Strategies for analysis of a complex
regulatory network in E. coli the acid stress response
Neil Burton
Matt Johnson
Peter Lund
My definition of the stress
response:
adaptive response of an entity to
any event which threatens its
optimal existence
What properties should a well-adapted (or wellengineered) stress response system have?
• It has to be able to:
– Detect the stress
– Convert the detection into a signal
– Respond appropriately (i.e., response must
ameliorate the stress in some way)
– Respond at an appropriate level
• This will include ending the response once the
stress has ended
A good understanding of a
network implies:
• Understanding of its components
• Understanding how the components interact
with each other (including dynamically)
• Understanding their adaptive value for the entity
• Understanding how the network may evolve
Technical approaches:
• ’omics approaches – how do the different ’omes
change during stress?
• Analysis of known genes in detail in wt and
different mutant backgrounds - esp. time series
• Comparisons between related strains that
respond differently to same stress
• Evolution of novel stress response phenotypes
in the lab
Acid resistance (AR) and enteric
bacteria
• Important for passage
through stomach (pH 1-3)
• E .coli can survive
remarkably well in
conditions of high acidity
(<pH2.5)
• Wide range of genes which
protect against acid are
induced by exposure to
mild acid stress (pH 5.7)
Mild acid
pH 5.7
Aims:
•
P EvgA
EvgS
RpoS
ydeO
•
•
gadE
gadBC
Other cytoplasmic and
periplasmic components
•
•
Describe and understand
the dynamics of the
system
Identify novel components
and their interactions
Understand the adaptive
value of the network and its
components
Understand how the
network might evolve given
new selective constraints
Choice of models:
– E. coli K-12 MG1655
– E. coli O157:H7 (Sakai)
Lux plasmid reporter system
Promoter
(intergenic region
+100-200bp)
Stops, RBS
P. luminescens
LuxCDABE
Operon
pLUX
Light
Low copy
pSC101
replication
• Non-invasive
• Automation for
high temporal
resolution
Questions
• Does this system give us usable data?
• Can it be used to explore different mutant
backgrounds/different strains?
• Can we uncover novel aspects of the
network using this approach?
General messages
• Promoter probes using lux/gfp provide reproducible, high
density time-series data, suitable for model fitting
• They can be used to identify potential new links in
regulatory networks
• However, the outputs don’t necessarily map directly onto
transcriptional outputs
• Lab-based evolution and whole genome resequencing
has great potential for understanding the nature and
evolution of stress response systems (and other
networks) at the whole organism level
Matt
Neil
UoB:
Nick Loman
George Weinstock,
Washington Uni
Dov Stekel
NIH