ARHGAP12 suppresses F-actin assembly to control epithelial tight junction mechanics and paracellular leak pathway permeability

Tight junctions (TJs) control the paracellular transport of ions, solutes, and macromolecules across epithelial barriers. There is evidence that claudin-based ion transport (the pore pathway) and the paracellular transport of macromolecules (the leak pathway) are controlled independently. However, how leak pathway flux is regulated is unclear. Here, we have identified the Cdc42/Rac GTPase-activating protein ARHGAP12 as a specific activator of the leak pathway. ARHGAP12 is recruited to TJs via an interaction between its Src homology (SH3) domain and the TJ protein ZO-2 to suppress N-WASP-mediated F-actin assembly. This dampens junctional tension and promotes the paracellular transport of macromolecules without affecting ion flux. Mechanistically, we demonstrate that the ARHGAP12 tandem WW domain interacts directly with PPxR motifs in the proline-rich domain of N-WASP and thereby attenuates SH3-domain-mediated N-WASP oligomerization and Arp2/3-driven F-actin assembly. Collectively, our data indicate that branched F-actin networks regulate junctional tension to fine-tune the TJ leak pathway.