Regulation of Cytokinesis by Exocyst Subunit SEC6 and KEULE in Arabidopsis thaliana

ABSTRACT Proper vesicle tethering and membrane fusion at the cell plate are essential for cytokinesis. Both the vesicle tethering complex exocyst and membrane fusion regulator KEULE were shown to function in cell plate formation, but the exact mechanisms still remain to be explored. In this study, using yeast two-hybrid (Y-2-H) assay, we found that SEC6 interacted with KEULE, and that a small portion of C-terminal region of KEULE was required for the interaction. The direct SEC6–KEULE interaction was supported by further studies using in vitro pull-down assay, immunoprecipitation, and in vivo bimolecular florescence complementation (BIFC) microscopy. sec6 mutants were male gametophytic lethal as reported; however, pollen-rescued sec6 mutants ( PRsec6 ) displayed cytokinesis defects in the embryonic cells and later in the leaf pavement cells and the guard cells. SEC6 and KEULE proteins were co-localized to the cell plate during cytokinesis in transgenic Arabidopsis . Furthermore, only SEC6 but not other exocyst subunits located in the cell plate interacted with KEULE in vitro . These results demonstrated that, like KEULE, SEC6 plays a physiological role in cytokinesis, and the SEC6–KEULE interaction may serve as a novel molecular linkage between arriving vesicles and membrane fusion machinery or directly regulate membrane fusion during cell plate formation in plants. SUMMARY SEC6 and KEULE interact directly, and may coordinate cytokinesis in Arabidopsis thaliana . The interaction may serve as a novel molecular linkage between arriving vesicles and membrane fusion machinery or directly regulate membrane fusion during cell plate formation in plants.