Zonula occludens (ZO)‐1 is required for epithelial morphogenesis and lumen development

Abstract

Epithelial morphogenesis involves establishment of cell polarity and formation of the tight and adherens junctions between adjacent cells. However, contributions of tight junctions to epithelial morphogenesis remain poorly understood. Studies of epithelia lacking the scaffold‐like tight junction protein ZO‐1 in 2D culture have identified reduced barrier function with few other defects, likely due to overlapping functions of ZO‐2. To address the role of ZO‐1 in 3D morphogenesis, we compared growth of control and ZO‐1 knockdown (kd) MDCK cells in 3D collagen gels. Over 90% of cysts formed by shRNA control MDCK displayed a single lumen phenotype, while >;90% of ZO‐1 kd cysts had multiple lumina. Consistent with the increased cellular mass required, ZO‐1 kd MDCK displayed increased proliferation (apoptosis was similar to controls). The multi‐lumen phenotype was not due to aberrant cellular polarization, as claudin‐2 (tight junction), E‐cadherin (adherens junction and basolateral membrane), and gp135 (apical membrane) were normally distributed in ZO‐1 kd cysts. Expression of ZO‐1 SH3‐GuK or ABR (actin binding region) domains as free proteins in wildtype MDCK resulted in multi‐lumen cysts similar to ZO‐1 kd. These data suggest that both SH3‐ GuK‐ and ABR‐mediated interactions are essential for morphogenesis of epithelial lumina via mechanisms that do not involve cell polarity and cannot be complemented by ZO‐2.