ZO-1 regulates Hippo-independent YAP activity and cell proliferation via a GEF-H1- and TBK1-regulated mechanosensitive signalling network

Tight junctions are a barrier-forming cell-cell adhesion complex and have been proposed to regulate cell proliferation. However, the underlying mechanisms are not well understood. Here, we used cells deficient in the junction scaffold ZO-1 alone or together with its paralog ZO-2, which disrupts the junctional barrier. We found that ZO-1 knockout increased cell proliferation, loss of cell density-dependent proliferation control, and promoted cell death. These phenotypes were enhanced by double ZO-1/ZO-2 knockout. Increased proliferation was dependent on YAP and ZONAB, two transcriptional regulators. ZO-1 knockout stimulated YAP nuclear translocation and activity without changes in Hippo-dependent phosphorylation. Knockout promoted TANK-binding Kinase 1 (TBK1) activation and increased expression of the RhoA activator GEF-H1. Knockdown of ZO-3, another paralog interacting with ZO1, was sufficient to induce GEF-H1 expression and YAP activity. GEF-H1, TBK1, and mechanotransduction at focal adhesions were required for YAP/TEAD activation in ZO-1-deficient cells. Thus, ZO-1 controls cell proliferation and Hippo-independent YAP activity by activating a GEF-H1- and TBK1-regulated mechanosensitive signalling network.