γ-Tubulin ring complexes (γ-TuRCs) nucleate microtubules. They are recruited to centrosomes in dividing cells via binding to N-terminal CM1 domains within γ-TuRC–tethering proteins, including Drosophila Centrosomin (Cnn). Binding promotes microtubule nucleation and is restricted to centrosomes in dividing cells, but the mechanism regulating binding remains unknown. Here, we identify an extreme N-terminal CM1 autoinhibition (CAI) domain found specifically within the centrosomal isoform of Cnn (Cnn-C) that inhibits γ-TuRC binding. Robust binding occurs after removal of the CAI domain or with the addition of phosphomimetic mutations, suggesting that phosphorylation helps relieve inhibition. We show that regulation of Cnn binding to γ-TuRCs is isoform specific and that misregulation of binding can result in ectopic cytosolic microtubules and major defects during cell division. We also find that human CDK5RAP2 is autoinhibited from binding γ-TuRCs, suggesting conservation across species. Overall, our results shed light on how and why CM1 domain binding to γ-TuRCs is regulated.
Bibliographical noteFunding Information:
We thank Jordan Raff for sharing reagents, Berthold Hedwig and Steve Rogers for help with needle pulling, Jens Lüders for sharing the pCMV-GFP vector, and Matt Castle for guidance on statistical analysis. We thank other members of the Conduit laboratory for their invaluable input. The work benefited from the Imaging Facility, Department of Zoology, University of Cambridge, supported by Matt Wayland and a Sir Isaac Newton Trust Research Grant (18.07ii[c]) from the ImagoSeine at the Institut Jacques-Monod, and from use of the Cambridge Centre for Proteomics Core Facility. For the purpose of open access, the author has applied a CC-BY public copyright license to any Author Accepted Manuscript (AAM) version arising from this submission.
This research was supported by a Wellcome Trust and Royal Society Sir Henry Dale fellowship (105653/Z/14/Z) and by IdEx Universitéde Paris (grant ANR-18-IDEX-0001 to P.T. Conduit), by a Glover Fund research fellowship from Clare College, University of Cambridge (to C.A. Tovey), by an Association pour la Recherche sur le Cancer grant (PJA 20181208148 to A. Guichet), and by the Centre National de la Recherche Scientifique. The authors declare no competing financial interests.
© 2021 Tovey et al.
- Cell cycle and division