Modeling the chemosensing system of E' coli

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Modeling the chemosensing system of E. coli

• Ned Wingreen Princeton
• Juan Keymer Princeton
• Robert Endres Princeton/NEC
• Yigal Meir Ben Gurion University

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Outline

• Introduction to chemotaxis in E. coli
• Runs and tumbles
• Signaling properties
• The chemotaxis network
• FRET
• New probe of receptor activity
• Two regimes of activity
• Receptors function collectively
• Modeling
• How does adaptation work?

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Signaling properties of the chemotaxis network

• Precise and robust adaptation range of 3-4
  orders of magnitude of attractant
• Signal integration multiple attractants
• Sensitivity amplification

Segall, Block, and Berg (1986)
CCW vs CW bias for tethered cells in response to
step in attractant
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The chemotaxis network
http//www.rowland.harvard.edu/labs/bacteria/proje
cts_fret.html
but signal integration and sensitivity are still
not well understood.
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Chemoreceptors
Dimer
Tar - aspartate, glutamate (900 copies) Tsr -
serine (1600) Trg - ribose, galactose (150) Tap
- dipeptides (150) (Aer - oxygen via FAD (150?))
Sensor
Transmembrane helices
Linker region

• Attractant binding inhibits phosphorylation of
  CheA
• Adaptation
• More attractant ? increased methylation by CheR ?
  increased rate of phosphorylation of CheA
• Less attractant ? increased demethylation by CheB
  ? decreased rate of phosphorylation of CheA

380 A
Methyl binding sites CheB, CheR
Cytoplasmic domain
CheA / CheW binding region
Stock (2000)
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Chemoreceptor clustering
Receptors are clustered globally into a large
array, and locally into trimers of dimers.
Gestwicki et al. (2000)
Kim et al. (1999) Studdert and Parkinson (2004)
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In vivo FRET studies of receptor activity
Real-time measurement of rate of phosphorylation
of CheY. FRET also allows subcellular imaging,
Vaknin And Berg (2004).
Sourjik and Berg (2002)
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FRET data two regimes of activity
Sourjik and Berg (2002)

• Regime I
• Activity moderate to low at zero ambient MeAsp
  (0.06,1)
• KD small and almost constant
• Regime II
• Activity high (saturated?) at zero ambient MeAsp
  (1.3-1.9)
• KD1 large and increasing with methylation
• Plateau in activity
• KD2 approximately constant

Two regimes of receptor activity consistent with
2-state receptor model.
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2-state receptor model

• Originally proposed by Asakura and Honda (1984).
• Modified by Barkai and Leibler (1998) to explain
  precise and robust adaptation
• Receptor complex has 2 states on, i.e. active
  as kinase, and off, i.e. inactive as kinase.
• Demethylation only occurs in on state, i.e.
  when receptor is active, so that
• Therefore, at steady state,
• Which implies precise and robust adaptation of
  each receptor complex to a fixed activity.

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Two regimes of a 2-state receptor
But first, a 1-state receptor
Regime I
Regime II

• Regime I
• Activity low to very low at zero ligand
  concentration
• KD KDoff
• Regime II
• Activity high (saturated) at zero ligand
  concentration
• KD increasing as eon ?

Off
Free Energy
Off
KD
Ligand
Ligand
Ligand
However, single receptor does not account for low
apparent KD in Regime I.
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Receptor-receptor coupling
Duke and Bray (1999)
Duke and Bray (1999) proposed that
receptor-receptor coupling could enhance
sensitivity to ligands.
Toy model if N receptors are all on or all
off together,

• Regime I (?e gt 0)
• Low activity e-N?e at zero ligand
  concentration
• KD KDoff / N
• Hill coefficient 1
• Regime II (?e lt 0)
• KD KDoff e-?e
• Hill coefficient N

Receptor-receptor coupling gives enhanced
sensitivity (low KD) in Regime I, and enhanced
cooperativity (high Hill coefficient ) in Regime
II.
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Review of FRET data

• Regime I
• Low, constant KD
• Activity low at zero ligand concentration
• Hill coefficient 1
• Regime II
• KD1 increasing with methylation
• Activity high at zero ligand concentration
• Hill coefficient 1 ?
• Plateau in activity ?

KDoff /N, value of N ?
Hill coefficient increases with receptor
homogeneity. Must consider mixture of different
receptor types.
Sourjik and Berg (2004)
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Model 1d mixed lattice of receptors

• Regime I
• KD set by coupling energy EJ
• Regime II
• Plateaus Tars off, Tsrs on
• Hill coefficient 1, no cooperativity because
  Tar receptors separated by Tsr receptors

Normalized Activity
Log(MeAsp)
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Receptor homogeneity and cooperativity

• Receptors are in Regime II
• Hill coefficient increases with homogeneity
  because clusters of identical receptors grow.
• KD (or KD1) increases as lattice becomes more
  mixed because of coupling EJ to on receptors.

Normalized Activity
Log(MeAsp)
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Adaptation
Adaptation uses methylation to return all
receptors to ?e 0, and thereby enhances
sensitivity.
?e gt 0
?e 0
?e 0
?e gt 0
?e gt 0
?e 0
?e 0
Off
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Precise adaptation
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Scaling of wild-type response data
Sourjik and Berg ?MeAsp ? ?FRETTar(QEQE)
Free energy scaling ?MeAsp ? ?(Fon Foff)
Sourjik and Berg (2002)
Includes zero-ambient data!
Doesnt collapse zero-ambient data.
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Predictions

• For homogeneous lattice
• Transition from Regime I to Regime II with
  methylation
• Adaptation range set by KDon

Activity
MeAsp
Sourjik (unpublished)
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Open questions

• Lattice structure?
• Mechanism of receptor-receptor coupling?
• Do other receptors work this way?

Stock (2000)
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Conclusions

• Signaling properties of the chemotaxis network
• Precise and robust adaptation
• Signal integration
• Sensitivity
• FRET studies reveal two regimes of activity
• Regime I low activity and constant KD
• Regime II high activity and increasing KD
• Model of coupled 2-state receptors account for
  signaling properties, and for two regimes
• Regime I (?e gt 0) coupling ? enhanced
  sensitivity
• Regime II (?e lt 0) coupling ? enhanced
  cooperativity (but only for homogeneous clusters)
• Adaptation homogenizes receptors (?e 0) for
  enhanced sensitivity