A Perturbation Approach to Quantitatively Decipher Transcription Factor Kinetic Response to Target Gene Output In Vivo

In early metazoan development, cells must decide their fate to create robust spatial gene expression patterns leading to proper body segmentation. In the fruit fly, these early processes are tightly regulated by maternal and zygotic transcription factors (TFs). One of the classical maternal TF, Bicoid (Bcd), is known to organize spatial and temporal patterns of target genes called gap genes. Previous attempts by genetic and optogenetic approaches had limited temporal control to distinguish how TFs directly or indirectly regulate gap gene expression in space and time. Here we present a light-inducible system to regulate Bcd TF concentration in living fly embryos on second time-scale. Precise temporal optogenetic perturbation allows us to quantify the response time of gene activation to input factor perturbation in living embryos. We show how nuclear concentration and TF residence times affect the kinetic rates and expression levels of downstream gene targets. Furthermore, we explore how cells utilize differential levels of TF to make cell fate decisions. Our synthetic approach to modulate TF concentration while simultaneously recording transcriptional gene output in real-time presents a powerful tool to study transcriptional dynamics during development.