The Na/K ATPase pump, a signaling switch between life and death

This thesis is the first in-depth investigation into the roles of Na/K ATPase in assembly and maintenance of septate junctions (SJ) and tricellular junctions (TCJ). Together, these domains block the flow of fluids or pathogens across an epithelia. Loss of either domain (SJ or TCJ) leads to a loss of the permeability barriers. These domains contain large protein complexes that include both the alpha and beta subunits of Na/K ATPase of which the alpha subunit has the potential for scaffolding protein complexes and cell signaling. In cultured fibroblasts the alpha subunit binds and inhibits Src kinase and downstream signaling pathways, but the role in developing epithelia is unknown. Likewise how the Na/K ATPase interacts with other junctional components to establish, maintain and regulate SJs and TCJs is unknown. We examined the consequences of loss of Na/K ATPase in the simple epithelium of the Drosophila wing imaginal disc to understand the role of the pump in mediating cell signaling and scaffolding of SJ and TCJ components. Using RNAi-mediated knockdown of either Na/K ATPase subunit, we observed an increase in activation of the tyrosine kinases, Src and Abl, leading to JNK-mediated apoptosis and apoptosis-induced proliferation. The role of Na/K ATPase in regulating Src activation is conserved between vertebrates and invertebrates and functions to regulate epithelia in vivo. An additional finding was that RNAi knockdown of Na/K ATPase in the imaginal wing disc led to changes in SJ and TCJ proteins that were distinct from knockdown of another core protein, NeurexinIV. In particular the Na/K ATPase led to a specific increase of the TCJ proteins Discs-large and Gliotactin at the TCJ. Overexpression of Gliotactin leads to JNK-mediated apoptosis and cell spreading and these were suppressed by loss of Na/K ATPase but enhanced by loss of NrxIV. This suggests Na/K ATPase has a unique role in both regulation and maintenance of the TCJ. Overall our data support a role for Na/K ATPase both as a scaffolding protein that organizes a subset of SJ and TCJ proteins and as a signaling component in epithelial cell survival.Medicine, Faculty ofGraduat