Global maps of transcription factor properties reveal threshold-based formation of DNA-bound and mobile clusters

The relationship between bulk transcription factor concentration and DNA binding has been a central question in gene regulation for decades. Recent studies propose that DNA-bound transcription factor hubs, or clusters, aid in fast and precise transcriptional interpretation. Using live imaging techniques, we quantify the concentration, binding, and mobility of the morphogen Dorsal (Dl), both in bulk and in clusters, in Drosophila blastoderm embryo. Our experiments encompass multiple length and time scales, allowing us to obtain a nucleus-wide view of the mechanism connecting hub formation to bulk Dl concentration. Our results show that previously unobserved, slowly-moving clusters of Dl are present, in addition to the expected populations of freely mobile and DNA-bound Dl. Furthermore, both mobile clusters and DNA-bound Dl appear only once a threshold concentration in the nucleus is surpassed, a behavior consistent with liquid-liquid phase separation. Thus, our work elucidates how bulk transcription factor concentration dictates the formation and spatiotemporal changes of different populations needed for gene regulation.