laminin
Collegen
Elastin
What are fibronectins?
Large, secreted proteins
What domains do fibronectins have?
- Heparin binding domain - interact with cells
- Collagen binding domain
- Self-association domain
What is the structure of laminin?
Trimer
Many domains
What does laminin interact with at the C terminal domain?
- Intergrins
- Perlecan
- Dystroglycan
What is the structure of integrins?
- Alpha and beta subunits
- Each monomer has a single transmembrane domain
- Alpha subunit is cleaved and held together by disulphide bonds
- Extracellular cysteine rich domain
- Matrix binding domain at the amino terminus
- Intracellular component is anchored to actin
How do intergrins interact?
Heterodimerically
In integrins, what do extracellular cysteines allow?
- The formation of disulphide bridges
- To interact with different extracellular components
How do integrins interact with actin?
Using talin and vinculin
What does the amino terminus of integrins bind to?
What does this result in?
The ECM and divalent cations (Ca2+, Mg2+)
Results in activation of the integrin and activates intracellular signalling pathways that influence the behaviour of the cell
How many alpha and beta subunits are there of integrins?
How can they bind?
18 subtypes
Can be paired in many combinations - to make 24 varients
How are integrins acitvated?
In 2 ways:
1) Binding to the ECM
- Causing a conformational change (unfolding of extracellular domain)
- Become activated
- Activate the intracellular domain to bind to talin
2) If integrin interacts with intracellular domain - causes unfolding of the extracellular domain
What are FAKs?
Focal adhesion kinases
A type of tyrosine kinase
How are FAKs activated?
What do activated FAKs do?
Activated by integrin binding to the ECM, on the foramtion of adhesions
When activated - FAK phosphorylates tyrosine residues - docking regions for other proteins
What is the typical pathway that actin is polymerised?
- Fibronectin
- Integrin (extracellular)
- FAK (in the cytoplasm)
- Tyrosine phosphorylation (localised in the area of focal adhesion)
- Actin fibre polymerisation
What are focal adhesions?
- Protein complexes where the cell connects to the ECM
- Where mechanical force and regulatory signals are transmitted between the cytoplasm and the cell
What are FAKs involved with?
- MIGRATION (are made and formed)
- ANOIKIS (attachment-dependant cell death)
What do FAKs bind to?
The cytosolic tail of integrin, with the assistance of other proteins
Where to integrins cluster to?
What does this trigger?
The sites of the matrix contact
This triggers the assembly of cell-matrix junctions (focal adhesions)
What happens to cells if they are not attached to a substrate?
They die
What is motility linked with?
Adhesion
What forms the basis of cell motility in mammals?
Actin
What is the leading edge of a cell?
What is present in the leading edge and why?
The direction that the cell is moving in
Actin - pushes the cell forwards
Where are microtubules present in the cell?
In the centre
What are lamellipodium?
Actin projection at the leading edges
What happens to focal adhesions as the cell migrates?
The focal adhesions are recycled from the back of the cell to the front
What is the actin cortex?
What does it provide to the cell?
Skeleton underneath the membrane (surrounding the whole cell)
Maintains cell shape as the cell migrates
Prevents the cell becoming flat
Gives the migrating cell strength
How is the migrating cell polarised?
Different things happening at the front and rear:
- Recycling of focal adhesions
- Flow of actin to the front of the cell
What is ruffling of the leading edge and what is it important for?
- Actin polymerisation pushing the membrane up in different directions
- Important in sensing guidance cues
How is actin attached to the lamellipodia?
At the + end (distal to the cell)
What happens at the - end of lemellipodia?
Actin fillaments are removed and recycled to the + end
What is cytochalasin B?
A drug which binds to the + end of actin and prevents any further actin polymerisation
Can be used to study actin polymerisation
What complex is important in actin polymerisation?
The ARP complex
What is the structure of the ARP complex?
Made of many proteins (including Arp2 and Arp3)
What do Arp2 and Arp3 do?
Nucleate actin monomers by binding at the minus end and allowing actin to polymerise
What do Arp2 and Arp3 resemble?
Actin
What activates ARP complex activity? How?
Rac kinase - initiates the formation of ARP complexes
What is the result of Rac kinase?
Increase in formation of ARP complexes
Increase nucleation of actin filaments
Drive the leading edge forwards
How do ARP complexes form a branched networks?
What does this provide to the cell?
ARP complexes can bind to existing actin filaments
Provides stability and strength and pushes the leading edge forwards
What happens to focal adhesions when the cell migrates?
- Focal adhesions that interact with the ECM at the rear of the cell are broken
- Recycled to the leading edge to initiate new binding
Where in the migrating cell is stable adhesion?
At the leading edge
Where in the migrating cell is sliding adhesion?
At the rear
How are focal adhesions different?
They are all made of different molecules
Some are stable, whereas some are transient
What are the 2 different categories of focal adhesions?
What are the differences between the 2?
1) Low density
- Found around the leading edge
- Immobile (fixed within the cell)
2) High density
- Compacted
- Found away from the leading edge
- Can move within the membrane
What are low density focal adhesions?
rac1
cdc42-dependant
What are high density focal adhesions?
RhoA
Actin-myosin interaction-dependant
What are 3 signals that are used for motility?
1) Soluble/diffusible
2) Insoluble
3) Fibronectin
What are the soluble signals used for motility and how are they used?
Netrins
Released from an attraction point and diffuse away to attract cells to that point
What are the insoluble signals used for motility and how are they used?
- CAMs and components of the ECM
- Inserted into a tissue or the ECM and don’t diffuse away
- Act as contact attractants
Where is fibronectin secreted from?
What is this secreted fibronetin used for?
Secreted by migratory cells
To lay down a trail which is followed by other migratory cells
What 2 things inhibit fibronectin dependant migration?
1) Antibodies binding to fibronectin
2) RGD motif injection into migratory cels
What is the RGD motif and why does injecting it into migratory cells block fibronectin dependant migration?
Arginine - Glycine - Asparagine
It is important for fibronectin dependant migration - if artificially administered, it ‘swamps’ the binding site
What are 5 roles of endocytosis in motility?
1) Shaping chemotactic gradients
2) Membrane cycling
3) Confining signalling
4) Modulation of adhesive contact and ECM
5) Polarisation of endocytosis
What happens to the membrane of the cell when it is migrating?
- More membrane made to accommodate movement
- Membrane endocytosis around the cell and deposition (exocytosis) at the leading edge
What must happen to the membrane for the cell to move forwards?
It must be fixed by focal contacts to the ECM
What do integrins facilitate?
How?
Cell to extracellular matrix binding
Bind to components of the ECM, such as laminin and fibronectin
Bind to actin
