Department of Zoology, University of Cambridge, Cambridge, UK.
Université de Paris, Centre National de la Recherche Scientifique, Institut Jacques Monod, Paris, France.
J Cell Biol. 2021 Aug 2;220(8). doi: 10.1083/jcb.202010020. Epub 2021 May 27.
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.
γ-微管蛋白环复合物（γ-TuRCs）可起始微管的形成。在有丝分裂细胞中，它们通过与 γ-TuRC 连接蛋白的 N 端 CM1 结构域结合被招募到中心体，包括果蝇中心体蛋白（Cnn）。这种结合促进了微管的起始，并且仅局限于有丝分裂细胞的中心体，但调节结合的机制尚不清楚。在这里，我们鉴定了一个在 Cnn 的中心体同工型（Cnn-C）中特有的极端 N 端 CM1 自身抑制（CAI）结构域，它抑制了 γ-TuRC 的结合。在去除 CAI 结构域或添加磷酸模拟突变后，会发生强烈的结合，这表明磷酸化有助于解除抑制。我们表明，Cnn 与 γ-TuRC 结合的调节是同工型特异性的，并且结合的失调可能导致细胞分裂过程中胞质微管的异位和主要缺陷。我们还发现人类 CDK5RAP2 被自身抑制与 γ-TuRC 结合，表明这种抑制在物种间是保守的。总的来说，我们的研究结果阐明了 CM1 结构域与 γ-TuRC 结合的调节方式和原因。
