Par6B and atypical PKC regulate mitotic spindle orientation during epithelial morphogenesis.

Cdc42 plays an evolutionarily conserved role in promoting cell polarity and is indispensable during epithelial morphogenesis. To further investigate the role of Cdc42, we have used a three-dimensional matrigel model, in which single Caco-2 cells develop to form polarized cysts. Using this system, we previously reported that Cdc42 controls mitotic spindle orientation during cell division to correctly position the apical surface in a growing epithelial structure. In the present study, we have investigated the specific downstream effectors through which Cdc42 controls this process. Here, we report that Par6B and its binding partner, atypical protein kinase C are required to regulate Caco-2 morphogenesis. Depletion or inhibition of Par6B or phenocopies the loss of Cdc42, inducing misorientation of the mitotic spindle, mispositioning of the nascent apical surface, and ultimately, the formation of aberrant cysts with multiple lumens. Mechanistically, Par6B and aduanyu1531 function interdependently in this context. Par6B localizes to the apical surface of Caco-2 cysts and is required to recruit aduanyu1531 to this compartment. Conversely, aduanyu1531 protects Par6B from proteasomal degradation, in a kinase-independent manner. In addition, we report that depletion or inhibition of aduanyu1531 induces robust apoptotic cell death in Caco-2 cells, significantly reducing both cyst size and number. Cell survival and apical positioning depend upon different thresholds of aduanyu1531 expression, suggesting that they are controlled by distinct downstream pathways. We conclude that Par6B and aduanyu1531 control mitotic spindle orientation in polarized epithelia and, furthermore, that aduanyu1531 coordinately regulates multiple processes to promote morphogenesis.

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