The Eyes Absent proteins (EYA1-4) are a biochemically unique group of tyrosine phosphatases known to be tumour-promoting across a range of cancer types. To date, the targets of EYA phosphatase activity remain largely uncharacterised. Here, we identify Polo-like kinase 1 (PLK1) as an interactor and phosphatase substrate of EYA4 and EYA1, with pY445 on PLK1 being the primary target site. Dephosphorylation of pY445 in the G2 phase of the cell cycle is required for centrosome maturation, PLK1 localization to centrosomes, and polo-box domain (PBD) dependent interactions between PLK1 and PLK1-activation complexes. Molecular dynamics simulations support the rationale that pY445 confers a structural impairment to PBD-substrate interactions that is relieved by EYA-mediated dephosphorylation. Depletion of EYA4 or EYA1, or chemical inhibition of EYA phosphatase activity, dramatically reduces PLK1 activation, causing mitotic defects and cell death. Overall, we have characterized a phosphotyrosine signalling network governing PLK1 and mitosis.

缺眼蛋白 （EYA1-4） 是一组生化上独特的酪氨酸磷酸酶，已知在一系列癌症类型中具有促进肿瘤的作用。迄今为止，EYA 磷酸酶活性的靶标在很大程度上仍未表征。在这里，我们确定 Polo 样激酶 1 （PLK1） 是 EYA4 和 EYA1 的相互作用物和磷酸酶底物，其中 PLK1 上的 pY445 是主要靶位点。pY445 在细胞周期的 G2 期去磷酸化是中心体成熟、PLK1 定位到中心体以及 PLK1 和 PLK1 激活复合物之间的 polo-box 结构域 （PBD） 依赖性相互作用所必需的。分子动力学模拟支持 pY445 对 PBD-底物相互作用造成结构损伤的基本原理，而 EYA 介导的去磷酸化可以缓解这种损伤。EYA4 或 EYA1 的耗竭或 EYA 磷酸酶活性的化学抑制会显著降低 PLK1 的激活，从而导致有丝分裂缺陷和细胞死亡。总体而言，我们已经表征了控制 PLK1 和有丝分裂的磷酸酪氨酸信号网络。