Cell migration & invasion Flashcards
what are the four main roles of the actin cytoskeleton?
- driving membrane protrusion
- cell shape changes
- maintaing cell-ECM linkages
- cell contraction
how is TGFbeta1 involved in remodelling of the actin cytoskeleton?
Activation of TGFbeta1 activates RhoGTPases which contribute ot remodelling of the actin cytoskeleton
briefly describe the Rho GTPase cycle
inactive Rho is bound to GDP (and sometimes guanine nucelotide dissociation inhibitor GDI)
(GEF) guanine nucleotide exchange factor catalyses the dissociation of GDP from the Rho protein allowing GTP to bind, induce a conformation chagne in to the active form
(GAP) GTPase activating protein promotes the conversion of the active GTPase-GTP form to the GTPase-GDP form - removal of phosphate return the RhoGTPase to an inactive state
give three example of RhoGTPase family members
Rho A
Rac1
Cdc42
injection of constituatively active RhoA causes what changes in actin expression
it causes actin stress fibres and focal adhesions on the outer parts of the cell
injection of constituatively active Rac1 causes what changes in actin expression
it causes lamellipodia formation -> actin rich membrane ruffles and small focal adhesion contacts
injection of constituatively active Cdc42 causes what changes in actin expression
filipodia formation -> finger like actin protrusions
what are the main effectors and function of RhoA
RhoA activates the contractile phenotype of actin, a myocin dependent process.
(myocin walks along actin filaments and tightens them up)
- in addition it induces actin polymerisation (via formin) and stress fibre formation
what are the main effectors and function of Rac
Actin polymerisation and actin branching (lamellipodia)
via activation of (WAVE and then) Arp2/3
what are the main effectors and function of Cdc42?
actin polymerisation and filopodia
via activation of (WASP then) Arp2/3
what are focal adhesions?
A multicellualr protein complex at the plasma membrane interface that links the extracellular environment to intracellular cytoplasm
interaction with ECM is mediated by integrin receptors on cell surface
what proteins are involved in focal adhesions?
- actin-binding proteins [a-actinin, vinculin, myosin],
- signalling proteins [p130Cas, Src, focal adhesion kinase (FAK)],
- structural proteins [paxillin, talin],
- integrin receptor
what are the functions of focal adhesions?
provides tensile strength
cell shape
facilitates membrane protrusion
cell migration
promotes cancer metastasis
what are integrin receptors?
heterodimeric receptors comprising an alpha chain and beta chain - link internal actin cytoskeleton to ECM (except a6b4)
different combinations interact with specific extracellular matrix proteins -> provide links from inside to outide of cell
what are the four main proteins involved in focal adhesion proteins?
- FAK - structural support and signalling platform
- Src - tyrosin kinase (RhoGTPase activation)
- vinvulin and talin - actin binding proteins
- paxillin - adaptor protein
what are the four stages of actin faccilitated cell movement
- Membrane extension facilitated by actin polyerisation and branching (lamelipodia and filipodia)
- Adhesion by focal adhesion formation
- Translocation (retraction facilitated by increased tension/contraction)
- De-adhesion
describe the actin treadmilling effect
- A linear actin filament (F-actin) is made up of many individual globular actin (G-actin) monomers
- These G-actin monomers get preferentially added to the growing (plus) end of a filament (called polymerisation) and disassemble at the retreating (minus) end of a filament
- Each molecule of actin is bound to either ATP or ADP
- ATP-G-actin is added to the growing end of the filament and ADP-G-actin is disassembled from the retreating end of the actin filament
what is the difference in organisation between lamellipodium and filopodium?
Lamellipodium is made up of branched actin filaments (branches via Arp2/3 complex)
Filopodium is made up of parallel actin filaments connected by actin cross linking protein
what occurs at the trailing edge of a cell that is migrating?
RhoA signals via ROCK and pMLC t oincrease myosin contractility
the uropod is rich in myosin 2 which cross links actin and contributes to contractility
what happens at the leading edge of a migrating cell?
Filipodia form via Cdc42 signalling to formin to promote actin polymerisation
Lamellipodium form via Rac1 and Cdc42 signalling via WAVE and WASP respectively activating Arp2/3 to promote branched actin
what are the four different types of actin organisation within a migrating cell?
Lamellipodium
Filopodium
Focal adhesion
Lamella
what are the four types of cell migration?
- Collective migration
- Mesenchymal cell migration
- Amoeboid migration
- Scaffold cell dependent migration
what are the similarities and differences between mesenchymal and collective cell migration?
- similarities - both are path generating
- differences -
collective retains cadherin cell-cell junctions while only exhibiting focalised cell-matrix adhesion and ECM degredation in leader cells.
mesenchymal only has cell-ECM junctions and travels as a single cell degrading the ECM as it goes
what are the differences between mesenchymal and amoeboid cell migration
mesenchymal: path generating, proteolytic ECM degradation, focalised cell-ECM interaction; integrin receptor clustered (focal adhesions)
amoeboid: path finding, morphological adaptation (actin contriction), pseudopod, diffuse cell-ECM interactions; integrin receptors non-clustered
what matrix metalloprotease is always membrane bound?
MT1-MMP
what are matrix metalloproteases?
- Zn2+ dependent proteases secreted by cells into extracellular space
- secreted as an inactive pro-form and following activation in the extracellular space they cleave extracellular matrix proteins and/or activate latent TGFb1
- Regulate migration, invasion, proliferation, differentiation, angiogenesis
What is the function of MT1-MMP in cancer cell
MT1-MMP directly degrades ECM (eg basal lamina), allowing cells to invade into underlying connective tissue layer
MT1-MMP can activate MMP-2 within connective tissue layer, thus futher facilitating cancer cells ability to migrate and invade surrounding tissue
what two ways does MT1-MMP remodel extracellular matrix
directly; remodels extracellular matrix (fibronectin, collagen, laminin, vitronectin)
indirect; remodelling through activation of MMP2
what is the natural inhibitor of MMP activity?
TIMP-2
describe the activation of MMP-2 via MT1-MMP
- MT1-MMP monomer expression on the cell surface
- MT1-MMP dimerisation through Hpx and TM domain
- formation of activation complex of dimer MT1-MMP, TIMP-2 and proMMP-2
- Propeptide cleavage by the MT1-MMP that isnt bound by TIMP-2
- MMP-2 activation followed by release of active MMP-2
how do levels of Tissue inhibitor of metalloproteinase (TIMP)-2 regulate MMP activation?
what are invadopodium?
actin rich cell protrusions of the plasma membrane that secrete MMPs and degrade extracellular matrix
