An RNA-Based Negative Autoregulation Circuit

The central focus of synthetic biology is to develop programmable genetic regulatory systems to control gene expression. Recently, regulatory RNAs have emerged as powerful components of the synthetic biology toolbox for constructing genetic circuits. RNA-only circuits have the advantage of fast signal propagation that is set by the rapid degradation rate of RNA molecules. Negative autoregulation (NAR) as a network motif is an essential part of several designs of genetic oscillators. NARs can also decrease the response time of genetic circuits, and promote the robustness of steady state expression levels. Being able to engineer an RNA-only negative autoregulatory circuit expands the possibilities to engineer more sophisticated RNA-only circuitry. In this work, we used a model-guided design to construct an RNA negative autoregulatory circuit with transcriptional sRNA repressors. We showed that the RNA-only transcriptional negative autoregulator speeds circuit response time both in vivo and in ell free transcription and translation systems (TX-TL). We also used an effective model to describe the negative autoregulator’s dynamics in TX-TL, and parameterized unknown parameters to predict future designs. This work presents the first known RNA-only negative autoregulatory circuit, and broadens our ability to construct increasingly sophisticated RNA genetic circuitry.