p115RhoGEF activates RhoA to support tight junction maintenance and remodeling.

In vertebrates, epithelial cell-cell junctions must rapidly remodel to maintain barrier function as cells undergo dynamic shape-change events. Consequently, localized leaks sometimes arise within the tight junction (TJ) barrier, which are repaired by short-lived activations of RhoA, called "Rho flares." However, how RhoA is activated at leak sites remains unknown. Here we asked which guanine nucleotide exchange factor (GEF) localizes to TJs to initiate Rho activity at Rho flares. We find that p115RhoGEF locally activates Rho flares at sites of TJ loss. Knockdown of p115RhoGEF leads to diminished Rho flare intensity and impaired TJ remodeling. p115RhoGEF knockdown also decreases junctional active RhoA levels, thus compromising the apical actomyosin array and junctional complex. Furthermore, p115RhoGEF is necessary to promote local leak repair to maintain TJ barrier function. In all, our work demonstrates a central role for p115RhoGEF in activating junctional RhoA to preserve barrier function and direct local TJ remodeling.

SNAP- and Halo-tagging and dye introduction protocol for live microscopy in Xenopus embryos.

Traditional fluorescent proteins exhibit limitations in brightness and photostability that hinder optimal characterization of the dynamic cellular behavior of proteins of interest. SNAP- and Halo-tagging are alternatives to traditional fluorescent protein tagging utilizing bright, stable chemical dyes, which may improve signal-to-noise ratio. However, there has been limited use of this approach in vivo in developing organisms. Here, we present a protocol for implementing SNAP- and Halo-tagging in gastrula-stage Xenopus laevis embryos for live confocal microscopy. For complete details on the use and execution of this protocol, please refer to Varadarajan et al. (2022).

Closing the gap: Tricellulin/α-catenin interaction maintains epithelial integrity at vertices.

Tricellular junctions play a critical role in regulating epithelial barrier function. In this issue, Cho et al. (2022. J. Cell Biol.https://doi.org/10.1083/jcb.202009037) demonstrate a novel interaction between tricellulin and α-catenin, which connects tricellular junctions to the actomyosin cytoskeleton, thus supporting the epithelial barrier at cell vertices.