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Poster Full Abstracts - Systems and Quantitative Biology
Poster board number is above title. The first author is the presenter
367
Transcriptional mechanisms that compensate for the cost of bistability.
Alistair N. Boettiger
1
, Jacques Bothma
2,3
, Michael Perry
3
, Michael Levine
3
. 1)
Chemistry and Chemical Biology, Harvard University, Cambridge, Ma; 2) Biophysics Grad. Group, UC Berkeley, Berkeley CA; 3) Molecular and Cell
Biology, UC Berkeley, Berkeley CA.
Bistable switches govern a variety of cellular and developmental processes, including sporulation in Bacillus subtilis and rhodopsin expression in the
ommatidia of adult flies. Yet, they are inherently costly since a small change in the levels of a signaling molecule or transcription factor can produce a
catastrophic change in gene expression and cell identity. Here we explore the transcriptional mechanisms responsible for the reliable deployment of the
bistable switch controlling the establishment of the sharp mesoderm/ectoderm boundary in the early Drosophila embryo. We introduce an automated analysis
to simultaneously track many hundred thousand individual snail mRNA transcripts in separate cells of whole mount embryos. With this method, we find
wildtype mesodermal cells are able to transcribe snail and a surprisingly rapid rate, within a factor of two of the theoretical limit.
Transgenic expression experiments with snail show that this bistable regulatory architecture is exquisitely sensitive to molecular noise, and that small
quantitative differences in the levels of early snail mRNA resolve into large qualitative differences in embryo morophology and cell fate. We present
evidence that the rapid rate of transcription of snail in wildtype embryos minimizes gene expression noise for snail resulting from promoter switching. This
rapid and precise expression arises through the use of both paused polII and dual enhancers, and is essential for the formation of a sharp and accurately
positioned mesoderm-ectoderm boundary.
884B
Topological Dynamics of the Gap Gene System in
Drosophila Melanogaster
.
Lena Panok
1,2
, Konstantin Kozlov
6
, Svetlana Surkova
6
, Vitaly Gursky
7
,
John Reinitz
1,3,4,5
. 1) Department of Ecology and Evolution, University of Chicago; 2) Department of Applied Mathematics and Statistics, and Center for
Developmental Genetics, Stony Brook University, Stony Brook, New York; 3) Department of Statistics, University of Chicago; 4) Department of Molecular
Genetics and Cell Biology, University of Chicago; 5) Chicago Center for Systems Biology, University of Chicago; 6) Department of Computational
Biology, Center for Advanced Studies, St. Petersburg State Polytechnical University, St. Petersburg, Russia; 7) Theoretical Department, The Ioffe Physico-
Technical Institute of the Russian Academy of Sciences, St. Petersburg, Russia.
Pattern formation in
Drosophila Melanogaster
is a robust developmental process. The parallels between robustness in biological systems and the
mathematical concept of structural stability suggest that we analyze development with the aid of tools from dynamical systems. In this poster we present a
dynamical model that agrees with the properties of a biological system studied, and analyze what geometric structure(s) ensure the stability of the system.
Our analysis was carried out by smoothly varying the concentration of two maternal factors and determining how both stable and unstable equilibria points
responded. To further understand how our system responded to variation of upstream maternal factors (Bicoid and Caudal) we carried out a bifurcation
analysis. The goal of such analysis is to give a global understanding of the behavior of the system with respect to these parameters. The parametric portrait,
which subdivides [Bicoid]-[Caudal] space into regions with different dynamics is also computed. Finally four different mechanisms of pattern formation are
presented. Our ongoing work is to understand Kr mutants.
885C
New Applications of Synthetic DNA Technology: Testing the Combinatorial Effects of Co-Occurring Cancer Genes through RNAi Double
Knockdown.
Jennifer R. Moran, Xiaoyue Wang, Kevin P. White. Institute for Genomics and Systems Biology, University of Chicago, Chicago, IL.
The freedom to synthesize DNA molecules that do not exist in nature opens up exciting new experimental possibilities. The IGSB Recombineering Core
Facility is now able to synthesize DNA molecules up to 1 kb in length through the annealing and extension of commercially-available synthetic oligos.
These products may then be joined using a One-Step Isothermal Assembly reaction to form DNA molecules up to 10 kb in length, or cloned directly into
expression vectors, attB integration vectors, or other vehicles. We have developed a system that permits production of complex RNAi constructs through in
vitro DNA synthesis. We can readily create a double-shmiR (short hairpin micro-RNA) construct that enables us to test the combinatorial effects of multiple
gene knockdown in a high-throughput manner in Drosophila. These constructs, based on two shmiRs in a single transcript, are functional in Drosophila. We
are beginning to test combinations of candidate "cancer genes" identified through cancer sequencing projects using this method.
886A
Design and validation of novel gene regulatory functions by perturbing existing cis-regulatory elements.
Ben Vincent
1
, Tara Martin
1
, Garth Isley
2
,
Zeba Wunderlich
1
, Meghan Bragdon
1
, Kelly Eckenrode
1
, Nick Luscombe
2
, Angela DePace
1
. 1) Department of Systems Biology, Harvard Medical School,
Boston, MA; 2) European Bioinformatics Institute, Cambridge UK.
Transcriptional regulation is critical for the specification of individual cell types and the control of complex cellular processes. In animals, cis-regulatory
elements (CREs) encode gene regulatory functions (GRFs) that integrate inputs from upstream regulators and output precise spatio-temporal expression
patterns. Despite recent advances in the identification of individual CREs and their component transcription factor binding sites, it remains difficult to
predict how regulatory sequence variants will affect GRF output. To address this problem, we build new GRFs by perturbing existing CRE sequences. The
even-skipped
(
eve
) locus is an ideal system for this approach: its expression pattern is produced by a set of known CREs; the expression patterns of upstream
regulators have been measured at cellular resolution; and the binding preferences of these regulators are known. By fitting functions that relate the
concentration of upstream regulators to expression output, we predict novel GRFs that are feasible within this system. The parameters of these models reflect
the role of regulating TFs, and therefore predict how modifications to existing CRMs could produce new GRFs. We have predicted that it is possible to
construct a GRF which outputs stripe 2 and stripe 5 by modifying the minimal stripe 2 CRE. We have constructed transgenic lines containing perturbed
minimal CREs upstream of a LacZ reporter and have determined output at cellular resolution with fluorescent in situ hybridization and 2-photon imaging.
Our results suggest that the 7-stripe
eve
expression pattern may be encoded in many different ways, and we are exploring this flexibility by applying this
general approach to other novel GRFs.