The Transforming Rho Family GTPase Wrch-1 Disrupts Epithelial Cell Tight Junctions and Epithelial Morphogenesis

Abstract

Wrch-1, an atypical and transforming Rho GTPase, regulates cellular activities including proliferation and actin organization, but its functions and effectors remain poorly characterized. We show here that Wrch-1 distributes along the apical and basolateral membranes in MDCK cells and binds the cell polarity protein Par6 in a GTP-dependent manner. Activated Wrch-1 negatively regulates the kinetics of tight junction (TJ) assembly during epithelial cell polarization but has no detectable effect on overall cell polarity in confluent monolayers. It also causes a dramatic cytoskeletal reorganization and multilayering in cells grown in two-dimensional culture and disrupts cystogenesis of cells grown in three-dimensional (3D) culture. Similarly, short hairpin RNA-mediated knockdown of Wrch-1 perturbs cystogenesis in 3D culture, suggesting that tight regulation of Wrch-1 activity is necessary for normal epithelial morphogenesis. A weakly transforming effector domain mutant of activated Wrch-1 that inhibits Par6 binding abrogates the ability of Wrch-1 to disrupt TJ formation, actin organization, and epithelial morphogenesis. We hypothesize that Wrch-1-induced morphological and growth transformation may occur in part through Par6-mediated disruption of TJs and actin organization.

Rhou maintains the epithelial architecture and facilitates differentiation of the foregut endoderm

“…Consistent with the cell culture results, in Xenopus embryos, in which Rhou is expressed in migrating cranial neural crest cells, Rhou knockdown impairs cell migration (Fort et al, 2011). In MDCK (canine kidney epithelial) cells, overexpression of Rhou disrupts tight junction formation and the distribution of F-actin, and depletion of Rhou disrupts lumen formation in cysts (Brady et al, 2009). The findings of these studies suggest that Rhou plays a role in regulating epithelial cell shape and structure and modulating cellular behaviour.…”

“…Reduced Rhou activity affects cellular morphology and cytoskeletal organization in cultured cells Brady et al, 2009;Saras et al, 2004). Of particular note, Rhou knockdown in MDCK cells disrupts epithelial cyst formation in a three-dimensional matrix (Brady et al, 2009). These findings suggest that Rhou activity is essential for the maintenance of cellular and tissue architecture.…”

CCM3 signaling through sterile 20–like kinases plays an essential role during zebrafish cardiovascular development and cerebral cavernous malformations

“…Recent studies of endothelial cells lacking CCM1 or CCM2 (10,13,21) have demonstrated an increased formation of actin stress fibers and an increase in the activity of Rho, a small GTPase known to negatively regulate both endothelial and epithelial cell junctions (26)(27)(28)(29). We therefore next measured the effect of loss of CCM3 and the STKs on stress fiber formation and Rho activity.…”

Regulation and functions of RhoU and RhoV

“…By contrast, RhoV does not contain PRMs in its N-terminal extension. The sequences in RhoU are PAFPDR (residues 8-13), PPVPPRR (20)(21)(22)(23)(24)(25)(26) and PGEPGGR (36-42) (Figure 4). The N-terminal region of RhoU binds to various SH3-domain containing adaptor proteins, including Grb2, 5,14 Nck2 5,6 and phospholipase Cg.…”