(1000-08) Computer-aided chemical screening identifies SOFTI as a novel small molecule inhibitor of protein O-fucosyltransferase SPINDLY in Arabidopsis
Graduate student Stanford University and Carnegie Institute for Science Stanford, California
Body of Abstract: Protein O-glycosylation plays a crucial role in maintaining cellular homeostasis across different species. Two homologs of O-GlcNAc transferase, SPINDLY (SPY) and SECRET AGENT (SEC), mediate the addition of O-fucose and O-GlcNAc to Serine/Threonine residues of a variety of target proteins in Arabidopsis. Loss of both SPY and SEC causes lethality in Arabidopsis whereas mutants are unavailable in many crops. Therefore, chemical inhibitors of SPY and SEC are crucial for functional studies of O-glycosylation. Here, combining computational and experimental screening of chemical libraries, we identify a SPY O-fucosyltransferase inhibitor (SOFTI). Computational analyses predict that SOFTI binds to the GDP-fucose-binding pocket of SPY and competitively inhibits GDP-fucose binding. In vitro assays confirm that SOFTI interacts with SPY protein and inhibits its O-fucosyltransferase activity. SOFTI treatment of Arabidopsis seedlings decreases the O-fucosylation of the SPY and NEDD1 proteins and causes phenotypes similar to the spy mutants, including a reduced sensitivity to plant hormone cytokinin, enhanced root hair density, and a defect in sugar-dependent growth in the darkness. These results demonstrate that SOFTI is a specific SPY O-fucosyltransferase inhibitor and a valuable chemical tool for functional studies of O-fucosylation and growth management in agriculture.