Notch is a conserved regulator of stem cells and differentiation. Upon activation by ligand, the Notch receptor is cleaved, releasing its intracellular domain (NICD) for assembly of a nuclear complex and transcriptional activation of downstream targets. Our work focuses on Notch signaling in the C. elegans germline stem cell niche, where well-defined signaling and receiving cells facilitate quantitative analyses. We recently reported that the transcriptional response to GLP-1/Notch activation occurs in a steep gradient across the germline stem cell pool (Lee et al., 2016). An outstanding question is whether this transcriptional gradient reflects a corresponding gradient in GLP-1/Notch activation. We are visualizing nuclear GLP-1/Notch NICD as a readout of GLP-1/Notch activation. Our reagents include MOS-SCI transgenes encoding a tagged GLP-1 receptor that rescues a
glp-1 null allele as well as CRISPR-inserted tags at the endogenous locus. To date, we have detected signal with Myc, V5, OLLAS and Halo tags. The signal is low, however, and we are working on methods to bring the signal into a quantifiable range without changing receptor abundance. Our progress will be reported at the meeting.