Quantitative measurement of synthetic repression curves reveals design challenges for genetic circuit engineering under growth arrest

Abstract

Despite the fact that microbes in natural environments spend most of their time in growth arrest, we understand little about how this physiological state affects the performance of engineered genetic circuits. Here, we measure repression curves from a library of genetic NOT gates at single-cell resolution in Escherichia coli under both active growth and growth arrest to systematically investigate how growth arrest affects circuit behavior. We find that the impact of growth arrest on circuit performance is almost entirely dominated by a single effect: a >100-fold reduction in unrepressed expression levels. Growth arrest caused gene expression noise to increase moderately and had only minimal impacts on the sensitivity and sharpness of the repression curves. Our work shows both that conventional genetic circuit design paradigms are currently insufficient to develop circuits that can function properly under growth arrest, but also that addressing the reduction in just a single performance parameter would be sufficient to resolve this problem. This work expands our understanding of bacterial gene regulation under growth arrest and lays the groundwork for new design paradigms that will be essential in ensuring the safe and reliable performance of synthetic biology systems in real-world environments. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC="FIGDIR/small/703179v1_ufig1.gif" ALT="Figure 1"> View larger version (14K): org.highwire.dtl.DTLVardef@104ecd2org.highwire.dtl.DTLVardef@57f992org.highwire.dtl.DTLVardef@1c215c2org.highwire.dtl.DTLVardef@122a8c5_HPS_FORMAT_FIGEXP M_FIG C_FIG