lecture 24 Flashcards
1
Q
What are cell adhesion molecules?
A
- cell adhesion is critical in morphogenesis
- for the formation and maintenance of cellular structures (e.g. epithelia)
- for the ability of cells to move with respect to their environment (e.g. cell migration and axon guidance)
Cell-cell adhesion: e.g. E-cadherin—E-cadherin in adherens junctions
Cell-ECM adhesion: e.g. integrin to laminin
2
Q
What is the role of the cytoskeleton in morphogenesis?
A
- regulation of the actin and microtubule cytoskeleton is crucial for many morphogenetic cell behaviours
- important for changing shape, movement etc
3
Q
What sort of control is required for cell migration?
A
- molecular control of cell migration is complex
- cell migration and other motile behaviours (e.g. axon guidance, branching, morphogenesis) involve highly complex regulation of many cellular processes such as cytoskeletal remodelling, receptor-ligand interactions, and vesicle trafficking
4
Q
What is Cell Migration at a Glance?
A
- cell has polarity: front and back/trailing and leading edge, definitely heading in one direction
- one thing migrating cells do is extend protrusions
- thin protrusions = filopodia
- broad, sheet-like protrusions = lamellipodia
- actin filaments sliding against each other under the influence of myosins
- focal adhesions controlled by integrins
- adhesion dyanmics
- integrin to actin cytoskeleton
- polarity regulated by complexes like CDC42, apical complex
- adhesions at the back need to be released, microtubules bring something that dissolves the adhesions
5
Q
What is regulation of actin polymerisation and contractility?
A
- the assembly and contraction of actin filaments
- crucial for many cell behaviours such as cytokinesis, cell shape changes, and cellular protrusions
- myosin II is found in muscle and these migrating cells/normal cells (e.g. cytokinesis)
- heads pull on actin filament to make them slide together
- Myosin light chain regulates whether the myosin is contractile or not
- myosin regulatory molecule is active when phosphorylated
- MLC kinase and MLC phosphatase regulate phosphorylation of light chain
- two main types of polymerisation
- molecules help a polymer add monomers to the growing end (even though they will also spontaneously grow)
- diaphenous
- Arp2/3 complex (actin related protein)
- binds to an existing filament and then nucleates a new filament that comes off at 70º exactly
- contraction of networks of actin filaments is controlled by myosin II . myosin is comprised of a heavy chain (MHC) and a regulatory light chain (MLC)
- myosin contractility depends on MLC phosphorylation, which is regulated by MLC kinase and MLC phosphatase
- elongation of actin filaments is promoted by formins such as Diaphanous (mDia1) which bind to the end of actin filaments
- formation of new branches is promoted by the Arp2/3 complex which binds to the side of actin filaments
6
Q
What are Rho small GTPases?
A
- regulate actin structures
- Rho GTPases are molecular switches that cycle between an inactive, GDP-bound form and an active, GTP-bound form
- when active they can bind, and activate downstream effector proteins
- they are activated by Rho GTP exchange factors (RhoGEFs)
- they are inactivated by Rho GTP Activating Proteins (RhoGAPs)
- different Rho GTPases control formation of different actin structures
e. g. Cdc42-GTP = filopodia, Rac1-GTP = lamellipodia, RhoA-GTP = stress-fibres
7
Q
How do Rho small GTPases regulate the actin cytoskeleton?
A
three key GTPases are:
- RhoA: activates myosin contractility, via Rho kinase, which inhibits MLC phosphatase, and polymerisation through mDia
- Cdc42: stimulates polymerisation through mDia and formation of new branches via WASP, which activates the Arp2/3 complex
- Rac: stimulates new branches, via WAVE, which activates the Arp2/3 complex, and inhibits contraction through the kinase PAK
8
Q
What is genetic regulation of cell behaviour?
A
- cells have a large range of behaviours, but how are these behaviours controlled during development?
- answer: differential gene expression
9
Q
What is an example of genetic regulation of cell behaviour?
A
- Twist regulates mesodermal cell behaviour
- the Drosophila presumptive mesodermal cells express the transcription factor Twist
- during gastrulation these cells fold inwards (i.e. invaginate), undergo an EMT, and then migrate out over the ectoderm
- all of these behaviours are controlled by twist
10
Q
What does Twist do?
A
- transcription factor in presumptive mesodermal cells
- turns on 3 key genes (fog, snail and heartless)
- fog: initial furrowing, bending
- myosin apical constriction
- snail: EMT
- inhibits E-cadherin, epithelial cell-cell adhesion
- heartless: migration
- FGF-receptor, motility
11
Q
What is Fog?
A
- target of Twist
- controls epithelial sheet folding
- normally myosin II accumulates on the apical side of cells
- in folded gastrulation (fog) mutants, myosin localisation is patchy, and invagination of the mesoderm fails
- twist
- fog
- RhoGEF2
- Rho1
- Rho kinase
- myosin localisation constriction
12
Q
How does Fog control myosin II localisation?
A
- secreted Fog is thought to bind to an unknown receptor which activates Rho1 which activates myosin contraction, leading to localisation of myosin filaments
13
Q
What is snail?
A
- target of twist
- controls loss of epithelial adhesion
- the EMT is too rapid (~15 mins) to be accounted for only by transcriptional mechanisms, so there are likely to be other Twist targets, which regulate the dissolution of existing adherens junction complexes
- snail is turned on by Twist in the ventral presumptive mesoderm
- E-cadherin normally repressed in the mesoderm
- repression depends on snail
14
Q
What is Heartless?
A
- twist target, aka FGFR
- regulates migration
- heartless is an FGF receptor tyrosine kinase expressed in the mesoderm
- its FGF ligands are expressed in the dorsolateral ectoderm under the control of the transcription factor Dorsal
- mesodermal cells expressing heartless migrate towards the source of the FGFs
15
Q
What are the molecular mechanisms acting downstream of Heartless?
A
- unknown
- in other situations, activation of receptor tyrosine kinases, like the FGF-receptor, can stimulate F-actin polymerisation via signalling pathways
other pathway
- Cdc42 activates WASP - affects polymerisation of actin
- RTK phosphorylated has PI3-Kinase bind to its tail, phosphorylates PIP2 –> PIP3, can be bound by GEF, GEF activates Cdc42
- theoretical
16
Q
What is a theoretical pathway from growth factor to F-actin?
A
- activated FGF-receptor becomes phosphorylated
- PI3 kinase binds to a phosphorylated FGF-receptor
- PI3K converts to PIP2 (i.e. PtdIns(4,5)P2) to PIP3
- GEF is recruited to membrane by binding to PIP3
- GEF activates Cdc42, which activates WASP, which activates Arp2/3 complex, which stimulates formation of F-actin branches
