@article {Kaur114686, author = {Prameet Kaur and Timothy E. Saunders and Nicholas S. Tolwinski}, title = {Coupling optogenetics and light-sheet microscopy to study signal transduction in vivo}, elocation-id = {114686}, year = {2017}, doi = {10.1101/114686}, publisher = {Cold Spring Harbor Laboratory}, abstract = {Optogenetics allows precise, fast and reversible intervention in biological processes. Coupled with light-sheet microscopy, this approach allows unparalleled insight into the regulation of signaling pathways and cellular processes from a spatial and temporal perspective in vivo. To develop this method, we investigated the spatial and temporal regulation of canonical Wnt signaling during anterior-posterior patterning of the Drosophila embryonic epidermis. Cryptochrome C (Cry2) from Arabidopsis Thaliana was fused to mCherry fluorescent protein and Drosophila β{\textendash}catenin to form an easy to visualize optogenetic switch. Blue light illumination caused reversible oligomerization of the fusion protein and inhibited downstream Wnt signaling in vitro and in vivo. Temporal inactivation of β{\textendash}catenin showed that Wnt signaling is required not only for Drosophila patterning formation, but also for maintenance throughout development. By applying light-sheet microscopy to activate the pathway and then record subsequent behavior in toto, we show precise spatial regulation of Wnt signaling in vivo. We anticipate that this method will be easily extendable to other developmental signaling pathways and many other experimental systems.}, URL = {https://www.biorxiv.org/content/early/2017/03/08/114686}, eprint = {https://www.biorxiv.org/content/early/2017/03/08/114686.full.pdf}, journal = {bioRxiv} }