Negative feedback regulation in Drosophila dorsal-ventral patterning
Authors: Allison Schloop 1; Gregory Reeves 2
Affiliations: 1) NC State University, Raleigh, NC; 2) Texas A&M University, College Station, TX
Keywords: a. axis specification; b. live imaging
Development of an organism is dependent upon proper regulation of gene expression. Initiation of gene expression often relies on long-range signals referred to as morphogens; these morphogens form concentration gradients that aid in specific activation of genes responsible for proper body patterning. In Drosophila, one such morphogen is Dorsal (Dl), a transcription factor that helps with patterning of the dorsal-ventral (DV) axis in the early embryo. The impact of Dl is further refined by gene regulatory loops that help to control the dynamics of the Dl gradient. One regulatory loop of interest is the negative feedback loop with Cactus (Cact). Cact is initially bound to Dl, sequestering it to the cytoplasm, but Toll signaling on the ventral side of the cell degrades Cact and allows Dl to enter the nucleus. There, Dl can activate target genes, one of which is Cact, suggesting that Dl may regulate its own inhibition.
Our work currently focuses on establishing a system through which Cact can be examined in live embryos. Protein expression and use during development is very rapid; the turnover of Cact happens too quickly for standard live imaging techniques, like fluorescent protein fusions. Fluorescent proteins like GFP do not have enough time to mature and fluoresce before the associated protein is degraded. To work around this, we have utilized LlamaTags (Bothma et al. 2018) to image the dynamics of Cactus. We show that Cactus, while predominantly cytoplasmic, is also in the nucleus and shows a recoverable pattern after photobleaching. This provides initial evidence for an identifiable role of nuclear Cactus.