Determinants in the LIN-12/Notch Intracellular Domain That Govern Its Activity and Stability During Caenorhabditis...

Upon ligand binding, the LIN-12/Notch intracellular domain is released from its transmembrane tether to function in a nuclear complex that activates transcription of target genes. During Caenorhabditis elegans vulval development, LIN-12/Notch is activated by ligand in two of six multipotential vulval precursor cells (VPCs), specifying the "secondary vulval fate" and descendants that contribute to the vulva. If LIN-12 is ectopically activated in other VPCs, they also adopt the secondary fate, dividing to produce extra vulval cells, resulting in a "Multivulva" phenotype. Here, we identify determinants in the LIN-12 intracellular domain ["LIN-12(intra)"] that govern its activity and stability during C. elegans vulval development; we assayed activity of mutant forms based on their ability to cause a Multivulva phenotype and stability using a GFP tag to visualize their accumulation. Our analysis has revealed that, while the ubiquitin ligase SEL-10/Fbw7 promotes LIN-12(intra) downregulation in VPCs, there is a distinct mechanism for downregulation of LIN-12(intra) in VPC descendants. Our analysis also revealed that LIN-12(intra) must be in the nuclear complex to be regulated appropriately in VPCs and their descendants, and that the structure or conformation of the carboxy-terminal region influences stability as well. Although activity and stability are generally well-correlated, exceptions where they are uncoupled suggest that there may be roles for the carboxy-terminal region and sel-10 that are independent of their roles in regulating LIN-12(intra) stability.