DNA looping causes stability and
robustness in the bacteriophage λ
genetic switch
Marco Morelli
Division of Ecology and Evolutionary Biology
Glasgow University
PNAS, 106, 8101 (2009)
Bacteriophage λ
The Genetic Switch
•  Two alternative pathways
•  Lysogeny -> Lysis transitions
induced by RecA
Lysis
•  In RecA- mutants we should
expect spontaneous, noisedriven transitions
Lysogeny
M. Ptashne, A Genetic Switch: Gene Control and Phage λ
(Cell Press & Blackwell Scientific Publications, 1986)
The Regulatory Network
•  Two mutually repressing
genes -> two proteins
abundant in the two states
Lysogeny
•  A structured operator
•  Cooperative interactions
•  Different promoter
strengths
M. Ptashne, A Genetic Switch: Gene Control and Phage λ
(Cell Press & Blackwell Scientific Publications, 1986)
Lysis
The DNA looping interaction
Another operator (OL, 2400bp from OR) bind cI
A cI octamer can be formed via DNA loop
Effects on the switch???
I. B. Dodd et al, Genes and Development 18, 344 (2004)
B. Revet et al, Curr. Biol. 9, 151 (1999)
I. B. Dodd et al, Genes and Development 15, 3013 (2001)
Puzzles
Measurement of the spontaneous switching rate (lysogeny ->
lysis): < 10-9 per generation per cell
J. W. Little et al, EMBO J. 18, 4299 (1999)
Stochastic models assuming equilibrium in binding of regulating
proteins to the operator predict much more frequent flips
E. Aurell et al, Phys. Rev. E 65, 051914 (2002)
How is it so stable?
Promoter activity measurements reveal only a little cI is
available at OR - probably non-specifically bound
I. B. Dodd et al, Genes and Development 18, 344 (2004)
I. B. Dodd et al, Genes and Development 15, 3013 (2001)
How can it be so robust to cI depletion?
A stochastic model
•  Gillespie algorithm on the core genetic network
–  Specific binding to single operator sites (OR and OL)
–  Dimerisation of transcription factors (cI, cro)
–  RNApolymerase binding to promoters
–  Production and degradation of mRNA and proteins
–  DNA loop formation
–  Nonspecific binding of transcription factors to DNA
•  Rate constants from (abundant) literature
•  Vary strength of DNA looping and non-specific binding interactions
Results/1 - Bistability
Steady state levels of regulating proteins are compatible
with measured values
ΔGNSB =-4.1 kcal/mol ΔGloop=-3.7 kcal/mol
Measuring the Switching Rate
Spontaneous switching events are rare -> hard to sample
Forward Flux Sampling method
T. S. van Erp et al, J. Chem. Phys. 118, 7762 (2003)
R. J. Allen et al, Phys. Rev. Lett. 94, 018104 (2005)
R. J. Allen et al, J. Chem. Phys. 124, 024102 (2006)
kAB = ΦA ,1P1,B
€
Order parameter: (total number of cI) - (total number of cro)
Results/2 - Switching rates
A simulation with explicit operator binding agrees with experiments!!
The DNA loop stabilises the switch against non-specific DNA binding
Results/3 - Robustness
DNA looping increases the bistable region, conferring
robustness to nonspecific binding
Conclusions
•  Stochastic fluctuations play a crucial role in the stability of
the bacteriophage λ genetic switch
•  The DNA loop provide stability and robustness to
nonspecific binding of regulating molecules to DNA
•  Our model predicts a switching rate in agreement with
experiments
•  Predictions:
–  A mutant with no DNA loop will form a much less stable lysogen
–  Depleting free cI will destabilise a non-looping mutant but have
a much less severe effect on the wild-type lysogen
Biochemical Group@AMOLF
Pieter Rein ten Wolde
Sorin Tanase-Nicola
School of Chemistry@Edinburgh
David Dryden
School of Physics@Edinburgh
Rosalind Allen
EPCC@Edinburgh
Alan Gray
High Performance Computing
Transnational Access Program