Numb Localizes at Endosomes and Controls the Endosomal Sorting of Notch after Asymmetric Division in Drosophila

Abstract

Numb acts as a cell-fate determinant during asymmetric and stem cell divisions in both vertebrates and invertebrates [1, 2]. In Drosophila, Numb is unequally segregated in asymmetrically dividing sensory organ precursor cells (SOPs). Numb is inherited by the pIIb cell (Notch OFF) and is absent from the pIIa cell (Notch ON) [3, 4]. Numb is required to establish directional Notch signaling during cytokinesis [3, 5-7]. Using real-time imaging of a functional GFP-tagged Numb, we show that Numb relocalizes during cytokinesis from the basal cortex of pIIb to subapical endosomes. This relocalization appeared to depend on its interaction with the α-adaptin [8, 9]. Live imaging of Sanpodo (Spdo), a membrane protein interacting with Numb and regulating the trafficking of Notch [6, 7, 10-15], revealed that Spdo is internalized during cytokinesis and coaccumulates with Numb in pIIb endosomes. Using a GFP-tagged Notch [6], we found that Notch coaccumulates with Spdo in a Numb-dependent manner in these pIIb endosomes. Numb was, however, dispensable for the internalization of Notch and Spdo. We propose that Numb interacts with internalized Spdo-Notch oligomers at sorting endosomes and inhibits the recycling of Notch, thereby creating an asymmetry in Notch distribution along the pIIa-pIIb interface and regulating binary fate choice.

Inhibition of notch signaling pathway prevents cholestatic liver fibrosis by decreasing the differentiation of hepatic progenitor cells into cholangiocytes

“…30 Numb is a cell-fate determinant that is localized to endosomes, controlling cell-fate choices by asymmetrically partitioning at mitosis and antagonizing Notch family plasma membrane receptor activity. [32][33][34] According to our results, Notch receptors Notch-2, -3, -4, and Notch ligand Jagged-1, -2, Deltal-3, as well as RBP-Jκ mRNA and protein expression significantly increased after BDL. So, Notch signaling enhanced bile duct proliferation.…”

Inhibition of Notch recycling by Numb: Relevance and mechanism(s)

“…5 These observations indicated that Numb regulates the endosomal trafficking of Notch during cytokinesis, and suggested that Numb breaks the symmetry of Notch distribution along the cytokinetic furrow between pIIa and pIIb, hence leading to directional signaling and binary fate decision.…”

“…4,6 Live imaging analysis of a functional Sanpodo-GFP protein combined with antibody uptake assays revealed that Notch and Sanpodo are co-internalized during cytokinesis, suggesting that Notch is endocytosed as Notch-Sanpodo oligomers. 4,5 Since Numb regulates the endosomal accumulation of Notch and Sanpodo in pIIb endosomes, it is likely that Numb somehow regulates the accumulation of Notch in sorting endosomes through its interaction with Sanpodo. Since Numb physically interacts with the AP-2 complex, one possibility would be that Numb regulates the AP-2-dependent internalization of Notch-Sanpodo oligomers.…”

“…7 However, Numb was found to be dispensable for the internalization of both Notch and Sanpodo, indicating that Numb does not inhibit Notch by promoting its internalization from the pIIb plasma membrane. 5,8 So, how does Numb regulate the trafficking of Notch-Sanpodo oligomers? Using a functional Numb-GFP protein, we showed that Numb relocalized during cytokinesis from the basal anterior cortex to the apical sorting endosomes that contain internalized Notch and Sanpodo.…”

Endosomal sorting of Notch receptors through COMMD9-dependent pathways modulates Notch signaling

“…Notch is constantly endocytosed (McGill et al, 2009), and the signaling outcome arising from this internalized pool of Notch is carefully controlled by several factors. On the one hand, the protein Numb is a conserved inhibitor of the pathway that acts at this level (Cotton et al, 2013;Couturier et al, 2013Couturier et al, , 2014Reichardt and Knoblich, 2013). This factor interacts with components of the AP-2 complex and prevents endosome-to-plasma membrane recycling of Notch; in addition, Numb also promotes the lysosomal degradation of the receptor (McGill et al, 2009).…”