Biological clocks in theory and experiments www.amillar.org

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Biological clocks in theory and experiments
www.amillar.org

• Current
• Megan Southern
• Laszlo Kozma-Bognar
• Kieron Edwards
• Vacancy !
• Paul Brown
• James Locke
• Domingo Salazar
• Ozgur Akman
• Vacancy !

Past Simon Thain Kamal Swarup Ruth
Bastow Harriet McWatters Shigeru Hanano Seth
Davis Mandy Dowson-Day Giovanni Murtas Neeraj
Salathia Maria Eriksson Anthony Hall Alex
Morton Boris Shulgin Nickiesha Bromley Victoria
Hibberd
Collaborators IPCR - Matthew Turner (Physics)
David Rand (Maths) Bärbel Finkenstädt
(Statistics) Mark Muldoon and David Broomhead
(Manchester) Lorenz Wernisch (Birkbeck) Antony
Dodd, Alex Webb, Julian Hibberd
(Cambridge) Ferenc Nagy (Szeged) Eberhard
Schäfer, Stefan Kircher (Freiburg) Mark Doyle,
Scott Michaels, Rick Amasino (Madison) Graham
King (HRI), Mike Kearsey (Birmingham)
Funding BBSRC, Gatsby, EPSRC, DTI
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Genome-wide circadian rhythms

• 12 RNA transcripts rhythmic in white light
• 3,000 genes of 22,000 on array
• Functional clustering
• 68 of rhythmic transcripts also
  stress-regulated, (Kreps et al. 2002)

Edwards et al. unpublished
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Mutant plants identify genes in the clockwork
?
Alabadi et al., 2001

• Negative regulation during the day - CCA1/LHY
• Positive regulation at night
• Mathematical model to test potential for
  regulation

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Luciferase (LUC) reporter

• Luminescence reflects transcription rate of
  promoter
• Unstable activity reports dynamic regulation
• Spatial resolution, high throughput

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LUC identifies mutants in clock genes
EARLY-FLOWERING 3 (elf3) arhythmic in light
Hicks et al, Science, 1996 McWatters et al.,
Nature, 2000 Reed et al., Plant Phys. 2000
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The circadian clock in Arabidopsis
TOC1
3600 genes OvertRhythms
LHYCCA1
Oscillator
Input

• Modelling
• Design principles
• Extension of network

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Projects in circadian rhythms - Current
Data
Models
Data analysis
Data preparation Rhythm detection Parameter
estimation (MCMC, cost functions)
Reporter genes RNA (PCR, arrays) Mutant plants
Central loop ODE, SDE, Simplified, etc.
Software
Clock mechanism. Functions of interlocking
loops, multiple light inputs
Understanding
Model analysis
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(No Transcript)
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Single-loop network model

• Locke, Millar and Turner, J Theor Biol, 2005.
  Global parameter search.

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Interlocking loop model for Arabidopsis clock

• Model- J. Locke
• Hypothetical components X, Y
• Cost function fit to WT and mutant behaviour
• Predicts X and Y expression

WT
cca1lhy
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Experiments to identify Y turn up a good candidate

• Prediction dashed line
• M. Southern tested candidate genes by qRT-PCR.
  Data crosses
• Unexpected light response of GIGANTEA RNA matches
  prediction
• GI also matches other predictions for Y from
  literature

WT
cca1lhy
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Projects in circadian rhythms - Current
Data
Models
Data analysis
Data preparation Rhythm detection Parameter
estimation (MCMC, cost functions)
Central loop ODE, SDE, Simplified, etc.
Understanding
Model analysis
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Data analysis CABLUC in 16h L8h D
Morton, Finkenstadt
raw
prepared
synthesis rate
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Parameter estimation simple model for synthesis
rate
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Comparison of WT and elf3 mutant waveforms
WT
elf3 mutant
clock effect

• Quantify distinct features within the timeseries
• Now apply to parameters of simple clock models

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Projects in circadian rhythms - Current
Data
Models
Data analysis
Data preparation Rhythm detection Parameter
estimation (MCMC, cost functions)
Reporter genes RNA (PCR, arrays) Mutant plants
Central loop ODE, SDE, Simplified, etc.
Software
Clock mechanism. Functions of interlocking
loops, multiple light inputs
Understanding
Model analysis
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Projects in circadian rhythms Future
collaboration
Data
Models
Data analysis
Reduced/synthetic systems Protein data (2-D
gels) Biochemistry (parameters)
Fitting directlyto multiple data types Network
inference (dynamic Bayes nets)
Database
Photoreceptors, secondary loops
Software
Stochastic processes Noise (internal and external)
Understanding
Model analysis