Mechanisms of Mitotic Spindle Orientation by Plexin-B2

Cells show certain asymmetries in morphology and molecular organization, a characteristic that is known as cell polarity. Polarity is generally regulated by protein complexes and Rho GTPases like Cdc42, Rho or Rac. Epithelial cells are polarized along the apicobasal axis, and this apicobasal polarity influences many cellular processes, including cell division. Polarized mitosis in epithelia is controlled by the orientation of the mitotic spindle. This is crucial in epithelial cells during development for a correct tissue morphogenesis, but also in the adult for maintenance of tissue homeostasis or damage repair. The orientation of the spindle is controlled by a protein complex that includes NuMA, which mediates pulling from the spindle poles and LGN, which links NuMA to the correct regions of the cell cortex. Semaphorin-Plexin signaling is a cell-cell communication pathway involved in many tissues and processes mainly through regulation of the cytoskeleton and adhesion. It has been shown that Plexin-B2 regulates mitotic spindle orientation in kidney epithelial cells and that this regulation is relevant for kidney morphogenesis and repair. However, the molecular mechanisms through which Plexin-B2 controls spindle orientation are still largely unclear. In this work, I demonstrate that Plexin-B2 localizes to cell-cell contacts in the kidney epithelium and in epithelial cell lines in 2D and 3D. Furthermore, I show that Plexin-B2 remains polarized during mitosis. I add that the basolateral localization of Plexin-B2 in the kidney is independent of its ligands, but depends on its intracellular juxtamembrane domain in 3D cultures. Furthermore, I show that this region contains a unique basolateral targeting motif present in all murine class B plexins and conserved in human Plexin-B2. Using CRISPR-Cas genome editing, I confirm that the deletion of Plexin-B2 impairs correct lumen formation in epithelial cell cysts. Additionally, I show that the lack of Plexin-B2 does not influence growth of these cells, but increases the ...