The Drosophila Dgt6 Protein Interacts with Msps/ XMAP215 to Promote Kinetochore-Driven MT Formation.

We analyzed the mitotic role of Dgt6, a member the Drosophila augmin complex required for spindle formation. Dgt6 exhibits a highly dynamic localization pattern; it is enriched at the center of prophase asters, at the kinetochore fibers (k-fibers) and at the central spindle. RNAi-mediated knockdown of Dgt6 resulted in spindles with a low MT density lacking robust k-fibers. These spindles were mostly unable to mediate sister chromatid separation but, nonetheless, they managed to elongate and assume ana-telophase configurations. To define the role of Dgt6 in kMT formation, we analyzed MT regrowth in prometaphases/metaphases following cold-induced MT depolymerization. In control cells returned to room temperature for 15-30 seconds, we observed short, randomly oriented k-fibers enriched in both Dgt6 and Msps. In Dgt6-depleted cells, k-fiber formation after return to room temperature was severely impaired, suggesting that the aberrant spindles observed in these cells are generated by a primary defect in kinetochore-driven k-fiber assembly. We also observed that Msps and D-TACC fail to accumulate at the centrosomes in Dgt6 RNAi cells. Consistent with these results, we found that Dgt6 co-immunoprecipitates with Msps, D- TACC and the kinetochore component Ndc80/Hec1. Collectively, our results suggest a model on the role of Dgt6 in spindle assembly. We propose that Dgt6 binds dynamic MTs and recruits Msps/XMAP215, which in turn promotes rapid MT polymerization by catalyzing the addition of tubulin dimers to the growing plus ends.