TACC3 regulates microtubule nucleation by affecting {gamma}-tubulin ring complexes [Gene Regulation]

September 22nd, 2014 by Singh, P., Thomas, G. E., Gireesh, K. K., Manna, T.

Centrosome-mediated microtubule nucleation is essential for spindle assembly during mitosis. Although γ-tubulin complexes have primarily been implicated in the nucleation process, details of the underlying mechanisms remain poorly understood. Here, we demonstrated that member of the human transforming acidic coiled-coil (TACC) protein family, TACC3, plays critical role in microtubule nucleation at the centrosome. In mitotic cells, TACC3 knock-down substantially affected the assembly of microtubules in the astral region and impaired microtubule nucleation at the centrosomes. The TACC3-depletion-induced mitotic phenotype was rescued by expression of the TACC3 C-terminus predominantly consisting of the TACC domain, suggesting that the TACC domain plays an important role in microtubule assembly. Consistently, experiments with the recombinant TACC domain of TACC3 demonstrated that this domain possesses intrinsic microtubule-nucleating activity. Co-immunoprecipitation and sedimentation experiments revealed that TACC3 mediates interactions with proteins of both the γ-tubulin ring complex (γ-TuRC) and the γ-tubulin small complex (γ-TuSC). Interestingly, TACC3 depletion resulted in reduced levels of γ-TuRC and increased levels of γ-TuSC, indicating that the assembly of γ-TuRC from γ-TuSC requires TACC3. Detailed analyses suggested that TACC3 facilitates the association of γ-TuSC-specific proteins with the proteins known to be involved in the assembly of γ-TuRC. Consistent with such a role for TACC3, the suppression of TACC3 disrupted localization of γ-TuRC proteins to the centrosome. Our findings reveal that TACC3 is involved in the regulation of microtubule nucleation at the centrosome and functions in the stabilization of the γ-tubulin ring complex assembly.