RhoA‐dependent tight junction regulation is associated with dysregulated Akt and ERK signaling and increased claudin‐2 expression

Previous studies have demonstrated that expression of either constitutively‐active (CA) or dominant negative (DN) RhoA reduced tight junction barrier function in cultured MDCK monolayers (Jou et al. 1998). However, the mechanisms by which RhoA disruption causes barrier loss have not been defined. The aims of this study were to determine whether RhoA regulates barrier function in mammary epithelia and to identify the molecular events responsible. V14 and N19 (CA and DN, respectively) RhoA were expressed in EpH4 mammary epithelial cells. RhoA (CA or DN) expression in confluent monolayers caused a dose‐dependent loss of transepithelial electrical resistance. Neither trafficking of ZO‐1 and occludin to the tight junction nor cell viability were affected by CA or DN RhoA expression, but western blots showed that ERK and Akt were phosphorylated (consistent with activation) after either CA or DN RhoA expression. Transcript content of several claudins was altered after CA or DN RhoA expression. Most notably, claudin‐2 mRNA and protein expression were increased following either CA or DN RhoA expression. These data demonstrate that RhoA regulates the barrier in mammary epithelia and suggest that increased claudin‐2 expression contributes to RhoA‐mediated tight junction regulation. Supported by Charite Universitaetsmedizin Berlin.