Mini Spindles TOG1 Maintains Microtubule Polymerization Rates and Mitotic Spindle Formation

Abstract

Microtubules (MTs) are dynamic polymers that regulate cellular processes including intracellular trafficking and mitotic spindle formation. MT-associated proteins (MAPs) control MT dynamics in space and time in order to ensure proper cellular function. Members of the XMAP215 protein family are conserved MAPs that utilize arrays of αβ-tubulin-binding tumor overexpressed gene (TOG) domains to promote rapid MT polymerization. Higher eukaryotic XMAP215 proteins possess a pentameric TOG array that is structurally diverse despite being positionally conserved across species. Previous studies show that a TOG1-2-5 array is sufficient to promote endogenous MT polymerization rates. Furthermore, multiple studies of XMAP215 family members demonstrate that removing TOG1 and TOG2 from the pentameric array completely abrogates MT growth rates. However, the significance of TOG1 alone in regulating MT dynamics remains uncertain. We investigated TOG1’s role in promoting MT polymerization and mitotic spindle formation using the Drosophila melanogaster (D.m.) XMAP215 family member Mini Spindles (Msps) as a model. Endogenous Msps was depleted from D.m. S2 cells using dsRNA and a Msps construct with TOG1 deleted (ΔTOG1) from the pentameric TOG domain array was expressed. ΔTOG1-expressing cells exhibited dramatically reduced MT polymerization rates in comparison to control cells. Furthermore, ΔTOG1 expression yielded a significantly decreased incidence of normal bipolar spindle phenotypes. These data suggest that TOG1 is critical for proper XMAP215-mediated MT dynamics and mitotic spindle formation. Together, these findings lead to a more comprehensive model of how distinct TOG domains contribute to XMAP215 function. These results also provide a foundation on which to study other TOG-domain containing MAPs.Bachelor of Scienc