Tutorial: Exploitation of the actin cytoskeleton by listeria

Question 1

Explain the formation of actin filaments

Answer 1

1. Nucleation: Three G-actin (Globular actin monomers) come together and form a filament nucleus for further elongation
2. Elongation
   
   • ​​actin monomers add to preexisting filaments at either end. There is a fast growing (ATP) and a slow growing (ADP) end

Question 2

How does polymerisatio of actin lead to force production?

Answer 2

The individual actin filaments of the comet tail of the listeria stay stationary within the cell (because they are crosslinked

• to each other
• to cellular filaments with alpha actinin
• other crosslinking filaments

When new filamenta re created (at the end of the trial clostest to the bacterium) : the bacteria pushes forward

Question 3

Explain through which mechanism Listeria modulates the cytoskeleton

Answer 3

Listeria expresses ActA at one pole of the bacterium

• binds to actin directly (but only limited movement) Mainly:
• Binds to VASP –> adaptor porotein causes Profilin accumulation and therefore actin filament formation

Question 4

What is VASP?

What is its role in listeria infection?

Answer 4

It is an adapotr protein that enables movement of the bacterium intracellulatrly

• Binds to ActA on bacterial surface
• Regruitement of Arp complexes (nuclei for new actin filaments)
• Profilin accumulation (supply of ATP G-filaments for polymerisation)

Question 5

Where does the energy for movement of the listerium inside the cell come from?

Answer 5

From Hydrolysis of ATP rich Actin to ADP

Question 6

Explain the life cycle of Listeria

Answer 6

1. Binds to extracellular adhesion molecule (E-cadherin) –> Triggers internalisation
2. In lysosome of cell–> listeria gets activated and breaks down enzyme of lysosome
3. Gets into the cytoplasm and moves
4. Migrates from cell to cell without leaving the intracellular compartment

Question 7

How does the life-cylce of listeria relates to the risk of infection?

Answer 7

Can cross placenta and BBB

1. It is present everywhere (e.g. soil) and can easily be eaten (fresh fruit or vegetables, unpasteurised milk, soft cheeses) and can proliferate at normal fridge temperature
2. It is an intracellular pathogen –> no antibody immune response
3. –> Needs cell-mediated immune response and therefore puts people with weak immune system at risk
   
   • pregnant
   • young
   • old
   • immunosupression

Question 8

Which structural components of the Bacterium Listeria allow it to exploit the host cytoskeleton?

Answer 8

It has ActA proteins at one end –> mimics host zyxin (which is an endogenous host protein)

• both: Zygin and Acta cause VASP bding (for vasodilator-stimulated phosphoprotein) which in turn binds profilin and Arp complex
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