Molecular steps in TAO kinase regulation

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Despite their discovery over 20 years ago, TAO protein kinases (TAO1-3) remain understudied. In particular, we lack an understanding of the mechanisms regulating TAO activation, how they recognize substrates, and how these kinases contribute to key physiological processes. We originally identified TAOs through a large- scale effort to find membrane proximal components of MAPK cascades and showed that TAOs are MAP3Ks in the p38 pathway. Recent work now reveals that TAOs are essential for schistosome and viral infections and viral RNA processing and export from the nucleus. These findings suggest that TAOs are drug targets for a range of pathophysiological conditions and beg for a greater understanding of how their activities are controlled. Here, we propose to develop a paradigm for the biochemical regulation of TAO protein kinases through their integration with phosphoprotein phosphatase 1 (PP1). These results should advance future drug discovery efforts to provide new therapeutic entry points for treating a range of diverse pathological conditions. An obstacle in realizing their therapeutic potential is the limited knowledge of their regulation and partners. In searching for regulatory interactions, we found that TAOs directly bind PP1 and its R7 regulatory subunit. While PP1 dephosphorylates half the proteins in the cell, its activity is largely restricted within heteromeric complexes by regulatory subunits. Recent literature indicates that R7 maintains PP1 in an inactive state. We propose that TAOs modulate PP1 phosphatase activity via direct interactions and by R7 phosphorylation. The connection with PP1 offers TAOs wide opportunities to impact cellular control mechanisms. Our specific aims are to 1) determine how the TAO-PP1 complex regulates TAO activity; and 2) determine how TAO through R7 regulates PP1 activity. Biochemical and cell biological studies will take advantage of a model of a TAO2-PP1 complex based on our crystal structure of the TAO2 kinase domain that shows the PP1 binding motif on TAO. The relevance of this interaction is supported by our recent work revealing co-localization of TAO2 and PP1 in structures in the nucleus and the cytoplasm. Our extensive experience in identifying and characterizing TAOs, determining the structure of the TAO kinase domain, identifying chemically tractable inhibitors, and elucidating TAO-dependent pathways, since our discovery of these kinases, puts us in a unique position to determine biochemical processes that will provide a foundation for TAO kinases as subjects of drug development.