POSTERS - BLAST X - University of Utah

BLAST X Tue. Morning Session
THE ROLE OF POSITIVE FEEDBACK IN CONTROLLING FLAGELLA ASSEMBLY
DYNAMICS
Supreet Saini 1 , Christy Aldridge 2 , Jonathan Brown 2 , Philip Aldridge 2 , Christopher Rao 1
1
Department of Chemical and Biomolecular Engineering, University of Illinois, Urbana, IL 61801,
United States
2
Institute for Cell and Molecular Biosciences, Newcastle University, Framlington Place,
Newcastle upon Tyne NE2 4HH, United Kingdom
Flagellar assembly in Salmonella enterica serovar Typhimurium (S. typhimurium)
proceeds in a sequential manner, starting from the base of the flagella and concluding at the
filament tip. A key regulatory step in the assembly process is the σ 28 -FlgM checkpoint, which
prevents the activation of σ 28 -dependent Pclass3 promoters prior to completion of the hook basal
body. This regulatory checkpoint is typically assumed to involve a binary decision process:
either proceed with Pclass3 gene expression or not, depending on the state of assembly.
However, mathematical modeling suggests that this binary checkpoint may in fact result in more
subtle, rheostat-like control. σ 28 is involved in a positive feedback loop, as ιτ positively regulates
its own expression along with the expression of FliZ, an activator of Pclass2 gene expression. In
addition, the ability of σ 28 to regulate gene expression depends on the concentration of FlgM in
the cell. This suggests that the response of the σ 28 positive feedback loop is tuned by late
protein secretion. In fact, our modeling and experimental results suggests that σ 28 and FlgM are
not only involved in establishing the binary checkpoint between Pclass2 and Pclass3 gene
expression but are also involved in fine tuning the relative timing of expression. In particular, the
positive feedback loop involving σ 28 and FliZ establishes the delay between Pclass2 and Pclass3
gene expression.
In this talk, we will discuss our recent work investigating the positive feedback loops
involving σ 28 and FliZ. We have recently shown that FliZ is an FlhD4C2-dependent activator of
Pclass2 gene expression. In addition, our results indicate the FliZ speeds up the induction of Pclass2
genes in a secretion-dependent manner. With regards to σ 28 , we have found that autoregulation
controls the relative timing of gene expression. In cells lacking FlgM, both Pclass2 and Pclass3
genes are induced at the same times. Conversely, when the rate of FlgM secretion is reduced,
the delay between Pclass2 and Pclass3 gene expression is exaggerated. These results suggest that
timing is responsive to the rate of late protein secretion. Moreover, mathematical modeling
predicts that this control is due to autoregulation by σ 28 . To test this model, we have rewired the
flagellar gene circuit by replacing the native PfliA promoter with Pclass1/Pclass2/Pclass3 promoters.
Consistent with the model predictions, these promoter replacement experiments show that
autoregulation plays a key role in enforcing the timing of flagellar gene expression. Last, we also
investigated gene expression dynamics at single-cell resolution, and our preliminary results
suggest that the dynamics may exhibit bistability, consistent with control involving feedback.
Collectively, our results suggest that the regulation of flagellar gene expression is complex and
involves multiple layers of control.
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