The correct establishment and maintenance of unidirectional Notch signaling are 
critical for the homeostasis of various stem cell lineages. However, the 
molecular mechanisms that prevent cell-autonomous ectopic Notch signaling 
activation and deleterious cell fate decisions remain unclear. Here we show that 
the retromer complex directly and specifically regulates Notch receptor 
retrograde trafficking in Drosophila neuroblast lineages to ensure the 
unidirectional Notch signaling from neural progenitors to neuroblasts. Notch 
polyubiquitination mediated by E3 ubiquitin ligase Itch/Su(dx) is inherently 
inefficient within neural progenitors, relying on retromer-mediated trafficking 
to avoid aberrant endosomal accumulation of Notch and cell-autonomous signaling 
activation. Upon retromer dysfunction, hypo-ubiquitinated Notch accumulates in 
Rab7+ enlarged endosomes, where it is ectopically processed and activated in a 
ligand-dependent manner, causing progenitor-originated tumorigenesis. Our 
results therefore unveil a safeguard mechanism whereby retromer retrieves 
potentially harmful Notch receptors in a timely manner to prevent aberrant Notch 
activation-induced neural progenitor dedifferentiation and brain tumor 
formation.