Synthetic gene networks

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Engineered Gene Circuits Jeff Hasty
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• How do we predict cellular behavior from the
  genome? Sequence data gives us the components,
  now how do we understand the full system?
• How can we control or monitor cellular behavior?
  Diseases, pathogenic invasions involve
  alterations of natural dynamics - can we
  reestablish normal function?

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Gene Regulation
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Gene regulatory networks

• Proteins affect rates of production of other
  proteins (or themselves)
• This allows formations of networks of interacting
  genes/proteins
• Sets of genes whose expression levels are
  interdependent

B
A
C
D
E
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John Tysons Analogy
Using gene and protein network wiring diagrams
to try to deduce cellular behavior is akin to
using a VCR circuit diagram to try to deduce how
to program it. Mathematical models are needed
to translate gene-protein wiring diagrams into
manuals explaining cellular processes.
But how do we construct reliable and useful
mesoscopic models?
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Engineered Gene Circuits
Faithful modeling of large-scale networks is
difficult Alternative Design and build simpler
networks Decouple complexity Use model to
design experiments Systematic comparison of
model and experiment Forward Engineering of
useful circuits Design networks to perform
tasks Couple to host - control or monitor
cellular function
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Engineered Toggle Switch
Model - design criteria
Construction/experiments
Gardner, Cantor Collins, Nature 403339 (2001)
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The Repressilator
Elowitz and Leibler, Nature 403335 (2001)
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A Detailed Example Single-Gene Autoregulatory
Module
Well-characterized Kinetic parms known Tunable
control CI857 denatures with temp Build network
with off-the-shelf molecular biology Theoretical
predictions Bistablity and hysteresis (Hasty
et al PNAS 972075, 2000)
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Biochemical Reactions
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Rate Eqs For cI Monomers and GFP Reporter
Model predictions as the temperature is varied?
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Model Prediction Multistability
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Experimental Protocol
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Bistability Results
Prediction
Observation
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Model the Fluctuations
- OK when fluctuations dominated by production
and degradation - Distributions numerically check
with Monte Carlo gold standard - Still working
on systematic demonstration of validity
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Model Versus Experiment
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Coefficient of Variation
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Genetic Relaxation Oscillator
Hasty et al, Chaos (2001)
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Relaxation Oscillator Analysis
Design network so that y is a slow variable
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Drive Oscillator With Cell Division Cycle
Identify known oscillating gene product and its
target promoter
SWI4 forms a complex and activates the HO promoter
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(No Transcript)
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Resonant Dynamics
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Summary

• Use of biochemical kinetics to describe gene
  regulation (in bacteria)
• Models can be used to develop tailor-made
  circuits
• Gene circuits lead naturally to problems relevant
  to nonlinear dynamics, statistical physics and
  engineering
• Noise from small molecular numbers is a dominant
  source
• Genetic states accessed through fluctuations
  (noise-induced transitions between attractors)

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Collaborators
Milos Dolnik (Brandeis) David McMillen (Boston
University) Vivi Rottschafer (Leiden)
Farren Isaacs (BU) Charles Cantor (BU-UCSD) Jim
Collins (BU)
Funding NSF, DARPA and the Fetzer Institute
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The Human Genome Project
Secret of Life Solved!
Cells fully understood!
Molecular biology finished!

• Why is this not true?
• Network dynamics not yet understood