Curated Optogenetic Publication Database

Abstract: Gene expression is thought to be affected not only by the concentration of transcription factors (TFs) but also the dynamics of their nuclear translocation. Testing this hypothesis requires direct control of TF dynamics. Here, we engineer CLASP, an optogenetic tool for rapid and tunable translocation of a TF of interest. Using CLASP fused to Crz1, we observe that, for the same integrated concentration of nuclear TF over time, changing input dynamics changes target gene expression: pulsatile inputs yield higher expression than continuous inputs, or vice versa, depending on the target gene. Computational modeling reveals that a dose-response saturating at low TF input can yield higher gene expression for pulsatile versus continuous input, and that multi-state promoter activation can yield the opposite behavior. Our integrated tool development and modeling approach characterize promoter responses to Crz1 nuclear translocation dynamics, extracting quantitative features that may help explain the differential expression of target genes.

Abstract: In eukaryotes, protein kinase A (PKA) is a master regulator of cell proliferation and survival. The activity of PKA is subject to elaborate control and exhibits complex time dynamics. To probe the quantitative attributes of PKA dynamics in the yeast Saccharomyces cerevisiae, we developed an optogenetic strategy that uses a photoactivatable adenylate cyclase to achieve real-time regulation of cAMP and the PKA pathway. We capitalize on the precise and rapid control afforded by this optogenetic tool, together with quantitative computational modeling, to study the properties of feedback in the PKA signaling network and dissect the nonintuitive dynamic effects that ensue from perturbing its components. Our analyses reveal that negative feedback channeled through the Ras1/2 GTPase is delayed, pinpointing its time scale and its contribution to the dynamic features of the cAMP/PKA signaling network.

Abstract: We show that difficulties in regulating cellular behavior with synthetic biological circuits may be circumvented using in silico feedback control. By tracking a circuit's output in Saccharomyces cerevisiae in real time, we precisely control its behavior using an in silico feedback algorithm to compute regulatory inputs implemented through a genetically encoded light-responsive module. Moving control functions outside the cell should enable more sophisticated manipulation of cellular processes whenever real-time measurements of cellular variables are possible.