invasion - regulation of cell cycle mobility Flashcards
what are the molecular mechanisms that regulate motility *
microfilaments
regulation of actin dynamics
cytoskeletal proteins
signalling proteins
What are the steps in tumour progression *
homeostastis
genetic alterations
hyper-proliferation
de-differentation - disassemble cell-cell contacts, lose polarity
can form a tumour mass and invade BM
invasion - increased motility and cleavage of ECM proteins
(from hyperproliferation to invasion is the change from benign to malignant tumour)
what is de-differentiation in tumour progression *
losing the knowledge that the cells are epiuthelial cells
normally tightly cohesive and highly polarised (ie nucleus at abse of cell)
normally cells are separated from stroma by the basement membrane which provides support for the epithelial cells
describe invasion in tumour progression *
increased motility
cleave ECM proteins so cells find canals to migrate through the ECM
describe the sequence of events in metastasis *
epi cells in primary tumours are tightly bound together - but not as organised as neighbouring normal cells
they cleave the BM
metastatic tomour cells become mobile mesenchyme type cells, enter the stroma, and then enter the bloodstream
metastatic cells then travel through the bloodstream to a new location in the body
they then exit the circulation through the endothelial cells and invade a new organ
the cells lose their mesenchymal characteristics and form a new tumour
sometimes the cells reverse and reacquire the neighbour-neighbour contacts
describe the types of tumour cell migration *
tumours can travel alone or as a collective (cluster of cells - they maintain their attachment ti each other and have a higher metastatic potential)
what are the different migration strategies *
ameoboid - individual tumour type - usually migrate as round structure but depends on the type - tumour tyoes are lymphoma, leukaemia, SCLC
mesenchymal - either single cell/chain - found in fibrosarcoma, glioblastoma, anaplastic tumour
cluster/cohorts - travel as a group through the ECM - found in epi cancer and melanomas
multicellular strands/sheets - larger number of cells - in epi cancer or vascular tumours
describe the signalling involved in different tumour cell migration strategies *
different types need different signalling processes
all involve integrins and proteases - proteases digest the ECM, integrins receptor of ECM proteins (like foot of cell so cell can migrate)
for cluster migration - coordination is fone by the gap junctions between epi cells (they allow communication between the cells), also cadherins induce differentation and hold the cells together so they can drag each other along (they are different to the cadherins in normal tissue)
describe how tumour cell metastasis mimics morphogenetic events *
2d sheet - cells migrate and drag cells behind
vascualr sprouting - need to regrow bv - tip cell drive newly formed vessel to a site where it needs to reconnect with the vasculature
branching morphogenesis - in mammary gland need spur of growth - need to proliferate and differentiate the epithelium - this is done by collective migration - form terminal end bud that brings cells together to invade and form the lactating gland
multicellular 3d invasion strands - also uses tip cells
detached cluster - have cells at the front and bring other cells along
border cells - in fly ovary cells - 1 side of ovary have cluster of nurse cells that feed the ovary - these celsl need to migrate through the egg so that they do their business at the front - migrate as a cluster - similar regulation in malignancies
explain the migration of primary glial cells in a scratch assay *
scratch the assay of cells
teh cells recognise there is a gap so proliferate to fill it
when the cells reach each other, they stop growing
this takes 16hours
describe the migration of a glial tumour cell line on a scratch assay *
the cells are not compact - loose cells close together
when they sense the space the cells migrate randomly and alone
when cells reach each other they continue to grow
this is much faster than in normal cells
compare the expression profile of invasive cells v primary tumours *
in invasive cells there is an upregulation of genes involved in cytoskeleton regulation and motility machinery
to test - put EGFL in injection (potent mitogenic and induces motility) - so the tumour cells grow up into needles
collect teh cells from the heterogenous tumour mass that are highly likely to metastisise because could sense EGF and leave primary tumour
compare the expression of these cells to that of the primary tumour - measure the RNA - higher expression of EGFR etc
this upregulation is surprising because the level of oncogenes was similar
what are stimulants for cell movement *
organogeneisis and morphogenesis
wounding
GF/chemoattractants - chemoattractants stimulate immune cells to move
dedifferentiation in tumour cells
what happens to cells when they move *
they have to change their shape - become polarised - so they get a front/leading edge
teh organelles are directed to dorection of motility
how do cells know where to go *
directionality - towards chemoattractant/space available
can either be to a specific place or with no purpose
how do epithelial cells know when to stop *
contact-inhibition motility
tehy recognise their neighnours
how do cells move *
they have specialised structures - focal adhesions, lamellae and filopodium
describe the attachment of cells to the substratum - ECM proteins *
when moving the cells form focal adhesions - very close contact with the RCM
the actin filaments are organised as bundles - they finish at focal adhesions
the cytoskeleton (actin bundles) are coordinated to go to the focal adhesions - this provides a hook to make traction forces for the cells to move
describe the complex joining the ECM to the cells *
the receptor is integrin - there are dimers of an a and b subunit, different combinations of the monomers will form different types of integrins
at cytoplasmic site - very short tail with no enzymatic activity so to do function it organises plaque of cytoskeletal proteins
the plaque acts as a signalling port and connects to the cytoskeleton
describe filopodia *
tehy are finger like prtusions rich in actin filaments
vinculin is a protein that binds to filamentous actin - one of the main components of the integrin plaque - there is a high concentration of vinculin forming the filopodia
filopodia are used to sense the surroundings of the cells
describe lamellipodia *
sheet like protusions rich in actin filaments
cells form contacts from large areas of membrane
the membrane attaches then comes back to the cell so that the cell can move
what control is needed in cell movement and why *
- within a cell- to know what is happening in differnet parts of the cell
- regulate adhesion/release of ECM receptors
- control froom outside to respond to external influences - ie sensors (know where the GF and nutrients are) and cues for directionality
needed because if cant coordinate different parts, the cells will move in opposite directions and split
what are the different types of motility *
hapoptatic - no purpose
chemotatic - purpose
describe the steps in cell motility *
1 - extension - extend the lamellipodium
2 - adhesion - form a new focal adhesion - this is like a step
3 - translocation - contract the back of the cell and move the cell body forward
4 - de-adhesion - de-attach the last focal adhesion so you can move forward
describe the relationship between G-actin and F-actin *
G-actin are small soluble subunits
F-actin are large filamentous polymers with polarity - have positive and negative ends so different ends have differnet functions
the small monomers can polymerase quickly
for example - a signal eg a nutrient causes disassembly of F-actin filaments quickly and rapid diffusion of subunits to side where nutrients are; then rapid reassembly of the filaments at a new site
describe actin filament organisation ion differnt structures *
actin is a filamentous cabel - exerts force so cells can move
in filopodium the fibres are in parallel bundles - this allows movement
in lamellipodium - the filaments are short, branched and crosslinked forming filaments
behind the lamellipodium are arched filaments where the stress fibres are - they are antiparallel contractile structures - focal adhesion structure is at end
describe the different processes involved in remodelling actin *
sequestering - maintains G-actin
bindling
severing - cut the filaments
nucleating
cross linking
capping
side binding
motor proteins - allows contraction
in order to react to a stimulus and disassemble then reassemble everything cells have to engage all these processes and proteins
steps in actib remodelling *
- nucleation
- elongation
- capping
- severing
- cross-linking and bundling
- branching
- gel-sol transition by actin filament severing
describe nucleation *
this is the limiting step in actin dynamics
need 3 actin monomers together to start remodelling - not stable
so instead use nucleators (Arp3 and 2) that resemble actin - they bind to actin and the cell thinks that 3 actin monomers are bound together = begin polymerisation
G-actin monomers are added to the plus end of actin chain
describe elongation *
when have actin filament - the arp complex can come off
actin-profilin complex fascilitates the monomer bidning
thymosin binds to G-actin and keeps it so it cannot bind to actin chain - restrict proliferation
prolifin competes for actin monomers with thymosine
molecules that inhibit actin binding are B4-thymosin, ADF/Cofilin
describe capping of actin *
it tells actin to stop growing - hooks onto the end of filaments to stop monomers being added
then filament becomes depolymerised from the otehr end, unless it is blocked
caps on +end - Cap Z, gelsolin, fragmin/severin
-end - tropomodulin, Arp complex
describe severing *
in unsevered population, actin filaments grow and shrink relatively slowly
in severed population actin filaements grow and shrink more rapidly
severing proteijns - gelsolin, ADF/cofilin, fragmin/severin
describe the cooperation of actin function to generate filaments *
profilin-actin complex cause elongation at +ve end (barbed end)
filament severing at opposite end - barbed end capped and depolarisation and monomer recycling at otehr end
annealing - join together 2 short fragments
can have growth from pre-existing ends
describe cross-linking and bundling *
group filaments in different states
fascin (mutated in melanomas)
a-actinin - dimer that cross links filaments
spectrin (stabalise cortical structure) and filamin - bring filaments at an angle - form a mesh
dystrophin - join actin to plasma mem, mutated in muscle wastage diseases
other proteins involved - fimbrin, villin, vinculin
this occurs when the actin filament is not severed so it is stabalised in the cells
filaments that are joined by myosin can be buckled if there is a contraction when the orientation of filaments is in opposite directions - opp polarity allows the filaments to slide over each other and cells contract
describe branching *
filaments branch at 70degrees
up to 3 filaments bind and then polymerise a new branch
protein involved is the Arp complex
describe gel-sol transition by actin filament severing *
network maintain rigidity of membrane in cytosol
but cell needs to protude so the network needs to open up - so cleave the filaments at certain sites = cross linked filaments that are not connected to each other
there is a transition from the gel phase to the sol phase
the protein that does this is gelsolin
what phases of actin remodelling are at different stages of cell movement *
extension - polymerisation (disassembly, nucleation, branching, severing, capping, bundling)
adhesion - gel-sol transition, attachment to the ECM
translocation - contraction
de-adhesion - deattachment
describe the processes involved in lamellae protusion *
branched filaments at 70degrees
there is capping so the filaments are short
polymerisation brings the membrane forward
back needs to disassemble and be clipped = form G-actin
G-actin goes to front of lamellae so it can be reassembled
this involves spatial, temporal, and molecular regulation
describe actin regulation in filopodia *
in parallel bundles
comprehensive actin assembly at tip - need protein to bundle them
retrograde flow - so get deassembled at back
when filopodia form focal adhesions, need to stop growing so are capped - also disassembled at end so retract beck to the cell body
what are the differnt cell shape and actin organisations *
if you have bristles/hairs - have fascin/forked heads
microvilli - cross linked filopodia proteins are villin, fimbrin and esprin
stereocilia in ears - haev a progressive height - esprin and fimbrin
filopodia - fascin, a-actin
lamellipodia - arp2/3, filamin, capping protein, cofiolin, gelsolin
what are the signalling mechanisms that regulate teh actin cytoskeleton *
ion flux changes - intracellular ca - some cytoskeletal proteins bind ca = upregulated func
phosphonositide signalling - phospholipid binding
kinases/phosphtases - phosphophorylation cytoskeleton proteins
signalling cascades via small GTPases
describe signalling by small G proteins *
Rho family of GTPases belong to the Ras superfamily - family members Rac Rho and Cdc42 afre best known
they participate in a variety of cytoskeletal processes, are activaetd by TK, adhesion receptors and signal transduction pathways
expression levels are upregulated in different tumours - this activates the pathways indirectly
there are some point mutations on rho
what happens when rho family is activated 8
drive specific shapes
cdc42 = very long filopodia
rac = spread large lamellae in any direction
rho = 2 rho proteins - form stress fibres which are very pwerful contractors
mechanism of rho proteins *
they are the top of the chain
layer of effectors that are specific from Rho gtpases - Pak, PI5K (this is a lipid kinase), formin, IQGAP
the actin binding proteins that are regulated by Rac/cdc42 are WAVE, arp2/3, cofilin, profilin, WASP, arp2/3
WASP is downstream of Cdc42 and activates Arp2/3
WAVE is a partner of the Arp2/3 complex and helps polymerisation
after this actin polymerisation and organisation into filaments, Rho can come in and bundle them
where do GTPases fit in with the cell migration *
Rac activation restricted to one area of cell - lamellipodium
focal adhesion is rac/rho
assembly of stress fibres, control of myosin motor that gives tension to fibres, and contraction by myosin is contorlled by rho
rho is also de-adhesion
cdc42 - tell cell where to go by generating the filopodia and involved in polarised motility - receptors that sense chemoattractants need cdc42 to transduce the signal into the cell, important for actin pol because with raq controls the arp2/3 complexes
