DrosophiladTBCE recruits tubulin around chromatin to promote mitotic spindle assembly

Abstract

Proper assembly of mitotic spindles requires microtubule nucleation at centrosomes but also around chromatin. In this study, we reveal a novel mechanism by which an enrichment of tubulin in the nuclear space following nuclear envelope breakdown promotes nucleation of spindle microtubules. This event mediated by the tubulin-specific chaperone dTBCE, depends on its tubulin binding CAP-Gly motif and is regulated by Ran. Live imaging, proteomic and biochemical analyses suggest that dTBCE is enriched in the nucleus at nuclear envelope breakdown and interacts with nuclear pore proteins and the Ran machinery to create an environment that facilitates subsequent tubulin enrichment. We propose that dTBCEdependent increase in tubulin concentration in the nuclear space is an important mechanism for microtubule nucleation in organisms where compartmentalization prevents free diffusion of tubulin.

Live imaging of Drosophila melanogaster neural stem cells with photo-ablated centrosomes

“…Control of successful centrosome ablation must then be performed. Samples are immediately fixed following ablation and processed for immunofluorescences using anti-centrosomin and anti-tubulin antibodies ( Figure 2 A) following a method described before ( Gallaud et al., 2014 ; Métivier et al., 2020 ). The fixation method (see above) is time consuming and only one or two cells can be analyzed per brain.…”