Stratum recruits Rab8 at Golgi exit sites to regulate the basolateral sorting of Notch and Sanpodo

Abstract

In , the sensory organ precursor (SOP or pI cell) divides asymmetrically to give birth to daughter cells, the fates of which are governed by the differential activation of the Notch pathway. Proteolytic activation of Notch induced by ligand is based on the correct polarized sorting and localization of the Notch ligand Delta, the Notch receptor and its trafficking partner Sanpodo (Spdo). Here, we have identified Stratum (Strat), a presumptive guanine nucleotide exchange factor for Rab GTPases, as a regulator of Notch activation. Loss of Strat causes cell fate transformations associated with an accumulation of Notch, Delta and Spdo in the-Golgi network (TGN), and an apical accumulation of Spdo. The mutant phenotype is rescued by the catalytically active as well as the wild-type form of Rab8, suggesting a chaperone function for Strat rather than that of exchange factor. Strat is required to localize Rab8 at the TGN, and phenocopies We propose that Strat and Rab8 act at the exit of the Golgi apparatus to regulate the sorting and the polarized distribution of Notch, Delta and Spdo.

Clathrin adaptor AP-1 and Stratum act in parallel pathways to control Notch activation inDrosophilaSensory Organ Precursor Cells

“…Therefore Su(H) can be used as a read-out of cell fate transformations to monitor the effect of individual or simultaneous impairment of Strat and AP-1 ( Fig.1). In agreement with previous studies, mild Notch GOF phenotypes revealed by an excess of Su(H) positive socket cells were observed in 6,7% and 7,5% of SO mutant for strat or depleted of AP-1, respectively (Bellec et al, 2018;Benhra et al, 2011). However simultaneous loss of AP-1 and Strat led to a much stronger Notch GOF phenotype (49% of transformed SOs, Fig.1B-C).…”

“…Indeed, we found that Numb and Neur localize asymmetrically during prometaphase in absence of Strat and AP-1 ( Fig.2A-B). Liveimaging of Numb::GFP crispr (Bellec et al, 2018) revealed that Numb is unequally partitioned in the anterior SOP daughter cell, in a similar manner as in the control situation ( Fig.2C-D).…”

“…To test if AP-1 and Strat function in the same pathway to regulate the activation of Notch signaling pathway, we induced clones of cells homozygote mutant for a null mutation of strat generated by CRISPR (Bellec et al, 2018) in which the µ subunit of the AP-1 complex (also known as AP-47) was silenced using previously characterized tools (Benhra et al, 2011).…”

“…To further investigate the effect of AP-1 and Strat on the apico-basal distribution of Notch receptors, we monitored the dynamics of Notch, with a GFP tag in the Notch intracellular domain (Notch::GFP crispr (Bellec et al, 2018), hereafter called NiGFP) throughout the asymmetric division of the SOP, identified by the nuclear marker Histone2B::RFP expressed under a minimal neur promoter (H2B::RFP, Fig.3). Previously, elegant work from F.…”

“…This recycling event relies on AP-1 activity (Cotton et al, 2013). We also reported that Stratum (Strat), a chaperone regulating Rab8 recruitment, controls the exit from the Golgi apparatus and the basolateral targeting of Notch, Delta and Spdo (Bellec et al, 2018). As for AP-1, loss of Strat leads to an enrichment in Notch and Spdo at the apical pole of SOP daughter cells associated with a mild Notch GOF phenotype.…”