Rab35 GTPase couples cell division with initiation of epithelial apico-basal polarity and lumen opening

Klinkert, K., Rocancourt, M., Houdusse, A. & Echard, A.

Nature communications 7 (2016)

Speaker: Jia-Wen Chen (陳佳文)                                        Time: 14:00~15:00, May 03, 2017

Commentator: Yang-Kao Wang. Ph.D. (王仰高老師)         Place: Room 601

Abstract:

Many epithelial organs are composed of a polarized cell monolayer surrounding a central apical lumen. Dog renal Madin-Darby Canine Kidney (MDCK) cells¹ are widely used to study epithelial organs as they have clear apico-basolateral polarity, well defined cell junctions, rapid growth rate, and can polarize in 2D and 3D cell culture under confocal microscopy. The authors previously identified Rab35 as a unique small GTPase presented at the cleavage site that promotes cytokinetic abscission in HeLa cells. It is known that establishment and maintenance of apico-basal polarity in the epithelial organs tightly coupled with cell division, but the underlying molecular mechanisms are unclear. In this study, by using 3D cultures of MDCK cells (cysts), the authors found that Podocalyxin (PODXL, an apical transmembrane proteins) is a genuine Rab35-interacting protein, suggesting that Rab35 may play a critical role in early steps of cyst development. They investigated the role of Rab35 in apico-basal polarity establishment and PODXL-dependent lumen formation, and found that Rab35 depletion leads to a complete inversion of apico-basal polarity in cysts. They also found that inversion of polarity upon Rab35 depletion occurred in the initial cell–cell interface step of cyst development by using a PODXL mutant which is unable to interact with Rab35 and triggers the inversion of cyst polarity from this two-cell stage. Furthermore, the authors found that the active GTP-bound Rab35 tethers intracellular vesicles containing key apical markers (PODXL et al.) through physical interaction with the cytoplasmic tail of PODXL. Finally, the authors analyzed the consequences of delocalizing Rab35 at mitochondria during cystogenesis. They reveal that delocalizing GTP-bound Rab35 on mitochondria displaced PODXL-positive vesicles from the plasma membrane, which prevented the apical membrane initiation site (AMIS) formation and apico-basal polarity establishment. In summary, the authors disclosed a novel and unconventional mode of Rab-dependent vesicle targeting which triggers both initiation of apico-basal polarity and lumen opening at the centre of cysts.

References:

1.    Gaush, C. R., Hard, W. L. & Smith, T. F. Characterization of an established line of canine kidney cells (MDCK). Proc. Soc. Exp. Biol. Med. 122, 931-935 (1966).