Project Summary
Current approaches to designing and constructing synthetic gene circuits have reached a
dilemma due to the substantial heterogeneity driven by circuit-host interactions, especially for
large-scale gene circuits. The conventional trial-and-error iteration approach on synthetic gene
circuit development is regarded as inefficient since the assembled gene circuits often are
susceptible to experimental conditions. One fundamental reason is that the heterogeneity driven
by circuit-host interactions become significant with the increase of the number of components in
gene circuits but are often neglected. Moreover, the lack of quantitative frameworks for quantifying,
characterizing, and controlling heterogeneity in the host-aware synthetic gene circuits impedes
the progress in the field. My laboratory has been focusing on dissecting the mechanisms of how
the circuit-host mutual interactions affect the gene circuit functions and developing control
strategies targeting circuit-host interactions to optimize engineered synthetic gene circuits.
Recently we found a topology-dependent interference of synthetic gene circuit function by growth
feedback, which was published in Nature Chemical Biology. We also found winner-takes-all
resource competition that redirected cascading cell fate transitions, which is in revision to Nature
Communication. In the proposed projects, we will establish experimental and computational
frameworks to quantify, characterize, and control the gene expression heterogeneity in the host-
aware synthetic gene circuits. The heterogeneity can result from stochastic cellular resource
allocation, stochastic biochemical reactions in gene circuits, and stochastic cell divisions. These
heterogeneities are intertwined due to the complex interactions between the gene circuits and the
host organisms, creating another layer of challenge and complexity to engineering robust gene
circuits. We will integrate a microfluidics system for time-lapse live-cell analysis, a Turbidostat
platform with Python-based easy-to-use web interface for accurate growth rate control and
automatic yet remotely-controllable in-situ fluorescence measurement, and hybrid agent-based
modeling algorithms for stochastic simulation of all the single cells in the bacterial community to
characterize the heterogeneity from various noise sources in the host-aware synthetic gene
circuits. I have built up my research group with all the necessary expertise and capabilities to
complete the proposed projects. This work will provide a systematic in-depth mechanical
understanding of the heterogeneity driven by circuit-host interactions, and will greatly help us to
rationally design and control the synthetic gene circuits for sophisticated clinical applications in a
real-world environment, such as bacterial infection and tumor microenvironments.