Division of Gene Regulation, Institute for Advanced Medical Research, Department of Obstetrics and Gynecology, School of Medicine, Keio University, Shinjuku-ku, Tokyo 160-8582, Japan.
J Cell Biol. 2012 May 28;197(5):625-41. doi: 10.1083/jcb.201110110.
In the mitotic exit network of budding yeast, Dbf2 kinase phosphorylates and regulates Cdc14 phosphatase. In contrast, no phosphatase substrates of LATS1/WARTS kinase, the mammalian equivalent of Dbf2, has been reported. To address this discrepancy, we performed phosphoproteomic screening using LATS1 kinase. Screening identified MYPT1 (myosin phosphatase-targeting subunit 1) as a new substrate for LATS1. LATS1 directly and preferentially phosphorylated serine 445 (S445) of MYPT1. An MYPT1 mutant (S445A) failed to dephosphorylate Thr 210 of PLK1 (pololike kinase 1), thereby activating PLK1. This suggests that LATS1 promotes MYPT1 to antagonize PLK1 activity. Consistent with this, LATS1-depleted HeLa cells or fibroblasts from LATS1 knockout mice showed increased PLK1 activity. We also found deoxyribonucleic acid (DNA) damage-induced LATS1 activation caused PLK1 suppression via the phosphorylation of MYPT1 S445. Furthermore, LATS1 knockdown cells showed reduced G2 checkpoint arrest after DNA damage. These results indicate that LATS1 phosphorylates a phosphatase as does the yeast Dbf2 and demonstrate a novel role of LATS1 in controlling PLK1 at the G2 DNA damage checkpoint.
在芽殖酵母的有丝分裂退出网络中，Dbf2 激酶磷酸化并调节 Cdc14 磷酸酶。相比之下，尚未报道哺乳动物 Dbf2 的等效物 LATS1/WARTS 激酶的磷酸酶底物。为了解决这一差异，我们使用 LATS1 激酶进行了磷酸蛋白质组学筛选。筛选鉴定出 MYPT1（肌球蛋白磷酸酶靶向亚基 1）是 LATS1 的新底物。LATS1 直接且优先磷酸化 MYPT1 的丝氨酸 445（S445）。MYPT1 突变体（S445A）未能去磷酸化 PLK1（polo 样激酶 1）的 Thr 210，从而激活 PLK1。这表明 LATS1 促进 MYPT1 拮抗 PLK1 活性。与此一致，LATS1 耗尽的 HeLa 细胞或 LATS1 敲除小鼠的成纤维细胞显示出 PLK1 活性增加。我们还发现 DNA 损伤诱导的 LATS1 激活通过 MYPT1 S445 的磷酸化导致 PLK1 抑制。此外，LATS1 敲低细胞在 DNA 损伤后显示出减少的 G2 检查点阻滞。这些结果表明 LATS1 像酵母 Dbf2 一样磷酸化磷酸酶，并证明 LATS1 在控制 G2 DNA 损伤检查点中的 PLK1 方面具有新的作用。
