Bacterial Pathogenesis and the Cytoskeleton

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Bacterial Pathogenesis and the Cytoskeleton

• Bacteria utilize functions of the cytoskeleton to
  infect cells
• Bacterial protein production changes upon
  infection
• Specialized pathways tell about cellular
  functions

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Background Readings

• Regulating Bacterial Expression by Methylation
  (reviewed in Low et al., 2001. Infection and
  Immunity 697197-7204)
• Microbial pathogenesis and cytoskeletal function
  (reviewed in Gruenheid Finlay. 2003. Nature
  422775-81)
• Chapter 16 in Molecular Biology of the Cell 4
  (Alberts et al.)

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Control of Pilus Production by DNA Methylation
From Low et al., 2001
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Bacteria Bind Stronger with More Force
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Molecular motors in bacteria how do pili pull?

• Pilus function
• twitching motility
• (unspecific adhesion to surface)
• interaction with host cell
• (specific adhesion)
• horizontal gene transfer
• Modelsystem Neisseria gonorrhoeae

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Pilus-Dependent Propulsion
Neisseria gonorrhoeae produce type IV pili that
are 2-5 microns long
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Functions of Type IV Pili

• Transformation
• Genetic Transfer
• Conjugation
• Phage Infection
• Virulence
• Twitching and Colony Motility

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Pili Are Homopolymers of Pilin
6 nm
4 nm
Inner Membrane
Outer Membrane
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• Pilus is a helical polymer
• - monomer pilin
• - pitch 4nm, 5 pilin per turn
• length up to 5µm, diameter 6nm

Forest et al Gene, 1997
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Twitching Movements Involve Retraction of Fibers
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Does the host cell sense the amount of force
generated by pilus retraction?
Accumulation of panCD44
Merz, A. et al Mol. Microbiol. 1999
Bacterial adhesion evokes host cell response
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Current molecular model for pilus
retraction pilus is a dynamic polymer
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We conclude for the most likely molecular
mechanism
A single pilT complex generates 110pN by an ATP
driven conformational change.
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AAA ATPases
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Is motor oligomerization involved in the
generation of forces in excess of 100pN?

• PilT belongs to the family of AAA ATPases
• AAA ATPases are involved in the dissociation
• of protein complexes
• in vitro PilT forms hexamers

Electron micrograph of a PilT homolog (TrbB)
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Kaiser, D. Current Biol. 10R778 (2000)
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ATP-dependent motors
Kinesin transports vesicles and mitochondria on
microtubules
Direct measurement of steps showed basic
stepsize is 8nm Variance of translocation per
time showed 1 ATP is hydrolized per step at
negligible load Tight mechanochemical coupling
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dNTP-dependent motors
DNA polymerase replicates DNA
Replication rate follows a simple Arrhenius
law V v0exp(ndF/kBT) n 2nts or 4nts
B.M., D.Bensimon, V.Croquette, PNAS 2000
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Translocation ratchet
Simulation
Flux of segments
 
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Motors driven by proton-motive forces
Bacterial respiration creates a transmembrane pH
gradient. Proton flux from the outside to the
inside of the cell generates force. Example
E.coli flagella Induction of MotA expression.
The motor MotA is supposed to change conformation
upon proton translocation.
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Kinetic Scheme For Kinesin Movement (Processive)
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Kinetic Scheme for Myosin Movement on Actin
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Bacterial gene transfer

• Biological relevance of bacterial gene transfer
• Genetic diversity. Escheria coli contains 17
  foreign DNA.
• Virulence. Pathogenicity islands are easily
  transfered between bacteria.
• Antibiotic resistance. Oral streptococci act as
  genetic reservoirs and
• transfer antibiotic resistance to pathogenic
  bacteria.

Proposed molecular machine for DNA uptake
Promising tool to study the molecular mechanism
of DNA uptake