Summary Background Formation of epithelial sheets requires that cell division occurs in the plane of the sheet. During mitosis, spindle poles align so the astral microtubules contact the lateral cortex. Confinement of the mammalian Pins protein to the lateral cortex is essential for this process. Defects in signaling through Cdc42 and atypical protein kinase C (aPKC) also cause spindle misorientation. When epithelial cysts are grown in 3D cultures, mis-orientation creates multiple lumens. Results We now show that silencing of the polarity protein Par3 causes spindle mis-orientation in MDCK cell cysts. Silencing of Par3 also disrupts aPKC association with the apical cortex, but expression of an apically-tethered aPKC rescues normal lumen formation. During mitosis, Pins is mislocalized to the apical surface in the absence of Par3, or by inhibition of aPKC. Active aPKC increases Pins phosphorylation on Ser401, which recruits 14-3-3 protein. 14-3-3 binding inhibits association of Pins with Gαi, through which Pins attaches to the cortex. A Pins S401A mutant mislocalizes over the cell cortex and causes spindle orientation and lumen defects. Conclusions The Par3/aPKC polarity proteins ensure correct spindle pole orientation during epithelial cell division by excluding Pins from the apical cortex. Apical aPKC phosphorylates Pins, which results in the recruitment of 14-3-3 and inhibition of binding to Gαi, so the Pins falls off the cortex. In the absence of a functional exclusion mechanism, astral microtubules can associate with Pins over the entire epithelial cortex, resulting in randomized spindle pole orientation.

Abbreviations used in this paper: aPKC, atypical protein kinase C; Crb3, Crumbs3; G, the alpha subunit of the heterotrimeric G proteins; Gi, the alpha subunit of adenylyl cyclase-inhibitory heterotrimeric G proteins; GoLoco motif, G i/o -Loco motif; LGN, mammalian homologue of Pins; MT, microtubule; NuMA, nuclear mitotic apparatus protein; Pins, partner of inscuteable.

Despite the longer operative time, LA results in less postoperative pain, faster postoperative rehabilitation, a shorter hospital stay, and fewer postoperative complications than OA. Therefore, LA is worth recommending as an effective and safe procedure for acute appendicitis.