Characterization of PBIP1, a novel polo-box-interacting protein at the kinetocho

丝氨酸/苏氨酸蛋白激酶的波罗亚家族的成员已被证明在各种真核生物的有丝分裂进程中发挥关键作用。我们以前已经证明，进化上保守的polo激酶的polo盒结构域（PBD）是必不可少的，在特定的亚细胞结构的波罗激酶的催化活性的目标。为了鉴定polo盒结合蛋白，我们使用哺乳动物polo样激酶Plk 1的PBD作为诱饵进行了酵母双杂交筛选，并分离出一种新的kinetochore相关蛋白，我们称之为PBIP 1（Polo盒相互作用蛋白1）。PBIP 1在生理条件下与内源性Plk 1在体内相互作用，并且似乎是一种在有丝分裂中被Plk 1磷酸化和降解的着丝粒特异性有丝分裂抑制剂。缺乏Plk 1依赖性磷酸化位点的不可降解的PBIP 1的表达诱导有丝分裂停滞，表明PBIP 1的降解是适当的有丝分裂进程所必需的。PBIP 1与髓性白血病因子1（MLF 1）结合，MLF 1通过与核磷蛋白（NPM）融合而失调，促进了从骨髓增生异常（MDS）发展为急性髓性白血病（AML）。因此，通过MLF 1下调PBIP 1可能是AML发展的重要步骤。与这些观察结果一致，我们在白血病细胞系的PLK 1 ORF中鉴定了多个突变。这些发现提出了Plk 1失调导致PBIP 1功能异常和白血病发生的可能性，并为波罗激酶在白血病和其他肿瘤发生发展中的作用提供了新的观点。

Members of the polo subfamily of Ser/Thr protein kinases have been shown to play critical roles for mitotic progression in various eukaryotic organisms. We have previously demonstrated that the evolutionarily conserved polo-box domain (PBD) of polo kinases is essential in targeting the catalytic activity of the polo kinases to specific subcellular structures. In an effort to identify the polo-box-binding proteins, we carried out yeast two-hybrid screening using the PBD of the mammalian polo-like kinase Plk1 as a bait and isolated a novel kinetochore-associating protein that we termed PBIP1 (for Polo-Box-Interacting Protein 1). PBIP1 interacts with endogenous Plk1 in vivo under physiological conditions and appears to be a kinetochore-specific mitotic inhibitor that is phosphorylated and degraded by Plk1 in mitosis. Expression of the non-degradable PBIP1 lacking the Plk1-dependent phosphorylation sites induces a mitotic arrest, indicating that degradation of PBIP1 is required for proper mitotic progression. PBIP1 binds to myeloid leukemia factor 1 (MLF1), whose deregulation by fusing with nuclear phosmin (NPM) promotes the development of acute myeloid leukemia (AML) from myelodysplasia (MDS). Thus, down-regulation of PBIP1 by MLF1 could be an important step for the development of AML. In line with these observations, we identified multiple mutations in the PLK1 ORF of leukemia cell lines. These findings raise the possibility that deregulation of Plk1 leads to improper PBIP1 function and the genesis of leukemia, and provide new views on the role of polo kinase in the development of leukemia and other tumoregenesis.